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Ash K, Dev A. Harnessing nanotechnology in HIV therapy: Exploring molecular pathogenesis and treatment strategies with special reference to chemotherapy and immunotherapy. Microb Pathog 2025; 204:107625. [PMID: 40268149 DOI: 10.1016/j.micpath.2025.107625] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2025] [Revised: 04/18/2025] [Accepted: 04/21/2025] [Indexed: 04/25/2025]
Abstract
Human immunodeficiency virus (HIV) continues to be a global threat, contributing substantially to social and economic burdens worldwide. Synthetic ARV drugs are classified into six different classes viz NRTIs, NNRTIs, PIs, IIs, INSTIs, and FIs. Highly active anti-retroviral therapy (HAART) which is a combination of two or more ARV drugs from different classes is gaining immense popularity in the HIV therapy regimen due to its better therapeutic outcome. However, despite its successful endeavor in significant viral suppression, synthetic drugs are associated with numerous adverse effects. To mitigate this issue, scientists are exploring ARV agents derived from various natural sources like plants, and marine organisms that can exhibit potent anti-HIV activity with minimal side effects. Nevertheless, both synthetically and naturally derived ARV agents have failed to exhibit eradication of HIV from latent reservoirs. Henceforth, researchers are shifting their attention towards formulating a drug-encapsulated nano-delivery system to ensure a significant amount of drug delivery into these reservoirs. Additionally, the discovery of a novel HIV vaccine that can induce robust immune responses against multiple HIV strains and facilitate complete removal of the virus before the establishment of a latent reservoir is the need of an hour. Briefly, we discussed various synthetic and natural chemotherapeutic agents along with their specificity and limitations, different drug-delivery devices for ART, immunotherapy, vaccines, and lastly, challenges and strategies associated with vaccine development.
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Affiliation(s)
- Kaushiki Ash
- Department of Pharmaceutical Sciences and Technology, Birla Institute of Technology, Mesra, Jharkhand, India
| | - Abhimanyu Dev
- Department of Pharmaceutical Sciences and Technology, Birla Institute of Technology, Mesra, Jharkhand, India.
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Zhang X, Lim K, Qiu Y, Hazawa M, Wong RW. Strategies for the Viral Exploitation of Nuclear Pore Transport Pathways. Viruses 2025; 17:151. [PMID: 40006906 PMCID: PMC11860923 DOI: 10.3390/v17020151] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2024] [Revised: 01/20/2025] [Accepted: 01/21/2025] [Indexed: 02/27/2025] Open
Abstract
Viruses frequently exploit the host's nucleocytoplasmic trafficking machinery to facilitate their replication and evade immune defenses. By encoding specialized proteins and other components, they strategically target host nuclear transport receptors (NTRs) and nucleoporins within the spiderweb-like inner channel of the nuclear pore complex (NPC), enabling efficient access to the host nucleus. This review explores the intricate mechanisms governing the nuclear import and export of viral components, with a focus on the interplay between viral factors and host determinants that are essential for these processes. Given the pivotal role of nucleocytoplasmic shuttling in the viral life cycle, we also examine therapeutic strategies aimed at disrupting the host's nuclear transport pathways. This includes evaluating the efficacy of pharmacological inhibitors in impairing viral replication and assessing their potential as antiviral treatments. Furthermore, we emphasize the need for continued research to develop targeted therapies that leverage vulnerabilities in nucleocytoplasmic trafficking. Emerging high-resolution techniques, such as advanced imaging and computational modeling, are transforming our understanding of the dynamic interactions between viruses and the NPC. These cutting-edge tools are driving progress in identifying novel therapeutic opportunities and uncovering deeper insights into viral pathogenesis. This review highlights the importance of these advancements in paving the way for innovative antiviral strategies.
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Affiliation(s)
- Xin Zhang
- Division of Nano Life Science, Graduate School of Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan; (X.Z.); (Y.Q.)
| | - Keesiang Lim
- WPI-Nano Life Science Institute, Kanazawa University, Kanazawa 920-1192, Japan;
| | - Yujia Qiu
- Division of Nano Life Science, Graduate School of Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan; (X.Z.); (Y.Q.)
| | - Masaharu Hazawa
- WPI-Nano Life Science Institute, Kanazawa University, Kanazawa 920-1192, Japan;
- Cell-Bionomics Research Unit, Institute for Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan
| | - Richard W. Wong
- Division of Nano Life Science, Graduate School of Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan; (X.Z.); (Y.Q.)
- WPI-Nano Life Science Institute, Kanazawa University, Kanazawa 920-1192, Japan;
- Cell-Bionomics Research Unit, Institute for Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan
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Ding H, Nguyen HT, Li W, Deshpande A, Zhang S, Jiang F, Zhang Z, Anang S, Mothes W, Sodroski J, Kappes JC. Inducible cell lines producing replication-defective human immunodeficiency virus particles containing envelope glycoproteins stabilized in a pretriggered conformation. J Virol 2024; 98:e0172024. [PMID: 39508605 PMCID: PMC11650979 DOI: 10.1128/jvi.01720-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2024] [Accepted: 10/16/2024] [Indexed: 11/15/2024] Open
Abstract
During the process by which human immunodeficiency virus (HIV-1) enters cells, the envelope glycoprotein (Env) trimer on the virion surface engages host cell receptors. Binding to the receptor CD4 induces Env to undergo transitions from a pretriggered, "closed" (State-1) conformation to more "open" (State 2/3) conformations. Most broadly neutralizing antibodies (bNAbs), which are difficult to elicit, recognize the pretriggered (State-1) conformation. More open Env conformations are recognized by poorly neutralizing antibodies (pNAbs), which are readily elicited during natural infection and vaccination with current Env immunogens. Env heterogeneity likely contributes to HIV-1 persistence by skewing antibody responses away from the pretriggered conformation. The conformationally flexible gp160 Env precursor on the infected cell or virion surface potentially presents multiple pNAb epitopes to the host immune system. Although proteolytic cleavage to produce the functional, mature Env trimer [(gp120/gp41)3] stabilizes State-1, many primary HIV-1 Envs spontaneously sample more open conformations. Here, we establish inducible cell lines that produce replication-defective HIV-1 particles with Env trimers stabilized in a pretriggered conformation. The mature Env is enriched on virus-like particles (VLPs). Using complementary approaches, we estimate an average of 25-50 Env trimers on each VLP. The stabilizing changes in Env limit the natural conformational heterogeneity of the VLP Env trimers, allowing recognition by bNAbs but not pNAbs. These defective VLPs provide a more homogeneous source of pretriggered Env trimers in a native membrane environment. Thus, these VLPs may facilitate the characterization of this functionally important Env conformation and its interaction with the immune system.IMPORTANCEA major impediment to the development of an effective HIV/AIDS vaccine is the inefficiency with which human immunodeficiency virus (HIV-1) envelope glycoproteins elicit antibodies that neutralize multiple virus strains. Neutralizing antibodies recognize a particular shape of the envelope glycoproteins that resides on the viral membrane before the virus engages the host cell. Here, we report the creation of stable cell lines that inducibly produce non-infectious HIV-like particles. The normally flexible envelope glycoprotein spikes on these virus-like particles have been stabilized in a conformation that is recognized by broadly neutralizing antibodies. These virus-like particles allow the study of the envelope glycoprotein conformation, its modification by sugars, and its ability to elicit desired neutralizing antibodies.
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Affiliation(s)
- Haitao Ding
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama, USA
| | - Hanh T. Nguyen
- Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
- Department of Microbiology, Harvard Medical School, Boston, Massachusetts, USA
| | - Wenwei Li
- Department of Microbial Pathogenesis, Yale University, New Haven, Connecticut, USA
| | - Ashlesha Deshpande
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama, USA
| | - Shijian Zhang
- Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
- Department of Microbiology, Harvard Medical School, Boston, Massachusetts, USA
| | - Fan Jiang
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama, USA
| | - Zhiqing Zhang
- Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
- Department of Microbiology, Harvard Medical School, Boston, Massachusetts, USA
| | - Saumya Anang
- Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
- Department of Microbiology, Harvard Medical School, Boston, Massachusetts, USA
| | - Walther Mothes
- Department of Microbial Pathogenesis, Yale University, New Haven, Connecticut, USA
| | - Joseph Sodroski
- Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
- Department of Microbiology, Harvard Medical School, Boston, Massachusetts, USA
| | - John C. Kappes
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama, USA
- Birmingham Veterans Affairs Medical Center, Research Service, Birmingham, Alabama, USA
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Ortiz-Hernandez GL, Sanchez-Hernandez ES, Ochoa PT, Casiano CA. The Emerging Roles of the Stress Epigenetic Reader LEDGF/p75 in Cancer Biology and Therapy Resistance: Mechanisms and Targeting Opportunities. Cancers (Basel) 2024; 16:3957. [PMID: 39682146 DOI: 10.3390/cancers16233957] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2024] [Revised: 11/19/2024] [Accepted: 11/21/2024] [Indexed: 12/18/2024] Open
Abstract
The lens epithelium derived growth factor of 75 kD (LEDGF/p75) is a transcription co-activator and epigenetic reader that has emerged as a stress oncoprotein in multiple human cancers. Growing evidence indicates that it promotes tumor cell survival against certain therapeutic drugs. The amino (N)-terminal region of LEDGF/p75 contains a PWWP domain that reads methylated histone marks, critical for recognizing transcriptionally active chromatin sites. Its carboxyl (C)-terminus has an integrase binding domain (IBD) that serves as the binding site for the HIV-1 integrase and multiple oncogenic transcription factors. Acting as hubs for protein-protein interactions, both domains facilitate the tethering of oncogenic transcription factors and regulators to active chromatin to regulate mRNA splicing, promote DNA repair, and enhance the expression of stress and cancer-related genes that contribute to tumor cell aggressiveness and chemoresistance. This review summarizes our current knowledge of the emerging roles of LEDGF/p75 in cancer biology and therapy resistance and discusses its potential as a novel oncotherapeutic target in combinatorial treatments.
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Affiliation(s)
- Greisha L Ortiz-Hernandez
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Evelyn S Sanchez-Hernandez
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Pedro T Ochoa
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Carlos A Casiano
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
- Department of Medicine, Division of Rheumatology, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
- Cancer Center, Loma Linda University Health, Loma Linda, CA 92350, USA
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Padron A, Dwivedi R, Chakraborty R, Prakash P, Kim K, Shi J, Ahn J, Pandhare J, Luban J, Aiken C, Balasubramaniam M, Dash C. Cyclophilin A facilitates HIV-1 integration. J Virol 2024; 98:e0094724. [PMID: 39480090 PMCID: PMC11575316 DOI: 10.1128/jvi.00947-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2024] [Accepted: 09/25/2024] [Indexed: 11/02/2024] Open
Abstract
Cyclophilin A (CypA) binds to the HIV-1 capsid to facilitate reverse transcription and nuclear entry and counter the antiviral activity of TRIM5α. Interestingly, recent studies suggest that the capsid enters the nucleus of an infected cell and uncoats prior to integration. We have previously reported that the capsid protein regulates HIV-1 integration. Therefore, we probed whether CypA-capsid interaction also regulates this post-nuclear entry step. First, we challenged CypA-expressing (CypA+/+) and CypA-depleted (CypA-/-) cells with HIV-1 and quantified the levels of provirus. CypA-depletion significantly reduced integration, an effect that was independent of CypA's effect on reverse transcription, nuclear entry, and the presence or absence of TRIM5α. In addition, cyclosporin A, an inhibitor that disrupts CypA-capsid binding, inhibited proviral integration in CypA+/+ cells but not in CypA-/- cells. HIV-1 capsid mutants (G89V and P90A) deficient in CypA binding were also blocked at the integration step in CypA+/+ cells but not in CypA-/- cells. Then, to understand the mechanism, we assessed the integration activity of the HIV-1 preintegration complexes (PICs) extracted from acutely infected cells. PICs from CypA-/- cells retained lower integration activity in vitro compared to those from CypA+/+ cells. PICs from cells depleted of both CypA and TRIM5α also had lower activity, suggesting that CypA's effect on PIC was independent of TRIM5α. Finally, CypA protein specifically stimulated PIC activity, as this effect was significantly blocked by CsA. Collectively, these results provide strong evidence that CypA directly promotes HIV-1 integration, a previously unknown role of this host factor in the nucleus of an infected cell. IMPORTANCE Interaction between the HIV-1 capsid and host cellular factors is essential for infection. However, the molecular details and functional consequences of viral-host factor interactions during HIV-1 infection are not fully understood. Over 30 years ago, Cyclophilin A (CypA) was identified as the first host protein to bind to the HIV-1 capsid. Now it is established that CypA-capsid interaction promotes reverse transcription and nuclear entry of HIV-1. In addition, CypA blocks TRIM5α-mediated restriction of HIV-1. In this report, we show that CypA promotes the post-nuclear entry step of HIV-1 integration by binding to the viral capsid. Notably, we show that CypA stimulates the viral DNA integration activity of the HIV-1 preintegration complex. Collectively, our studies identify a novel role of CypA during the early steps of HIV-1 infection. This new knowledge is important because recent reports suggest that an operationally intact HIV-1 capsid enters the nucleus of an infected cell.
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Affiliation(s)
- Adrian Padron
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
- School of Graduate Studies, Meharry Medical College, Nashville, Tennessee, USA
| | - Richa Dwivedi
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
| | - Rajasree Chakraborty
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
| | - Prem Prakash
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
| | - Kyusik Kim
- Program in Molecular Medicine University of Massachusetts Medical School, Worcester, Massachusetts, USA
| | - Jiong Shi
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Jinwoo Ahn
- Department of Structural Biology, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Jui Pandhare
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
| | - Jeremy Luban
- Program in Molecular Medicine University of Massachusetts Medical School, Worcester, Massachusetts, USA
| | - Christopher Aiken
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Muthukumar Balasubramaniam
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Pharmacology, and Neuroscience, Meharry Medical College, Nashville, Tennessee, USA
| | - Chandravanu Dash
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
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M.Ravichandran S, M.McFadden W, A.Snyder A, G.Sarafianos S. State of the ART (antiretroviral therapy): Long-acting HIV-1 therapeutics. Glob Health Med 2024; 6:285-294. [PMID: 39483451 PMCID: PMC11514626 DOI: 10.35772/ghm.2024.01049] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2024] [Revised: 08/27/2024] [Accepted: 09/02/2024] [Indexed: 11/03/2024]
Abstract
Human immunodeficiency virus (HIV) impacts millions of individuals worldwide, and well over 2/3 of those living with HIV are accessing antiviral therapies that are successfully repressing viral replication. Most often, HIV treatments and prevention are administered in the form of daily pills as combinations of multiple drugs. An emergent and effective strategy for suppressing viral replication is the application of long-acting antiretroviral therapy (LAART), or antivirals that require less-frequent, non-daily doses. Thus far, the repertoire of LAARTs includes the widely used antiviral classes of non-nucleoside reverse transcriptase inhibitors (NNRTIs) and integrase strand transfer inhibitors (INSTIs) and has recently expanded to include a capsid-targeting antiviral. Possible future additions are nucleoside reverse transcriptase inhibitors (NRTIs) and nucleoside reverse transcriptase translocation inhibitors (NRTTIs). Here, we discuss the different strategies of using long-acting compounds to treat or prevent HIV-1 infection by targeting reverse transcriptase, integrase, and capsid.
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Affiliation(s)
- Shreya M.Ravichandran
- Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA
- Children's Healthcare of Atlanta, Atlanta, GA, USA
| | - William M.McFadden
- Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA
- Children's Healthcare of Atlanta, Atlanta, GA, USA
| | - Alexa A.Snyder
- Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA
- Children's Healthcare of Atlanta, Atlanta, GA, USA
| | - Stefan G.Sarafianos
- Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA
- Children's Healthcare of Atlanta, Atlanta, GA, USA
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Hossain MS, Alom MS, Kader MS, Hossain MA, Halim MA. Structure-Guided Antiviral Peptides Identification Targeting the HIV-1 Integrase. ACS PHYSICAL CHEMISTRY AU 2024; 4:464-475. [PMID: 39346608 PMCID: PMC11428276 DOI: 10.1021/acsphyschemau.4c00006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 01/21/2024] [Revised: 06/25/2024] [Accepted: 06/25/2024] [Indexed: 10/01/2024]
Abstract
HIV-1 integrase (IN), a major protein in the HIV life cycle responsible for integrating viral cDNA into the host DNA, represents a promising drug target. Small peptides have emerged as antiviral therapeutics for HIV because of their facile synthesis, highly selective nature, and fewer side effects. However, selecting the best candidates from a vast pool of peptides is a daunting task. In this study, multistep virtual screening was employed to identify potential peptides from a list of 280 HIV inhibitory peptides. Initially, 80 peptides were selected based on their minimum inhibitory concentrations (MIC). Then, molecular docking was performed to evaluate their binding scores compared to HIP000 and HIP00N which are experimentally validated HIV-1 integrase binding peptides that were used as a positive and negative control, respectively. The top-scoring docked complexes, namely, IN-HIP1113, IN-HIP1140, IN-HIP1142, IN-HIP678, IN-HIP776, and IN-HIP777, were subjected to initial 500 ns molecular dynamics (MD) simulations. Subsequently, HIP776, HIP777, and HIP1142 were selected for an in-depth mechanistic study of peptide interactions, with multiple simulations conducted for each complex spanning one microsecond. Independent simulations of the peptides, along with comparisons to the bound state, were performed to elucidate the conformational dynamics of the peptides. These peptides exhibit strong interactions with specific residues, as revealed by snapshot interaction analysis. Notably, LYS159, LYS156, VAL150, and GLU69 residues are prominently involved in these interactions. Additionally, residue-based binding free energy (BFE) calculations highlight the significance of HIS67, GLN148, GLN146, and SER147 residues within the binding pocket. Furthermore, the structure-activity relationship (SAR) analysis demonstrated that aromatic amino acids and the overall volume of peptides are the two major contributors to the docking scores. The best peptides will be validated experimentally by incorporating SAR properties, aiming to develop them as therapeutic agents and structural models for future peptide-based HIV-1 drug design, addressing the urgent need for effective HIV treatments.
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Affiliation(s)
- Md Shahadat Hossain
- Division of Infectious Diseases and Division of Computer Aided Drug Design, The Red-Green Research Centre, BICCB, 16 Tejkunipara, Tejgaon, Dhaka 1215, Bangladesh
- Department of Pharmacy, Faculty of Life Science, Mawlana Bhashani Science & Technology University, Tangail 1902, Bangladesh
| | - Md Siddik Alom
- Ohio State Biochemistry Program, The Ohio State University, Columbus, Ohio 43210, United States
- Center for RNA Biology, The Ohio State University, Columbus, Ohio 43210, United States
| | | | | | - Mohammad A Halim
- Department of Chemistry and Biochemistry, Kennesaw State University, Kennesaw, Georgia 30144, United States
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Padron A, Dwivedi R, Chakraborty R, Prakash P, Kim K, Shi J, Ahn J, Pandhare J, Luban J, Aiken C, Balasubramaniam M, Dash C. Cyclophilin A Facilitates HIV-1 DNA Integration. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.06.15.599180. [PMID: 38948800 PMCID: PMC11212919 DOI: 10.1101/2024.06.15.599180] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/02/2024]
Abstract
Cyclophilin A (CypA) promotes HIV-1 infection by facilitating reverse transcription, nuclear entry and by countering the antiviral activity of TRIM5α. These multifunctional roles of CypA are driven by its binding to the viral capsid. Interestingly, recent studies suggest that the HIV-1 capsid lattice enters the nucleus of an infected cell and uncoats just before integration. Therefore, we tested whether CypA-capsid interaction regulates post-nuclear entry steps of infection, particularly integration. First, we challenged CypA-expressing (CypA +/+ ) and CypA-depleted (CypA -/- ) cells with HIV-1 particles and quantified the resulting levels of provirus. Surprisingly, CypA-depletion significantly reduced integration, an effect that was independent of CypA's effect on reverse transcription, nuclear entry, and the presence or absence of TRIM5α. Additionally, cyclosporin A, an inhibitor that disrupts CypA-capsid binding, inhibited HIV-1 integration in CypA +/+ cells but not in CypA -/- cells. Accordingly, HIV-1 capsid mutants (G89V and P90A) deficient in CypA binding were also blocked at integration in CypA +/+ cells but not in CypA -/- cells. Then, to understand the mechanism, we assessed the integration activity of HIV-1 preintegration complexes (PICs) extracted from infected cells. The PICs from CypA -/- cells had lower activity in vitro compared to those from CypA +/+ cells. PICs from cells depleted for CypA and TRIM5α also had lower activity, suggesting that CypA's effect on PIC activity is independent of TRIM5α. Finally, addition of CypA protein significantly stimulated the integration activity of PICs extracted from both CypA +/+ and CypA -/- cells. Collectively, these results suggest that CypA promotes HIV-1 integration, a previously unknown role of this host factor. Importance HIV-1 capsid interaction with host cellular factors is essential for establishing a productive infection. However, the molecular details of such virus-host interactions are not fully understood. Cyclophilin A (CypA) is the first host protein identified to specifically bind to the HIV-1 capsid. Now it is established that CypA promotes reverse transcription and nuclear entry steps of HIV-1 infection. In this report, we show that CypA promotes HIV-1 integration by binding to the viral capsid. Specifically, our results demonstrate that CypA promotes HIV-1 integration by stimulating the activity of the viral preintegration complex and identifies a novel role of CypA during HIV-1 infection. This new knowledge is important because recent reports suggest that an operationally intact HIV-1 capsid enters the nucleus of an infected cell.
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Kilroy JM, Leal AA, Henderson AJ. Chronic HIV Transcription, Translation, and Persistent Inflammation. Viruses 2024; 16:751. [PMID: 38793632 PMCID: PMC11125830 DOI: 10.3390/v16050751] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2024] [Revised: 05/02/2024] [Accepted: 05/07/2024] [Indexed: 05/26/2024] Open
Abstract
People with HIV exhibit persistent inflammation that correlates with HIV-associated comorbidities including accelerated aging, increased risk of cardiovascular disease, and neuroinflammation. Mechanisms that perpetuate chronic inflammation in people with HIV undergoing antiretroviral treatments are poorly understood. One hypothesis is that the persistent low-level expression of HIV proviruses, including RNAs generated from defective proviral genomes, drives the immune dysfunction that is responsible for chronic HIV pathogenesis. We explore factors during HIV infection that contribute to the generation of a pool of defective proviruses as well as how HIV-1 mRNA and proteins alter immune function in people living with HIV.
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Affiliation(s)
- Jonathan M. Kilroy
- Department of Virology, Immunology, Microbiology, Boston University Chobanian and Avedisian School of Medicine, Boston, MA 02118, USA; (J.M.K.); (A.A.L.)
| | - Andrew A. Leal
- Department of Virology, Immunology, Microbiology, Boston University Chobanian and Avedisian School of Medicine, Boston, MA 02118, USA; (J.M.K.); (A.A.L.)
| | - Andrew J. Henderson
- Department of Virology, Immunology, Microbiology, Boston University Chobanian and Avedisian School of Medicine, Boston, MA 02118, USA; (J.M.K.); (A.A.L.)
- Department of Medicine and Virology, Immunology, Microbiology, Boston University Chobanian and Avedisian School of Medicine, Boston, MA 02118, USA
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10
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Guo X, Yu D, Liu M, Li H, Chen M, Wang X, Zhai X, Zhang B, Wang Y, Yang C, Wang C, Liu Y, Han J, Wang X, Li J, Jia L, Li L. A unified classification system for HIV-1 5' long terminal repeats. PLoS One 2024; 19:e0301809. [PMID: 38696412 PMCID: PMC11065288 DOI: 10.1371/journal.pone.0301809] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2023] [Accepted: 03/22/2024] [Indexed: 05/04/2024] Open
Abstract
The HIV-1 provirus mainly consists of internal coding region flanked by 1 long terminal repeats (LTRs) at each terminus. The LTRs play important roles in HIV-1 reverse transcription, integration, and transcription. However, despite of the significant study advances of the internal coding regions of HIV-1 by using definite reference classification, there are no systematic and phylogenetic classifications for HIV-1 5' LTRs, which hinders our elaboration on 5' LTR and a better understanding of the viral origin, spread and therapy. Here, by analyzing all available resources of 5' LTR sequences in public databases following 4 recognized principles for the reference classification, 83 representatives and 14 consensus sequences were identified as representatives of 2 groups, 6 subtypes, 6 sub-subtypes, and 9 CRFs. To test the reliability of the supplemented classification system, the constructed references were applied to identify the 5' LTR assignment of the 22 clinical isolates in China. The results revealed that 16 out of 22 tested strains showed a consistent subtype classification with the previous LTR-independent classification system. However, 6 strains, for which recombination events within 5' LTR were demonstrated, unexpectedly showed a different subtype classification, leading a significant change of binding sites for important transcription factors including SP1, p53, and NF-κB. The binding change of these transcriptional factors would probably affect the transcriptional activity of 5' LTR. This study supplemented a unified classification system for HIV-1 5' LTRs, which will facilitate HIV-1 characterization and be helpful for both basic and clinical research fields.
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Affiliation(s)
- Xing Guo
- Department of Microbiology, School of Basic Medicine, Anhui Medical University, Hefei, Anhui, China
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Dan Yu
- Laboratory of Dermatology, Key Laboratory of Major Diseases in Children, Ministry of Education, Beijing Pediatric Research Institute, Beijing Children’s Hospital, National Center for Children’s Health, Capital Medical University, Beijing, China
| | - Mengying Liu
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
- College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, China
| | - Hanping Li
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Mingyue Chen
- National 111 Center for Cellular Regulation and Molecular Pharmaceutics, Key Laboratory of Fermentation Engineering, Hubei University of Technology, Wuhan, Hubei, China
| | - Xinyu Wang
- Laboratory of Dermatology, Key Laboratory of Major Diseases in Children, Ministry of Education, Beijing Pediatric Research Institute, Beijing Children’s Hospital, National Center for Children’s Health, Capital Medical University, Beijing, China
| | - Xiuli Zhai
- Department of Microbiology, School of Basic Medicine, Anhui Medical University, Hefei, Anhui, China
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Bohan Zhang
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Yanglan Wang
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
- College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, China
| | - Caiqing Yang
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Chunlei Wang
- Department of Microbiology, School of Basic Medicine, Anhui Medical University, Hefei, Anhui, China
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Yongjian Liu
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Jingwan Han
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Xiaolin Wang
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Jingyun Li
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Lei Jia
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
| | - Lin Li
- Department of Virology, Beijing Institute of Microbiology and Epidemiology, Beijing, China
- State Key Laboratory of Pathogen and Biosecurity, Beijing, China
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11
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Dwivedi R, Prakash P, Kumbhar BV, Balasubramaniam M, Dash C. HIV-1 capsid and viral DNA integration. mBio 2024; 15:e0021222. [PMID: 38085100 PMCID: PMC10790781 DOI: 10.1128/mbio.00212-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/17/2024] Open
Abstract
IMPORTANCE HIV-1 capsid protein (CA)-independently or by recruiting host factors-mediates several key steps of virus replication in the cytoplasm and nucleus of the target cell. Research in the recent years have established that CA is multifunctional and genetically fragile of all the HIV-1 proteins. Accordingly, CA has emerged as a validated and high priority therapeutic target, and the first CA-targeting antiviral drug was recently approved for treating multi-drug resistant HIV-1 infection. However, development of next generation CA inhibitors depends on a better understanding of CA's known roles, as well as probing of CA's novel roles, in HIV-1 replication. In this timely review, we present an updated overview of the current state of our understanding of CA's multifunctional role in HIV-1 replication-with a special emphasis on CA's newfound post-nuclear roles, highlight the pressing knowledge gaps, and discuss directions for future research.
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Affiliation(s)
- Richa Dwivedi
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
| | - Prem Prakash
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Neuroscience and Pharmacology, Meharry Medical College, Nashville, Tennessee, USA
| | - Bajarang Vasant Kumbhar
- Department of Biological Sciences, Sunandan Divatia School of Science, NMIMS (Deemed to be) University, Mumbai, Maharashtra, India
| | - Muthukumar Balasubramaniam
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Neuroscience and Pharmacology, Meharry Medical College, Nashville, Tennessee, USA
| | - Chandravanu Dash
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Neuroscience and Pharmacology, Meharry Medical College, Nashville, Tennessee, USA
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12
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Anisenko A, Galkin S, Mikhaylov AA, Khrenova MG, Agapkina Y, Korolev S, Garkul L, Shirokova V, Ikonnikova VA, Korlyukov A, Dorovatovskii P, Baranov M, Gottikh M. KuINins as a New Class of HIV-1 Inhibitors That Block Post-Integration DNA Repair. Int J Mol Sci 2023; 24:17354. [PMID: 38139188 PMCID: PMC10744174 DOI: 10.3390/ijms242417354] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2023] [Revised: 12/04/2023] [Accepted: 12/04/2023] [Indexed: 12/24/2023] Open
Abstract
Integration of HIV-1 genomic cDNA results in the formation of single-strand breaks in cellular DNA, which must be repaired for efficient viral replication. Post-integration DNA repair mainly depends on the formation of the HIV-1 integrase complex with the Ku70 protein, which promotes DNA-PK assembly at sites of integration and its activation. Here, we have developed a first-class inhibitor of the integrase-Ku70 complex formation that inhibits HIV-1 replication in cell culture by acting at the stage of post-integration DNA repair. This inhibitor, named s17, does not affect the main cellular function of Ku70, namely its participation in the repair of double-strand DNA breaks through the non-homologous end-joining pathway. Using a molecular dynamics approach, we have constructed a model for the interaction of s17 with Ku70. According to this model, the interaction of two phenyl radicals of s17 with the L76 residue of Ku70 is important for this interaction. The requirement of two phenyl radicals in the structure of s17 for its inhibitory properties was confirmed using a set of s17 derivatives. We propose to stimulate compounds that inhibit post-integration repair by disrupting the integrase binding to Ku70 KuINins.
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Affiliation(s)
- Andrey Anisenko
- Chemistry Department, Lomonosov Moscow State University, 119992 Moscow, Russia; (M.G.K.); (Y.A.); (S.K.)
- Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, 119992 Moscow, Russia; (S.G.); (L.G.)
- Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia
| | - Simon Galkin
- Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, 119992 Moscow, Russia; (S.G.); (L.G.)
| | - Andrey A. Mikhaylov
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, 117997 Moscow, Russia (V.S.); (V.A.I.); (M.B.)
| | - Maria G. Khrenova
- Chemistry Department, Lomonosov Moscow State University, 119992 Moscow, Russia; (M.G.K.); (Y.A.); (S.K.)
- Federal Research Centre of Biotechnology, Russian Academy of Sciences, 119071 Moscow, Russia
| | - Yulia Agapkina
- Chemistry Department, Lomonosov Moscow State University, 119992 Moscow, Russia; (M.G.K.); (Y.A.); (S.K.)
- Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia
| | - Sergey Korolev
- Chemistry Department, Lomonosov Moscow State University, 119992 Moscow, Russia; (M.G.K.); (Y.A.); (S.K.)
- Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia
| | - Lidia Garkul
- Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, 119992 Moscow, Russia; (S.G.); (L.G.)
| | - Vasilissa Shirokova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, 117997 Moscow, Russia (V.S.); (V.A.I.); (M.B.)
- Higher Chemical College, D.I. Mendeleev University of Chemical Technology of Russia, 125047 Moscow, Russia
| | - Viktoria A. Ikonnikova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, 117997 Moscow, Russia (V.S.); (V.A.I.); (M.B.)
- Higher Chemical College, D.I. Mendeleev University of Chemical Technology of Russia, 125047 Moscow, Russia
| | - Alexander Korlyukov
- Nesmeyanov Institute of Organoelement Compounds, 119334 Moscow, Russia;
- Institute of Translational Medicine and Institute of Pharmacy and Medicinal Chemistry, Pirogov Russian National Research Medical University, 117997 Moscow, Russia
| | | | - Mikhail Baranov
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, 117997 Moscow, Russia (V.S.); (V.A.I.); (M.B.)
- Institute of Translational Medicine and Institute of Pharmacy and Medicinal Chemistry, Pirogov Russian National Research Medical University, 117997 Moscow, Russia
| | - Marina Gottikh
- Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, 119992 Moscow, Russia; (S.G.); (L.G.)
- Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia
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13
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Padron A, Prakash P, Pandhare J, Luban J, Aiken C, Balasubramaniam M, Dash C. Emerging role of cyclophilin A in HIV-1 infection: from producer cell to the target cell nucleus. J Virol 2023; 97:e0073223. [PMID: 37843371 PMCID: PMC10688351 DOI: 10.1128/jvi.00732-23] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/17/2023] Open
Abstract
The HIV-1 genome encodes a small number of proteins with structural, enzymatic, regulatory, and accessory functions. These viral proteins interact with a number of host factors to promote the early and late stages of HIV-1 infection. During the early stages of infection, interactions between the viral proteins and host factors enable HIV-1 to enter the target cell, traverse the cytosol, dock at the nuclear pore, gain access to the nucleus, and integrate into the host genome. Similarly, the viral proteins recruit another set of host factors during the late stages of infection to orchestrate HIV-1 transcription, translation, assembly, and release of progeny virions. Among the host factors implicated in HIV-1 infection, Cyclophilin A (CypA) was identified as the first host factor to be packaged within HIV-1 particles. It is now well established that CypA promotes HIV-1 infection by directly binding to the viral capsid. Mechanistic models to pinpoint CypA's role have spanned from an effect in the producer cell to the early steps of infection in the target cell. In this review, we will describe our understanding of the role(s) of CypA in HIV-1 infection, highlight the current knowledge gaps, and discuss the potential role of this host factor in the post-nuclear entry steps of HIV-1 infection.
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Affiliation(s)
- Adrian Padron
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
- School of Graduate Studies, Meharry Medical College, Nashville, Tennessee, USA
| | - Prem Prakash
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Pharmacology and Neuroscience, Meharry Medical College, Nashville, Tennessee, USA
| | - Jui Pandhare
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
- School of Graduate Studies, Meharry Medical College, Nashville, Tennessee, USA
| | - Jeremy Luban
- Department of Biochemistry and Molecular Biotechnology, University of Massachusetts Medical School, Worcester, Massachusetts, USA
| | - Chris Aiken
- Department of Pathology, Microbiology and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA
| | - Muthukumar Balasubramaniam
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Pharmacology and Neuroscience, Meharry Medical College, Nashville, Tennessee, USA
| | - Chandravanu Dash
- The Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, Tennessee, USA
- Department of Microbiology, Immunology, and Physiology, Meharry Medical College, Nashville, Tennessee, USA
- Department of Biochemistry, Cancer Biology, Pharmacology and Neuroscience, Meharry Medical College, Nashville, Tennessee, USA
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14
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Sun L, Zhang T, Xu S, Zhang X, Zhan P, Liu X. Bibliometric analysis and visualization of research trends on HIV-1 capsid inhibitors (2000-2022). Front Pharmacol 2023; 14:1282090. [PMID: 37936907 PMCID: PMC10626487 DOI: 10.3389/fphar.2023.1282090] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2023] [Accepted: 10/16/2023] [Indexed: 11/09/2023] Open
Abstract
Background: Acquired immunodeficiency syndrome (AIDS) has seriously endangered human life and health, the main pathogenic agent is human immunodeficiency virus type 1 (HIV-1). The combination antiretroviral therapy (cART) has shown serious drug resistance and side effects, and the discovery of HIV-1 capsid inhibitors is an effective way to solve the problem. Recent studies have shown significant progress in the research of HIV-1 capsid inhibitors. However, there is still a lack of comprehensive overview of bibliometric analysis in this field. This study aimed to provide the research trends and hotspots of HIV-1 capsid inhibitors. Method: Publications related to HIV-1 capsid inhibitors from 2000 to 2022 were searched on the Web of Science Core Collection (WoSCC) database and screened according to inclusion criteria. VOSviewer was conducted to evaluate the results. Results: 96 publications from 25 countries were finally included, and the number of annual publications related to HIV-1 capsid inhibitors showed an increasing trend. The United States was the most productive country with the most publication number, H-index, and total citation number, as well as the widest international cooperation. The most popular journal in this field was Journal of Virology. Drexel University was the most productive institution, and Simon Cocklin participated in the most publications. Keywords co-occurrence analysis exhibited that studying the molecular mechanism of capsid protein, discovering drug candidates, and improving antiretroviral therapy are the main and hot topics in this field. Conclusion: This is the first bibliometric study in the field of HIV-1 capsid inhibitors, which comprehensively analyzed the research trends and hotspots in this direction. This work is expected to provide the scientific community with new insights to promote the research of HIV-1 capsid inhibitors.
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Affiliation(s)
- Lin Sun
- Department of Pharmacy, Qilu Hospital of Shandong University, Jinan, Shandong, China
- Key Laboratory of Chemical Biology (Ministry of Education), Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan, Shandong, China
| | - Tongchao Zhang
- Clinical Research Center of Shandong University, Clinical Epidemiology Unit, Qilu Hospital of Shandong University, Jinan, Shandong, China
| | - Shujing Xu
- Key Laboratory of Chemical Biology (Ministry of Education), Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan, Shandong, China
| | - Xujie Zhang
- Key Laboratory of Chemical Biology (Ministry of Education), Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan, Shandong, China
| | - Peng Zhan
- Key Laboratory of Chemical Biology (Ministry of Education), Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan, Shandong, China
| | - Xinyong Liu
- Key Laboratory of Chemical Biology (Ministry of Education), Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan, Shandong, China
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15
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Zhang X, Chen J. HIV Reservoir: How to Measure It? Curr HIV/AIDS Rep 2023; 20:29-41. [PMID: 37004676 DOI: 10.1007/s11904-023-00653-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 02/08/2023] [Indexed: 04/04/2023]
Abstract
PURPOSEOF REVIEW In the current quest for a complete cure for HIV/AIDS, the persistence of a long-lived reservoir of cells carrying replication-competent proviruses is the major challenge. Here, we describe the main elements and characteristics of several widely used assays of HIV latent reservoir detection. RECENT FINDINGS To date, researchers have developed several different HIV latent reservoir detection assays. Among them, the in vitro quantitative viral outgrowth assay (QVOA) has been the gold standard for assessing latent HIV-1 viral load. The intact proviral DNA assay (IPDA) based on PCR also demonstrated the predominance of defective viruses. However, these assays all have some drawbacks and may still be inadequate in detecting the presence of ultralow levels of latent virus in many patients who were initially thought to have been cured, but eventually showed viral rebound. An accurate and precise measurement of the HIV reservoir is therefore needed to evaluate curative strategies, aimed to functional cure or sterilizing cure.
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Affiliation(s)
- Xinyu Zhang
- Scientific Research Center, Shanghai Public Health Clinical Center, Fudan University, Shanghai, China
| | - Jun Chen
- Department of Infectious Diseases and Immunology, Shanghai Public Health Clinical Center, Fudan University, Shanghai, China.
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16
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de Villartay JP, Pannier E, Sibiude J, Frange P, Tubiana R, Blanche S. Brief Report: T-Cell Receptor α Repertoire Diversity at Birth After in utero Exposure to HIV Integrase Strand-Transfer Inhibitors. J Acquir Immune Defic Syndr 2023; 92:260-262. [PMID: 36343360 DOI: 10.1097/qai.0000000000003130] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/09/2022] [Accepted: 10/24/2022] [Indexed: 11/09/2022]
Abstract
ABSTRACT Effectiveness of anti-HIV in the prevention of perinatal transmission has been established. Assessing the tolerance of drug exposure during pregnancy is of the utmost importance given the number of children exposed. HIV integrase and the recombinase-activating gene enzyme involved in the establishment of the T-lymphocyte repertoire show structural similarity. The inhibition of recombinase-activating (RAG) gene by anti-integrases is observed in vitro, in a variable way according to the molecules. Here, we show that in utero exposure to raltegravir did not alter the T-lymphocyte repertoire of 12 newborns. These reassuring data merit verification for other anti-integrases. ( ClinicalTrial.org NCT04024150).
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Affiliation(s)
- Jean Pierre de Villartay
- Imagine Institute, Laboratory "Genome Dynamics in the Immune System", INSERM UMR 11635, Paris, France.,Université Paris-Cité, Paris, France; and
| | - Emmanuelle Pannier
- Université Paris-Cité, Paris, France; and.,Assistance Publique-Hôpitaux de Paris (APHP), Paris and Colombes, France
| | - Jeanne Sibiude
- Université Paris-Cité, Paris, France; and.,Assistance Publique-Hôpitaux de Paris (APHP), Paris and Colombes, France
| | - Pierre Frange
- Université Paris-Cité, Paris, France; and.,Assistance Publique-Hôpitaux de Paris (APHP), Paris and Colombes, France
| | - Roland Tubiana
- Assistance Publique-Hôpitaux de Paris (APHP), Paris and Colombes, France
| | - Stéphane Blanche
- Université Paris-Cité, Paris, France; and.,Assistance Publique-Hôpitaux de Paris (APHP), Paris and Colombes, France
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17
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Sanchez-Hernandez ES, Ortiz-Hernandez GL, Ochoa PT, Reeves M, Bizzaro N, Andrade LEC, Mahler M, Casiano CA. The Nuclear Dense Fine Speckled (DFS) Immunofluorescence Pattern: Not All Roads Lead to DFS70/LEDGFp75. Diagnostics (Basel) 2023; 13:diagnostics13020222. [PMID: 36673033 PMCID: PMC9858485 DOI: 10.3390/diagnostics13020222] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2022] [Revised: 12/29/2022] [Accepted: 01/04/2023] [Indexed: 01/11/2023] Open
Abstract
The monospecific dense fine speckled (DFS) immunofluorescence assay (IFA) pattern is considered a potential marker to aid in exclusion of antinuclear antibody (ANA)-associated rheumatic diseases (AARD). This pattern is typically produced by autoantibodies against transcription co-activator DFS70/LEDGFp75, which are frequently found in healthy individuals and patients with miscellaneous inflammatory conditions. In AARD patients, these antibodies usually co-exist with disease-associated ANAs. Previous studies reported the occurrence of monospecific autoantibodies that generate a DFS-like or pseudo-DFS IFA pattern but do not react with DFS70/LEDGFp75. We characterized this pattern using confocal microscopy and immunoblotting. The target antigen associated with this pattern partially co-localized with DFS70/LEDGFp75 and its interacting partners H3K36me2, an active chromatin marker, and MLL, a transcription factor, in HEp-2 cells, suggesting a role in transcription. Immunoblotting did not reveal a common protein band immunoreactive with antibodies producing the pseudo-DFS pattern, suggesting they may recognize diverse proteins or conformational epitopes. Given the subjectivity of the HEp-2 IFA test, the awareness of pseudo-DFS autoantibodies reinforces recommendations for confirmatory testing when reporting patient antibodies producing a putative DFS pattern in a clinical setting. Future studies should focus on defining the potential diagnostic utility of the pseudo-DFS pattern and its associated antigen(s).
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Affiliation(s)
- Evelyn S. Sanchez-Hernandez
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Greisha L. Ortiz-Hernandez
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Pedro T. Ochoa
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Michael Reeves
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
| | - Nicola Bizzaro
- Laboratorio di Patologia Clinica, Ospedale San Antonio, Azienda Sanitaria Universitaria Integrata, 33100 Udine, Italy
| | - Luis E. C. Andrade
- Rheumatology Division, Department of Medicine, Federal University of Sao Paulo, São Paulo 04021-001, Brazil
- Immunology Division, Fleury Medicine and Health Laboratory, São Paulo 04023-062, Brazil
| | | | - Carlos A. Casiano
- Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
- Rheumatology Division, Department of Medicine, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA
- Correspondence: ; Tel.: +909-558-1000 (ext. 42759); Fax: +909-558-0196
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18
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HIV-1 Preintegration Complex Preferentially Integrates the Viral DNA into Nucleosomes Containing Trimethylated Histone 3-Lysine 36 Modification and Flanking Linker DNA. J Virol 2022; 96:e0101122. [PMID: 36094316 PMCID: PMC9517705 DOI: 10.1128/jvi.01011-22] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023] Open
Abstract
HIV-1 DNA is preferentially integrated into chromosomal hot spots by the preintegration complex (PIC). To understand the mechanism, we measured the DNA integration activity of PICs-extracted from infected cells-and intasomes, biochemically assembled PIC substructures using a number of relevant target substrates. We observed that PIC-mediated integration into human chromatin is preferred compared to genomic DNA. Surprisingly, nucleosomes lacking histone modifications were not preferred integration compared to the analogous naked DNA. Nucleosomes containing the trimethylated histone 3 lysine 36 (H3K36me3), an epigenetic mark linked to active transcription, significantly stimulated integration, but the levels remained lower than the naked DNA. Notably, H3K36me3-modified nucleosomes with linker DNA optimally supported integration mediated by the PIC but not by the intasome. Interestingly, optimal intasome-mediated integration required the cellular cofactor LEDGF. Unexpectedly, LEDGF minimally affected PIC-mediated integration into naked DNA but blocked integration into nucleosomes. The block for the PIC-mediated integration was significantly relieved by H3K36me3 modification. Mapping the integration sites in the preferred substrates revealed that specific features of the nucleosome-bound DNA are preferred for integration, whereas integration into naked DNA was random. Finally, biochemical and genetic studies demonstrate that DNA condensation by the H1 protein dramatically reduces integration, providing further evidence that features inherent to the open chromatin are preferred for HIV-1 integration. Collectively, these results identify the optimal target substrate for HIV-1 integration, report a mechanistic link between H3K36me3 and integration preference, and importantly, reveal distinct mechanisms utilized by the PIC for integration compared to the intasomes. IMPORTANCE HIV-1 infection is dependent on integration of the viral DNA into the host chromosomes. The preintegration complex (PIC) containing the viral DNA, the virally encoded integrase (IN) enzyme, and other viral/host factors carries out HIV-1 integration. HIV-1 integration is not dependent on the target DNA sequence, and yet the viral DNA is selectively inserted into specific "hot spots" of human chromosomes. A growing body of literature indicates that structural features of the human chromatin are important for integration targeting. However, the mechanisms that guide the PIC and enable insertion of the PIC-associated viral DNA into specific hot spots of the human chromosomes are not fully understood. In this study, we describe a biochemical mechanism for the preference of the HIV-1 DNA integration into open chromatin. Furthermore, our study defines a direct role for the histone epigenetic mark H3K36me3 in HIV-1 integration preference and identify an optimal substrate for HIV-1 PIC-mediated viral DNA integration.
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19
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Whelan M, Pelchat M. Role of RNA Polymerase II Promoter-Proximal Pausing in Viral Transcription. Viruses 2022; 14:v14092029. [PMID: 36146833 PMCID: PMC9503719 DOI: 10.3390/v14092029] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2022] [Revised: 09/09/2022] [Accepted: 09/11/2022] [Indexed: 11/16/2022] Open
Abstract
The promoter-proximal pause induced by the binding of the DRB sensitivity-inducing factor (DSIF) and the negative elongation factor (NELF) to RNAP II is a key step in the regulation of metazoan gene expression. It helps maintain a permissive chromatin landscape and ensures a quick transcriptional response from stimulus-responsive pathways such as the innate immune response. It is also involved in the biology of several RNA viruses such as the human immunodeficiency virus (HIV), the influenza A virus (IAV) and the hepatitis delta virus (HDV). HIV uses the pause as one of its mechanisms to enter and maintain latency, leading to the creation of viral reservoirs resistant to antiretrovirals. IAV, on the other hand, uses the pause to acquire the capped primers necessary to initiate viral transcription through cap-snatching. Finally, the HDV RNA genome is transcribed directly by RNAP II and requires the small hepatitis delta antigen to displace NELF from the polymerase and overcome the transcriptional block caused by RNAP II promoter-proximal pausing. In this review, we will discuss the RNAP II promoter-proximal pause and the roles it plays in the life cycle of RNA viruses such as HIV, IAV and HDV.
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Troyano-Hernáez P, Reinosa R, Holguín A. Genetic Diversity and Low Therapeutic Impact of Variant-Specific Markers in HIV-1 Pol Proteins. Front Microbiol 2022; 13:866705. [PMID: 35910645 PMCID: PMC9330395 DOI: 10.3389/fmicb.2022.866705] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2022] [Accepted: 06/06/2022] [Indexed: 11/13/2022] Open
Abstract
The emergence and spread of new HIV-1 variants pose a challenge for the effectiveness of antiretrovirals (ARV) targeting Pol proteins. During viral evolution, non-synonymous mutations have fixed along the viral genome, leading to amino acid (aa) changes that can be variant-specific (V-markers). Those V-markers fixed in positions associated with drug resistance mutations (DRM), or R-markers, can impact drug susceptibility and resistance pathways. All available HIV-1 Pol sequences from ARV-naïve subjects were downloaded from the United States Los Alamos HIV Sequence Database, selecting 59,733 protease (PR), 6,437 retrotranscriptase (RT), and 6,059 integrase (IN) complete sequences ascribed to the four HIV-1 groups and group M subtypes and circulating recombinant forms (CRFs). Using a bioinformatics tool developed in our laboratory (EpiMolBio), we inferred the consensus sequences for each Pol protein and HIV-1 variant to analyze the aa conservation in Pol. We analyzed the Wu–Kabat protein variability coefficient (WK) in PR, RT, and IN group M to study the susceptibility of each site to evolutionary replacements. We identified as V-markers the variant-specific aa changes present in >75% of the sequences in variants with >5 available sequences, considering R-markers those V-markers that corresponded to DRM according to the IAS-USA2019 and Stanford-Database 9.0. The mean aa conservation of HIV-1 and group M consensus was 82.60%/93.11% in PR, 88.81%/94.07% in RT, and 90.98%/96.02% in IN. The median group M WK was 10 in PR, 4 in RT, and 5 in IN. The residues involved in binding or catalytic sites showed a variability <0.5%. We identified 106 V-markers: 31 in PR, 28 in RT, and 47 in IN, present in 11, 12, and 13 variants, respectively. Among them, eight (7.5%) were R-markers, present in five variants, being minor DRM with little potential effect on ARV susceptibility. We present a thorough analysis of Pol variability among all HIV-1 variants circulating to date. The relatively high aa conservation observed in Pol proteins across HIV-1 variants highlights their critical role in the viral cycle. However, further studies are needed to understand the V-markers’ impact on the Pol proteins structure, viral cycle, or treatment strategies, and periodic variability surveillance studies are also required to understand PR, RT, and IN evolution.
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21
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Linden N, Jones RB. Potential multi-modal effects of provirus integration on HIV-1 persistence: lessons from other viruses. Trends Immunol 2022; 43:617-629. [PMID: 35817699 PMCID: PMC9429957 DOI: 10.1016/j.it.2022.06.001] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2022] [Revised: 06/10/2022] [Accepted: 06/12/2022] [Indexed: 11/29/2022]
Abstract
Despite antiretroviral therapy (ART), HIV-1 persists as proviruses integrated into the genomic DNA of CD4+ T cells. The mechanisms underlying the persistence and clonal expansion of these cells remain incompletely understood. Cases have been described in which proviral integration can alter host gene expression to drive cellular proliferation. Here, we review observations from other genome-integrating human viruses to propose additional putative modalities by which HIV-1 integration may alter cellular function to favor persistence, such as by altering susceptibility to cytotoxicity in virus-expressing cells. We propose that signals implicating such mechanisms may have been masked thus far by the preponderance of defective and/or nonreactivatable HIV-1 proviruses, but could be revealed by focusing on the integration sites of intact proviruses with expression potential.
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Affiliation(s)
- Noemi Linden
- Division of Infectious Diseases, Department of Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Immunology and Microbial Pathogenesis Graduate Program, Weill Cornell Graduate School of Medical Sciences, New York, NY 10021, USA; Department of Microbiology and Immunology, Weill Cornell Medicine, New York, NY 10021, USA
| | - R Brad Jones
- Division of Infectious Diseases, Department of Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Immunology and Microbial Pathogenesis Graduate Program, Weill Cornell Graduate School of Medical Sciences, New York, NY 10021, USA; Department of Microbiology and Immunology, Weill Cornell Medicine, New York, NY 10021, USA.
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22
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Lau CY, Adan MA, Maldarelli F. Why the HIV Reservoir Never Runs Dry: Clonal Expansion and the Characteristics of HIV-Infected Cells Challenge Strategies to Cure and Control HIV Infection. Viruses 2021; 13:2512. [PMID: 34960781 PMCID: PMC8708047 DOI: 10.3390/v13122512] [Citation(s) in RCA: 24] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2021] [Revised: 11/22/2021] [Accepted: 11/27/2021] [Indexed: 12/13/2022] Open
Abstract
Antiretroviral therapy (ART) effectively reduces cycles of viral replication but does not target proviral populations in cells that persist for prolonged periods and that can undergo clonal expansion. Consequently, chronic human immunodeficiency virus (HIV) infection is sustained during ART by a reservoir of long-lived latently infected cells and their progeny. This proviral landscape undergoes change over time on ART. One of the forces driving change in the landscape is the clonal expansion of infected CD4 T cells, which presents a key obstacle to HIV eradication. Potential mechanisms of clonal expansion include general immune activation, antigenic stimulation, homeostatic proliferation, and provirus-driven clonal expansion, each of which likely contributes in varying, and largely unmeasured, amounts to maintaining the reservoir. The role of clinical events, such as infections or neoplasms, in driving these mechanisms remains uncertain, but characterizing these forces may shed light on approaches to effectively eradicate HIV. A limited number of individuals have been cured of HIV infection in the setting of bone marrow transplant; information from these and other studies may identify the means to eradicate or control the virus without ART. In this review, we describe the mechanisms of HIV-1 persistence and clonal expansion, along with the attempts to modify these factors as part of reservoir reduction and cure strategies.
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Affiliation(s)
- Chuen-Yen Lau
- HIV Dynamics and Replication Program, NCI, NIH, Bethesda, MD 20892, USA; (C.-Y.L.); (M.A.A.)
| | - Matthew A. Adan
- HIV Dynamics and Replication Program, NCI, NIH, Bethesda, MD 20892, USA; (C.-Y.L.); (M.A.A.)
- Vagelos College of Physicians & Surgeons, Columbia University, New York, NY 10032, USA
| | - Frank Maldarelli
- HIV Dynamics and Replication Program, NCI, NIH, Bethesda, MD 20892, USA; (C.-Y.L.); (M.A.A.)
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23
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Coffin JM, Hughes SH. Clonal Expansion of Infected CD4+ T Cells in People Living with HIV. Viruses 2021; 13:v13102078. [PMID: 34696507 PMCID: PMC8537114 DOI: 10.3390/v13102078] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2021] [Revised: 09/28/2021] [Accepted: 10/05/2021] [Indexed: 01/16/2023] Open
Abstract
HIV infection is not curable with current antiretroviral therapy (ART) because a small fraction of CD4+ T cells infected prior to ART initiation persists. Understanding the nature of this latent reservoir and how it is created is essential to development of potentially curative strategies. The discovery that a large fraction of the persistently infected cells in individuals on suppressive ART are members of large clones greatly changed our view of the reservoir and how it arises. Rather than being the products of infection of resting cells, as was once thought, HIV persistence is largely or entirely a consequence of infection of cells that are either expanding or are destined to expand, primarily due to antigen-driven activation. Although most of the clones carry defective proviruses, some carry intact infectious proviruses; these clones comprise the majority of the reservoir. A large majority of both the defective and the intact infectious proviruses in clones of infected cells are transcriptionally silent; however, a small fraction expresses a few copies of unspliced HIV RNA. A much smaller fraction is responsible for production of low levels of infectious virus, which can rekindle infection when ART is stopped. Further understanding of the reservoir will be needed to clarify the mechanism(s) by which provirus expression is controlled in the clones of cells that constitute the reservoir.
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Affiliation(s)
- John M. Coffin
- Department of Molecular Biology and Microbiology, Tufts University, Boston, MA 02111, USA;
| | - Stephen H. Hughes
- HIV Dynamics and Replication Program, National Cancer Institute in Frederick, Frederick, MD 21702, USA
- Correspondence:
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24
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The LEDGF/p75 Integrase Binding Domain Interactome Contributes to the Survival, Clonogenicity, and Tumorsphere Formation of Docetaxel-Resistant Prostate Cancer Cells. Cells 2021; 10:cells10102723. [PMID: 34685704 PMCID: PMC8534522 DOI: 10.3390/cells10102723] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/04/2021] [Revised: 10/05/2021] [Accepted: 10/06/2021] [Indexed: 12/18/2022] Open
Abstract
Patients with prostate cancer (PCa) receiving docetaxel chemotherapy invariably develop chemoresistance. The transcription co-activator lens epithelium-derived growth factor p75 (LEDGF/p75), also known as DFS70 and PSIP1, is upregulated in several human cancers, including PCa and promotes resistance to docetaxel and other drugs. The C-terminal region of LEDGF/p75 contains an integrase binding domain (IBD) that tethers nuclear proteins, including the HIV-1 integrase and transcription factors, to active chromatin to promote viral integration and transcription of cellular survival genes. Here, we investigated the contribution of the LEDGF/p75 IBD interactome to PCa chemoresistance. Quantitative immunoblotting revealed that LEDGF/p75 and its IBD-interacting partners are endogenously upregulated in docetaxel-resistant PCa cell lines compared to docetaxel-sensitive parental cells. Using specific human autoantibodies, we co-immunoprecipitated LEDGF/p75 with its endogenous IBD-interacting partners JPO2, menin, MLL, IWS1, ASK1, and PogZ, as well as transcription factors c-MYC and HRP2, in docetaxel-resistant cells, and confirmed their nuclear co-localization by confocal microscopy. Depletion of LEDGF/p75 and selected interacting partners robustly decreased the survival, clonogenicity, and tumorsphere formation capacity of docetaxel-resistant cells. These results implicate the LEDGF/p75 IBD interactome in PCa chemoresistance and could lead to novel therapeutic strategies targeting this protein complex for the treatment of docetaxel-resistant tumors.
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25
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Xu S, Sun L, Dick A, Zalloum WA, Huang T, Meuser ME, Zhang X, Tao Y, Cherukupalli S, Ding D, Ding X, Gao S, Jiang X, Kang D, De Clercq E, Pannecouque C, Cocklin S, Liu X, Zhan P. Design, synthesis, and mechanistic investigations of phenylalanine derivatives containing a benzothiazole moiety as HIV-1 capsid inhibitors with improved metabolic stability. Eur J Med Chem 2021; 227:113903. [PMID: 34653770 DOI: 10.1016/j.ejmech.2021.113903] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2021] [Revised: 10/03/2021] [Accepted: 10/03/2021] [Indexed: 12/17/2022]
Abstract
Further clinical development of PF74, a lead compound targeting HIV-1 capsid, is impeded by low antiviral activity and inferior metabolic stability. By modifying the benzene (region I) and indole of PF74, we identified two potent compounds (7m and 7u) with significantly improved metabolic stability. Compared to PF74, 7u displayed greater metabolic stability in human liver microsomes (HLMs) with half-life (t1/2) 109-fold that of PF74. Moreover, mechanism of action (MOA) studies demonstrated that 7m and 7u effectively mirrored the MOA of compounds that interact within the PF74 interprotomer pocket, showing direct and robust interactions with recombinant CA, and 7u displaying antiviral effects in both the early and late stages of HIV-1 replication. Furthermore, MD simulation corroborated that 7u was bound to the PF74 binding site, and the results of the online molinspiration software predicted that 7m and 7u had desirable physicochemical properties. Unexpectedly, this series of compounds exhibited better antiviral activity than PF74 against HIV-2, represented by compound 7m whose anti-HIV-2 activity was almost 5 times increased potency over PF74. Therefore, we have rationally redesigned the PF74 chemotype to inhibitors with novel structures and enhanced metabolic stability in this study. We hope that these new compounds can serve as a blueprint for developing a new generation of HIV treatment regimens.
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Affiliation(s)
- Shujing Xu
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Lin Sun
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Alexej Dick
- Department of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, PA, 19102, USA
| | - Waleed A Zalloum
- Department of Pharmacy, Faculty of Health Science, American University of Madaba, P.O Box 2882, Amman, 11821, Jordan
| | - Tianguang Huang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Megan E Meuser
- Department of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, PA, 19102, USA
| | - Xujie Zhang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Yucen Tao
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Srinivasulu Cherukupalli
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Dang Ding
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Xiao Ding
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Shenghua Gao
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Xiangyi Jiang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Dongwei Kang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Erik De Clercq
- Rega Institute for Medical Research, Laboratory of Virology and Chemotherapy, K.U. Leuven, Herestraat 49 Postbus 1043 (09.A097), B-3000, Leuven, Belgium
| | - Christophe Pannecouque
- Rega Institute for Medical Research, Laboratory of Virology and Chemotherapy, K.U. Leuven, Herestraat 49 Postbus 1043 (09.A097), B-3000, Leuven, Belgium.
| | - Simon Cocklin
- Department of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, PA, 19102, USA.
| | - Xinyong Liu
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China.
| | - Peng Zhan
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China.
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26
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Targeting the Integrated Stress Response Kinase GCN2 to Modulate Retroviral Integration. Molecules 2021; 26:molecules26175423. [PMID: 34500856 PMCID: PMC8434491 DOI: 10.3390/molecules26175423] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2021] [Revised: 09/01/2021] [Accepted: 09/04/2021] [Indexed: 11/16/2022] Open
Abstract
Multiple viral targets are now available in the clinic to fight HIV infection. Even if this targeted therapy is highly effective at suppressing viral replication, caregivers are facing growing therapeutic failures in patients due to resistance, with or without treatment-adherence glitches. Accordingly, it is important to better understand how HIV and other retroviruses replicate in order to propose alternative antiviral strategies. Recent studies have shown that multiple cellular factors are implicated during the integration step and, more specifically, that integrase can be regulated through post-translational modifications. We have shown that integrase is phosphorylated by GCN2, a cellular protein kinase of the integrated stress response, leading to a restriction of HIV replication. In addition, we found that this mechanism is conserved among other retroviruses. Accordingly, we developed an in vitro interaction assay, based on the AlphaLISA technology, to monitor the integrase-GCN2 interaction. From an initial library of 133 FDA-approved molecules, we identified nine compounds that either inhibited or stimulated the interaction between GCN2 and HIV integrase. In vitro characterization of these nine hits validated this pilot screen and demonstrated that the GCN2-integrase interaction could be a viable solution for targeting integrase out of its active site.
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27
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Nickoloff-Bybel EA, Festa L, Meucci O, Gaskill PJ. Co-receptor signaling in the pathogenesis of neuroHIV. Retrovirology 2021; 18:24. [PMID: 34429135 PMCID: PMC8385912 DOI: 10.1186/s12977-021-00569-x] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/17/2021] [Accepted: 08/11/2021] [Indexed: 12/13/2022] Open
Abstract
The HIV co-receptors, CCR5 and CXCR4, are necessary for HIV entry into target cells, interacting with the HIV envelope protein, gp120, to initiate several signaling cascades thought to be important to the entry process. Co-receptor signaling may also promote the development of neuroHIV by contributing to both persistent neuroinflammation and indirect neurotoxicity. But despite the critical importance of CXCR4 and CCR5 signaling to HIV pathogenesis, there is only one therapeutic (the CCR5 inhibitor Maraviroc) that targets these receptors. Moreover, our understanding of co-receptor signaling in the specific context of neuroHIV is relatively poor. Research into co-receptor signaling has largely stalled in the past decade, possibly owing to the complexity of the signaling cascades and functions mediated by these receptors. Examining the many signaling pathways triggered by co-receptor activation has been challenging due to the lack of specific molecular tools targeting many of the proteins involved in these pathways and the wide array of model systems used across these experiments. Studies examining the impact of co-receptor signaling on HIV neuropathogenesis often show activation of multiple overlapping pathways by similar stimuli, leading to contradictory data on the effects of co-receptor activation. To address this, we will broadly review HIV infection and neuropathogenesis, examine different co-receptor mediated signaling pathways and functions, then discuss the HIV mediated signaling and the differences between activation induced by HIV and cognate ligands. We will assess the specific effects of co-receptor activation on neuropathogenesis, focusing on neuroinflammation. We will also explore how the use of substances of abuse, which are highly prevalent in people living with HIV, can exacerbate the neuropathogenic effects of co-receptor signaling. Finally, we will discuss the current state of therapeutics targeting co-receptors, highlighting challenges the field has faced and areas in which research into co-receptor signaling would yield the most therapeutic benefit in the context of HIV infection. This discussion will provide a comprehensive overview of what is known and what remains to be explored in regard to co-receptor signaling and HIV infection, and will emphasize the potential value of HIV co-receptors as a target for future therapeutic development. ![]()
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Affiliation(s)
- E A Nickoloff-Bybel
- Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA
| | - L Festa
- Department of Basic and Translational Sciences, School of Dental Medicine, University of Pennsylvania, 240 S. 40th Street, Philadelphia, PA, 19104, USA
| | - O Meucci
- Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA.,Department of Microbiology and Immunology, Drexel University College of Medicine, Philadelphia, PA, 19102, USA
| | - P J Gaskill
- Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA.
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28
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Sajidah ES, Lim K, Wong RW. How SARS-CoV-2 and Other Viruses Build an Invasion Route to Hijack the Host Nucleocytoplasmic Trafficking System. Cells 2021; 10:1424. [PMID: 34200500 PMCID: PMC8230057 DOI: 10.3390/cells10061424] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2021] [Revised: 05/31/2021] [Accepted: 06/03/2021] [Indexed: 12/14/2022] Open
Abstract
The host nucleocytoplasmic trafficking system is often hijacked by viruses to accomplish their replication and to suppress the host immune response. Viruses encode many factors that interact with the host nuclear transport receptors (NTRs) and the nucleoporins of the nuclear pore complex (NPC) to access the host nucleus. In this review, we discuss the viral factors and the host factors involved in the nuclear import and export of viral components. As nucleocytoplasmic shuttling is vital for the replication of many viruses, we also review several drugs that target the host nuclear transport machinery and discuss their feasibility for use in antiviral treatment.
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Affiliation(s)
- Elma Sakinatus Sajidah
- Division of Nano Life Science in the Graduate School of Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan;
| | - Keesiang Lim
- WPI-Nano Life Science Institute, Kanazawa University, Kanazawa 920-1192, Japan
| | - Richard W. Wong
- Division of Nano Life Science in the Graduate School of Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan;
- WPI-Nano Life Science Institute, Kanazawa University, Kanazawa 920-1192, Japan
- Cell-Bionomics Research Unit, Institute for Frontier Science Initiative, Kanazawa University, Kanazawa 920-1192, Japan
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29
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RNA Helicase DDX3: A Double-Edged Sword for Viral Replication and Immune Signaling. Microorganisms 2021; 9:microorganisms9061206. [PMID: 34204859 PMCID: PMC8227550 DOI: 10.3390/microorganisms9061206] [Citation(s) in RCA: 24] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2021] [Revised: 05/20/2021] [Accepted: 05/21/2021] [Indexed: 12/19/2022] Open
Abstract
DDX3 is a cellular ATP-dependent RNA helicase involved in different aspects of RNA metabolism ranging from transcription to translation and therefore, DDX3 participates in the regulation of key cellular processes including cell cycle progression, apoptosis, cancer and the antiviral immune response leading to type-I interferon production. DDX3 has also been described as an essential cellular factor for the replication of different viruses, including important human threats such HIV-1 or HCV, and different small molecules targeting DDX3 activity have been developed. Indeed, increasing evidence suggests that DDX3 can be considered not only a promising but also a viable target for anticancer and antiviral treatments. In this review, we summarize distinct functional aspects of DDX3 focusing on its participation as a double-edged sword in the host immune response and in the replication cycle of different viruses.
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30
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Arhel NJ. [An unprecedented interest for the human immunodeficiency virus capsid]. Med Sci (Paris) 2021; 37:549-552. [PMID: 34003105 DOI: 10.1051/medsci/2021050] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022] Open
Affiliation(s)
- Nathalie Jane Arhel
- Institut de recherche en infectiologie de Montpellier (IRIM), CNRS UMR9004, Université de Montpellier, 1919 route de Mende, 34090 Montpellier, France
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31
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Xue W, Zheng X, Hu X, Zhang Y. Research and Clinical Significance of the Differentially Expressed Genes TP63 and LMO4 in Human Immunodeficiency Virus-Related Penile Squamous Cell Carcinoma. Am J Mens Health 2021; 15:15579883211011380. [PMID: 33906487 PMCID: PMC8108076 DOI: 10.1177/15579883211011380] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
To study the differential gene expression and clinical significance in human immunodeficiency virus-infected individuals (HIVIIs) with penile squamous cell carcinoma. At our hospital from 2019 to 2020, we selected six samples of HIV-related penile squamous cell carcinoma for the experimental group and six samples of non-HIV-related penile squamous cell carcinoma for the control group. Transcriptome sequencing of sample mRNAs was performed by high-throughput sequencing. Differential gene expression analysis, differential Gene Ontology (GO) enrichment analysis and differential Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were carried out, and the reads per kilobase per million reads (RPKM) value was used as a measure of gene expression. A total of 2418 differentially expressed genes were obtained, of which 663 were upregulated and 1755 were downregulated (absolute value of logFC >1 and p value <.05). On the basis of the significance of the GO enrichment analysis, we found that the tumor protein p63 (TP63) gene was significantly upregulated and that the LIM domain only 4 (LMO4) gene was significantly downregulated in the experimental group compared with the control group. KEGG pathway analysis of the differentially expressed genes revealed that DNA replication was the most significant pathway associated with the upregulated genes and cell adhesion molecule (CAM) metabolism was the most significant pathway associated with the downregulated genes. The gene expression profiles of HIV-related penile squamous cell carcinoma and non-HIV-related penile squamous cell carcinoma are significantly different and involve significant GO enrichment and KEGG metabolic pathways, and this is very meaningful for the study of non-AIDS-defining cancers (NADCs). Differential expression of genes may be an important target for the prevention of penile squamous cell carcinoma in HIVIIs.
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Affiliation(s)
- Wenrui Xue
- Beijing Youan Hospital of Capital Medical University, Fengtai District, Beijing China
| | - Xin Zheng
- Beijing Youan Hospital of Capital Medical University, Fengtai District, Beijing China
| | - Xiaopeng Hu
- Beijing Chaoyang Hospital of Capital Medical University, Chaoyang District, Beijing China
| | - Yu Zhang
- Beijing Youan Hospital of Capital Medical University, Fengtai District, Beijing China
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32
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Integration in oncogenes plays only a minor role in determining the in vivo distribution of HIV integration sites before or during suppressive antiretroviral therapy. PLoS Pathog 2021; 17:e1009141. [PMID: 33826675 PMCID: PMC8055010 DOI: 10.1371/journal.ppat.1009141] [Citation(s) in RCA: 42] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2020] [Revised: 04/19/2021] [Accepted: 03/24/2021] [Indexed: 12/11/2022] Open
Abstract
HIV persists during antiretroviral therapy (ART) as integrated proviruses in cells descended from a small fraction of the CD4+ T cells infected prior to the initiation of ART. To better understand what controls HIV persistence and the distribution of integration sites (IS), we compared about 15,000 and 54,000 IS from individuals pre-ART and on ART, respectively, with approximately 395,000 IS from PBMC infected in vitro. The distribution of IS in vivo is quite similar to the distribution in PBMC, but modified by selection against proviruses in expressed genes, by selection for proviruses integrated into one of 7 specific genes, and by clonal expansion. Clones in which a provirus integrated in an oncogene contributed to cell survival comprised only a small fraction of the clones persisting in on ART. Mechanisms that do not involve the provirus, or its location in the host genome, are more important in determining which clones expand and persist. In HIV-infected individuals, a small fraction of the infected cells persist and divide. This reservoir persists during fully suppressive ART and can rekindle the infection if ART is discontinued. Because the number of possible sites of HIV DNA integration is very large, each infected cell, and all of its descendants, can be identified by the site where the provirus is integrated (IS). To understand the selective forces that determine the fates of infected cells in vivo, we compared the distribution of HIV IS in freshly-infected cells to cells from HIV-infected donors sampled both before and during ART. We found that, as previously reported, integration favors highly-expressed genes. However, over time, the fraction of cells with proviruses integrated in highly-expressed genes decreases, implying that they grow less well. There are exceptions to this broad negative selection. When a provirus is integrated in a specific region in one of seven genes, the proviruses affect the expression of the target gene, promoting growth and/or survival of the cell. Although this effect is striking, it is only a minor component of the forces that promote the growth and survival of the population of infected cells during ART.
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Zila V, Margiotta E, Turoňová B, Müller TG, Zimmerli CE, Mattei S, Allegretti M, Börner K, Rada J, Müller B, Lusic M, Kräusslich HG, Beck M. Cone-shaped HIV-1 capsids are transported through intact nuclear pores. Cell 2021; 184:1032-1046.e18. [PMID: 33571428 PMCID: PMC7895898 DOI: 10.1016/j.cell.2021.01.025] [Citation(s) in RCA: 217] [Impact Index Per Article: 54.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2020] [Revised: 11/20/2020] [Accepted: 01/19/2021] [Indexed: 12/14/2022]
Abstract
Human immunodeficiency virus (HIV-1) remains a major health threat. Viral capsid uncoating and nuclear import of the viral genome are critical for productive infection. The size of the HIV-1 capsid is generally believed to exceed the diameter of the nuclear pore complex (NPC), indicating that capsid uncoating has to occur prior to nuclear import. Here, we combined correlative light and electron microscopy with subtomogram averaging to capture the structural status of reverse transcription-competent HIV-1 complexes in infected T cells. We demonstrated that the diameter of the NPC in cellulo is sufficient for the import of apparently intact, cone-shaped capsids. Subsequent to nuclear import, we detected disrupted and empty capsid fragments, indicating that uncoating of the replication complex occurs by breaking the capsid open, and not by disassembly into individual subunits. Our data directly visualize a key step in HIV-1 replication and enhance our mechanistic understanding of the viral life cycle.
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Affiliation(s)
- Vojtech Zila
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany
| | - Erica Margiotta
- European Molecular Biology Laboratory, Structural and Computational Biology Unit, 69117 Heidelberg, Germany; Collaboration for joint PhD degree between EMBL and Heidelberg University, Faculty of Biosciences, Heidelberg, Germany
| | - Beata Turoňová
- European Molecular Biology Laboratory, Structural and Computational Biology Unit, 69117 Heidelberg, Germany
| | - Thorsten G Müller
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany
| | - Christian E Zimmerli
- European Molecular Biology Laboratory, Structural and Computational Biology Unit, 69117 Heidelberg, Germany
| | - Simone Mattei
- European Molecular Biology Laboratory, Structural and Computational Biology Unit, 69117 Heidelberg, Germany; Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zurich, 8092 Zurich, Switzerland; European Molecular Biology Laboratory, Imaging Center, 69117 Heidelberg, Germany
| | - Matteo Allegretti
- European Molecular Biology Laboratory, Structural and Computational Biology Unit, 69117 Heidelberg, Germany
| | - Kathleen Börner
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany; German Center for Infection Research, partner site Heidelberg, 69120 Heidelberg, Germany
| | - Jona Rada
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany
| | - Barbara Müller
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany
| | - Marina Lusic
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany; German Center for Infection Research, partner site Heidelberg, 69120 Heidelberg, Germany
| | - Hans-Georg Kräusslich
- Department of Infectious Diseases, Virology, University of Heidelberg, 69120 Heidelberg, Germany; German Center for Infection Research, partner site Heidelberg, 69120 Heidelberg, Germany.
| | - Martin Beck
- European Molecular Biology Laboratory, Structural and Computational Biology Unit, 69117 Heidelberg, Germany; Max Planck Institute of Biophysics, Department of Molecular Sociology, 60438 Frankfurt, Germany.
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Urak RZ, Soemardy C, Ray R, Li S, Shevchenko G, Scott T, Lim L, Wang X, Morris KV. Conditionally Replicating Vectors Mobilize Chimeric Antigen Receptors against HIV. MOLECULAR THERAPY-METHODS & CLINICAL DEVELOPMENT 2020; 19:285-294. [PMID: 33102620 PMCID: PMC7569266 DOI: 10.1016/j.omtm.2020.09.014] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/24/2020] [Accepted: 09/23/2020] [Indexed: 11/29/2022]
Abstract
Human immunodeficiency virus (HIV) is an attractive target for chimeric antigen receptor (CAR) therapy. CAR T cells have proved remarkably potent in targeted killing of cancer cells, and we surmised that CAR T cells could prove useful in eradicating HIV-infected cells. Toward this goal, we interrogate several neutralizing single-chain variable fragments (scFvs) that target different regions of the HIV envelope glycoprotein, gp120. We find here that CAR T cells with scFv from NIH45-46 antibody demonstrated the highest cytotoxicity. Although NIH45-46 CAR T cells are capable of eliminating antigen-expressing cells, we wanted to address HIV reactivation from ex vivo culture of HIV patient-derived CAR T cells. In order to capitalize on the HIV reactivation, we developed a conditionally replicating lentiviral vector (crLV). The crLV can hijack HIV machinery, forming a chimeric lentivirus (LV) instead of HIV and delivered to uninfected cells. We find that CAR T cells generated with crLVs have similar CAR-mediated functionality as traditional CARs. We also demonstrate crLVs' capability of expanding CAR percentage and protecting CD4 CAR T cell in HIV donors. Collectively, we demonstrate here that the novel crLV NIH45-46 CAR can serve as a strategy to combat HIV, as well as overcome HIV reactivation in CD4+ CAR T cells.
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Affiliation(s)
- Ryan Z Urak
- Center for Gene Therapy, Beckman Research Institute at the City of Hope, Duarte, CA, USA.,Irell & Manella Graduate School of Biological Sciences, City of Hope, Duarte, CA, USA
| | - Citradewi Soemardy
- Center for Gene Therapy, Beckman Research Institute at the City of Hope, Duarte, CA, USA
| | - Roslyn Ray
- City of Hope Center for Gene and Cell Therapy, Duarte, CA, USA
| | - Shirley Li
- City of Hope Center for Gene and Cell Therapy, Duarte, CA, USA
| | - Galina Shevchenko
- Center for Gene Therapy, Beckman Research Institute at the City of Hope, Duarte, CA, USA.,Irell & Manella Graduate School of Biological Sciences, City of Hope, Duarte, CA, USA
| | - Tristan Scott
- Center for Gene Therapy, Beckman Research Institute at the City of Hope, Duarte, CA, USA
| | - Laura Lim
- Department of Hematology and Hematopoietic Cell Transplantation, Duarte, CA, USA
| | - Xiuli Wang
- Department of Hematology and Hematopoietic Cell Transplantation, Duarte, CA, USA
| | - Kevin V Morris
- Center for Gene Therapy, Beckman Research Institute at the City of Hope, Duarte, CA, USA.,Hematological Malignancy and Stem Cell Transplantation Institute, City of Hope, Duarte, CA, USA.,School of Medical Science, Griffith University, Gold Coast Campus, Southport, QLD 4222 Australia
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35
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Wu VH, Nobles CL, Kuri-Cervantes L, McCormick K, Everett JK, Nguyen S, Del Rio Estrada PM, González-Navarro M, Torres-Ruiz F, Ávila-Ríos S, Reyes-Terán G, Bushman FD, Betts MR. Assessment of HIV-1 integration in tissues and subsets across infection stages. JCI Insight 2020; 5:139783. [PMID: 32970634 PMCID: PMC7605534 DOI: 10.1172/jci.insight.139783] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2020] [Accepted: 09/17/2020] [Indexed: 12/19/2022] Open
Abstract
The integration of HIV DNA into the host genome contributes to lifelong infection in most individuals. Few studies have examined integration in lymphoid tissue, where HIV predominantly persists before and after antiretroviral treatment (ART). Of particular interest is whether integration site distributions differ between infection stages with paired blood and tissue comparisons. Here, we profiled HIV integration site distributions in sorted memory, tissue-resident, and/or follicular helper CD4+ T cell subsets from paired blood and lymphoid tissue samples from acute, chronic, and ART-treated individuals. We observed minor differences in the frequency of nonintronic and nondistal intergenic sites, varying with tissue and residency phenotypes during ART. Genomic and epigenetic annotations were generally similar. Clonal expansion of cells marked by identical integration sites was detected, with increased detection in chronic and ART-treated individuals. However, overlap between or within CD4+ T cell subsets or tissue compartments was only observed in 8 unique sites of the 3540 sites studied. Together, these findings suggest that shared integration sites between blood and tissue may, depending on the tissue site, be the exception rather than the rule and indicate that additional studies are necessary to fully understand the heterogeneity of tissue-sequestered HIV reservoirs.
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Affiliation(s)
- Vincent H Wu
- Department of Microbiology and.,Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | | | - Leticia Kuri-Cervantes
- Department of Microbiology and.,Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | | | | | - Son Nguyen
- Department of Microbiology and.,Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Perla M Del Rio Estrada
- Centro de Investigación en Enfermedades Infecciosas, Instituto Nacional de Enfermedades Respiratorias, Mexico City, Mexico
| | - Mauricio González-Navarro
- Centro de Investigación en Enfermedades Infecciosas, Instituto Nacional de Enfermedades Respiratorias, Mexico City, Mexico
| | - Fernanda Torres-Ruiz
- Centro de Investigación en Enfermedades Infecciosas, Instituto Nacional de Enfermedades Respiratorias, Mexico City, Mexico
| | - Santiago Ávila-Ríos
- Centro de Investigación en Enfermedades Infecciosas, Instituto Nacional de Enfermedades Respiratorias, Mexico City, Mexico
| | - Gustavo Reyes-Terán
- Centro de Investigación en Enfermedades Infecciosas, Instituto Nacional de Enfermedades Respiratorias, Mexico City, Mexico
| | | | - Michael R Betts
- Department of Microbiology and.,Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
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36
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Mattio LM, Catinella G, Pinto A, Dallavalle S. Natural and nature-inspired stilbenoids as antiviral agents. Eur J Med Chem 2020; 202:112541. [PMID: 32652408 PMCID: PMC7335248 DOI: 10.1016/j.ejmech.2020.112541] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2020] [Revised: 05/24/2020] [Accepted: 06/04/2020] [Indexed: 12/12/2022]
Abstract
Viruses continue to be a major threat to human health. In the last century, pandemics occurred and resulted in significant mortality and morbidity. Natural products have been largely screened as source of inspiration for new antiviral agents. Within the huge class of plant secondary metabolites, resveratrol-derived stilbenoids present a wide structural diversity and mediate a great number of biological responses relevant for human health. However, whilst the antiviral activity of resveratrol has been extensively studied, little is known about the efficacy of its monomeric and oligomeric derivatives. The purpose of this review is to provide an overview of the achievements in this field, with particular emphasis on the source, chemical structures and the mechanism of action of resveratrol-derived stilbenoids against the most challenging viruses. The collected results highlight the therapeutic versatility of stilbene-containing compounds and provide a prospective insight into their potential development as antiviral agents.
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Affiliation(s)
- Luce M Mattio
- Department of Food, Environmental and Nutritional Sciences, Università Degli Studi di Milano, Via Celoria 2, 20133, Milano, Italy
| | - Giorgia Catinella
- Department of Food, Environmental and Nutritional Sciences, Università Degli Studi di Milano, Via Celoria 2, 20133, Milano, Italy
| | - Andrea Pinto
- Department of Food, Environmental and Nutritional Sciences, Università Degli Studi di Milano, Via Celoria 2, 20133, Milano, Italy
| | - Sabrina Dallavalle
- Department of Food, Environmental and Nutritional Sciences, Università Degli Studi di Milano, Via Celoria 2, 20133, Milano, Italy.
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37
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Xie L, Chen L, Zhong C, Yu T, Ju Z, Wang M, Xiong H, Zeng Y, Wang J, Hu H, Hou W, Feng Y. MxB impedes the NUP358-mediated HIV-1 pre-integration complex nuclear import and viral replication cooperatively with CPSF6. Retrovirology 2020; 17:16. [PMID: 32600399 PMCID: PMC7322711 DOI: 10.1186/s12977-020-00524-2] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/17/2020] [Accepted: 06/16/2020] [Indexed: 12/26/2022] Open
Abstract
BACKGROUND The human myxovirus resistance 2 (Mx2/MxB) protein was originally found to regulate cytoplasmic-nuclear transport but was recently reported to restrict HIV-1 replication by binding to HIV-1 capsid (CA), preventing uncoating, the nuclear import of pre-integration complex (PIC) and viral DNA integration. This work explores the mechanisms of MxB-mediated HIV-1 inhibition. RESULTS We demonstrated that MxB represses NUP358-mediated PIC nuclear import and HIV-1 replication. Moreover, MxB's effects on PIC nuclear import and HIV-1 replication depend critically on cofactor cleavage and polyadenylation specificity factor subunit 6 (CPSF6). MxB binds nucleoporin NUP358, blocks NUP358-CA interaction, thereby impeding the nuclear import of HIV-1 PIC with CPSF6 binding to PIC. More intriguingly, CPSF6's role in nuclear import depends on MxB, being a facilitator of HIV-1 nuclear import on its own, but becoming an inhibitor when MxB is present. CONCLUSIONS Our work establishes that MxB impedes the NUP358-mediated HIV-1 nuclear import and viral replication cooperatively with CPSF6.
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Affiliation(s)
- Linlin Xie
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Lang Chen
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Chaojie Zhong
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Ting Yu
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Zhao Ju
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Meirong Wang
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Hairong Xiong
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China
| | - Yan Zeng
- Department of Zoology, College of Life Sciences, Nanjing Agriculture University, Nanjing, Jiangsu, People's Republic of China
| | - Jianhua Wang
- Key Laboratory of Molecular Virology and Immunology, Institute Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, People's Republic of China
| | - Haitao Hu
- Department of Microbiology and Immunology, Sealy Center for Vaccine Development and Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX, USA
| | - Wei Hou
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China.
| | - Yong Feng
- State Key Laboratory of Virology/Institute of Medical Virology/Hubei Province Key Laboratory of Allergy & Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei, People's Republic of China.
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38
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Ivanov S, Lagunin A, Filimonov D, Tarasova O. Network-Based Analysis of OMICs Data to Understand the HIV-Host Interaction. Front Microbiol 2020; 11:1314. [PMID: 32625189 PMCID: PMC7311653 DOI: 10.3389/fmicb.2020.01314] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2020] [Accepted: 05/25/2020] [Indexed: 12/22/2022] Open
Abstract
The interaction of human immunodeficiency virus with human cells is responsible for all stages of the viral life cycle, from the infection of CD4+ cells to reverse transcription, integration, and the assembly of new viral particles. To date, a large amount of OMICs data as well as information from functional genomics screenings regarding the HIV–host interaction has been accumulated in the literature and in public databases. We processed databases containing HIV–host interactions and found 2910 HIV-1-human protein-protein interactions, mostly related to viral group M subtype B, 137 interactions between human and HIV-1 coding and non-coding RNAs, essential for viral lifecycle and cell defense mechanisms, 232 transcriptomics, 27 proteomics, and 34 epigenomics HIV-related experiments. Numerous studies regarding network-based analysis of corresponding OMICs data have been published in recent years. We overview various types of molecular networks, which can be created using OMICs data, including HIV–human protein–protein interaction networks, co-expression networks, gene regulatory and signaling networks, and approaches for the analysis of their topology and dynamics. The network-based analysis can be used to determine the critical pathways and key proteins involved in the HIV life cycle, cellular and immune responses to infection, viral escape from host defense mechanisms, and mechanisms mediating different susceptibility of humans to infection. The proteins and pathways identified in these studies represent a basis for developing new anti-HIV therapeutic strategies such as new drugs preventing infection of CD4+ cells and viral replication, effective vaccines, “shock and kill” and “block and lock” approaches to cure latent infection.
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Affiliation(s)
- Sergey Ivanov
- Department of Bioinformatics, Institute of Biomedical Chemistry, Moscow, Russia.,Department of Bioinformatics, Pirogov Russian National Research Medical University, Moscow, Russia
| | - Alexey Lagunin
- Department of Bioinformatics, Institute of Biomedical Chemistry, Moscow, Russia.,Department of Bioinformatics, Pirogov Russian National Research Medical University, Moscow, Russia
| | - Dmitry Filimonov
- Department of Bioinformatics, Institute of Biomedical Chemistry, Moscow, Russia
| | - Olga Tarasova
- Department of Bioinformatics, Institute of Biomedical Chemistry, Moscow, Russia
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39
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Dong B, Borjabad A, Kelschenbach J, Chao W, Volsky DJ, Potash MJ. Prevention and treatment of HIV infection and cognitive disease in mice by innate immune responses. Brain Behav Immun Health 2020; 3:100054. [PMID: 32699842 PMCID: PMC7375446 DOI: 10.1016/j.bbih.2020.100054] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2020] [Accepted: 02/18/2020] [Indexed: 12/20/2022] Open
Abstract
HIV associated neurocognitive impairment afflicts roughly half of infected individuals on antiretroviral therapy. This disease currently has no treatment. We have previously shown that type I interferon is induced by and partially controls infection and neuropathogenesis in mice infected by chimeric HIV, EcoHIV. Here we investigate the intentional ligation of the pattern recognition receptor Toll-like receptor 3 (TLR3) by polyinosinic-polycytidylic acid (poly I:C) for its ability to prevent or control infection and associated cognitive disease in EcoHIV infected mice. We tested topical, injection, and intranasal application of poly I:C in mice during primary infection through injection or sexual transmission or in established infection. We measured different forms of HIV DNA and RNA in tissues by real-time PCR and the development of HIV-associated cognitive disease by the radial arm water maze behavioral test. Our results indicate that poly I:C blocks primary EcoHIV infection of mice prior to reverse transcription and reduces established EcoHIV infection. Prevention or control of viral replication by poly I:C prevents or reverses HIV associated cognitive disease in mice. These findings indicate that poly I:C or other innate immune agonists may be useful in control of HIV cognitive disease.
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Affiliation(s)
- Baojun Dong
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Alejandra Borjabad
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Jennifer Kelschenbach
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Wei Chao
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - David J. Volsky
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Mary Jane Potash
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
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40
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Ortiz-Hernandez GL, Sanchez-Hernandez ES, Casiano CA. Twenty years of research on the DFS70/LEDGF autoantibody-autoantigen system: many lessons learned but still many questions. AUTOIMMUNITY HIGHLIGHTS 2020; 11:3. [PMID: 32127038 PMCID: PMC7065333 DOI: 10.1186/s13317-020-0126-4] [Citation(s) in RCA: 32] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/15/2019] [Accepted: 01/14/2020] [Indexed: 12/24/2022]
Abstract
The discovery and initial characterization 20 years ago of antinuclear autoantibodies (ANAs) presenting a dense fine speckled (DFS) nuclear pattern with strong staining of mitotic chromosomes, detected by indirect immunofluorescence assay in HEp-2 cells (HEp-2 IIFA test), has transformed our view on ANAs. Traditionally, ANAs have been considered as reporters of abnormal immunological events associated with the onset and progression of systemic autoimmune rheumatic diseases (SARD), also called ANA-associated rheumatic diseases (AARD), as well as clinical biomarkers for the differential diagnosis of these diseases. However, based on our current knowledge, it is not apparent that autoantibodies presenting the DFS IIF pattern fall into these categories. These antibodies invariably target a chromatin-associated protein designated as dense fine speckled protein of 70 kD (DFS70), also known as lens epithelium-derived growth factor protein of 75 kD (LEDGF/p75) and PC4 and SFRS1 Interacting protein 1 (PSIP1). This multi-functional protein, hereafter referred to as DFS70/LEDGF, plays important roles in the formation of transcription complexes in active chromatin, transcriptional activation of specific genes, regulation of mRNA splicing, DNA repair, and cellular survival against stress. Due to its multiple functions, it has emerged as a key protein contributing to several human pathologies, including acquired immunodeficiency syndrome (AIDS), leukemia, cancer, ocular diseases, and Rett syndrome. Unlike other ANAs, "monospecific" anti-DFS70/LEDGF autoantibodies (only detectable ANA in serum) are not associated with SARD and have been detected in healthy individuals and some patients with non-SARD inflammatory conditions. These observations have led to the hypotheses that these antibodies could be considered as negative biomarkers of SARD and might even play a protective or beneficial role. In spite of 20 years of research on this autoantibody-autoantigen system, its biological and clinical significance still remains enigmatic. Here we review the current state of knowledge of this system, focusing on the lessons learned and posing emerging questions that await further scrutiny as we continue our quest to unravel its significance and potential clinical and therapeutic utility.
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Affiliation(s)
- Greisha L Ortiz-Hernandez
- Center for Health Disparities and Molecular Medicine, Loma Linda University School of Medicine, Loma Linda, CA, 92350, USA.,Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, USA
| | - Evelyn S Sanchez-Hernandez
- Center for Health Disparities and Molecular Medicine, Loma Linda University School of Medicine, Loma Linda, CA, 92350, USA.,Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, USA
| | - Carlos A Casiano
- Center for Health Disparities and Molecular Medicine, Loma Linda University School of Medicine, Loma Linda, CA, 92350, USA. .,Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, USA. .,Department of Medicine/Division of Rheumatology, Loma Linda University School of Medicine, Loma Linda, USA.
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41
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Trivedi J, Mahajan D, Jaffe RJ, Acharya A, Mitra D, Byrareddy SN. Recent Advances in the Development of Integrase Inhibitors for HIV Treatment. Curr HIV/AIDS Rep 2020; 17:63-75. [PMID: 31965427 PMCID: PMC7004278 DOI: 10.1007/s11904-019-00480-3] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023]
Abstract
PURPOSE OF THE REVIEW The complex multistep life cycle of HIV allows it to proliferate within the host and integrate its genome in to the host chromosomal DNA. This provirus can remain dormant for an indefinite period. The process of integration, governed by integrase (IN), is highly conserved across the Retroviridae family. Hence, targeting integration is not only expected to block HIV replication but may also reveal new therapeutic strategies to treat HIV as well as other retrovirus infections. RECENT FINDINGS HIV integrase (IN) has gained attention as the most promising therapeutic target as there are no equivalent homologues of IN that has been discovered in humans. Although current nano-formulated long-acting IN inhibitors have demonstrated the phenomenal ability to block HIV integration and replication with extraordinary half-life, they also have certain limitations. In this review, we have summarized the current literature on clinically established IN inhibitors, their mechanism of action, the advantages and disadvantages associated with their therapeutic application, and finally current HIV cure strategies using these inhibitors.
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Affiliation(s)
- Jay Trivedi
- National Centre for Cell Science, Pune University Campus, Pune, Maharashtra, India
- Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA
| | - Dinesh Mahajan
- Drug Discovery Research Centre, Translational Health Science and Technology Institute, NCR Biotech Science Cluster, 3rd Milestone, Faridabad, Haryana, India
| | - Russell J Jaffe
- Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA
| | - Arpan Acharya
- Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA
| | - Debashis Mitra
- National Centre for Cell Science, Pune University Campus, Pune, Maharashtra, India.
- Centre for DNA Fingerprinting and Diagnostics, Uppal Telangana state, Hyderabad, India.
| | - Siddappa N Byrareddy
- Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA.
- Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical Center, Omaha, NE, USA.
- Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE, USA.
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Abstract
The HIV-1 capsid performs essential functions during early viral replication and is an interesting target for novel antivirals. Thus, understanding molecular and structural details of capsid function will be important for elucidating early HIV-1 (and retroviral in general) replication in relevant target cells and may also aid antiviral development. Here, we show that HIV-1 capsids stay largely intact during transport to the nucleus of infected T cells but appear to uncoat upon entry into the nucleoplasm. These results support the hypothesis that capsids protect the HIV-1 genome from cytoplasmic defense mechanisms and target the genome toward the nucleus. A protective role of the capsid could be a paradigm that also applies to other viruses. Our findings raise the question of how reverse transcription of the HIV-1 genome is accomplished in the context of the capsid structure and whether the process is completed before the capsid is uncoated at the nuclear pore. HIV-1 infects host cells by fusion at the plasma membrane, leading to cytoplasmic entry of the viral capsid encasing the genome and replication machinery. The capsid eventually needs to disassemble, but time and location of uncoating are not fully characterized and may vary depending on the host cell. To study the fate of the capsid by fluorescence and superresolution (STED) microscopy, we established an experimental system that allows discrimination of subviral structures in the cytosol from intact virions at the plasma membrane or in endosomes without genetic modification of the virus. Quantitative microscopy of infected SupT1-R5 cells revealed that the CA signal on cytosolic HIV-1 complexes corresponded to ∼50% of that found in virions at the cell surface, in agreement with dissociation of nonassembled CA molecules from entering capsids after membrane fusion. The relative amount of CA in postfusion complexes remained stable until they reached the nuclear pore complex, while subviral structures in the nucleus of infected cells lacked detectable CA. An HIV-1 variant defective in binding of the host protein cleavage and polyadenylation specificity factor 6 (CPSF6) exhibited accumulation of CA-positive subviral complexes close to the nuclear envelope without loss of infectivity; STED microscopy revealed direct association of these complexes with nuclear pores. These results support previous observations indicating capsid uncoating at the nuclear pore in infected T-cell lines. They suggest that largely intact HIV-1 capsids dock at the nuclear pore in infected SupT1-R5 cells, with CPSF6 being a facilitator of nucleoplasmic entry in this cell type, as has been observed for infected macrophages.
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Transportin-1 binds to the HIV-1 capsid via a nuclear localization signal and triggers uncoating. Nat Microbiol 2019; 4:1840-1850. [PMID: 31611641 DOI: 10.1038/s41564-019-0575-6] [Citation(s) in RCA: 75] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2018] [Accepted: 08/30/2019] [Indexed: 01/05/2023]
Abstract
The initial steps of HIV replication in host cells prime the virus for passage through the nuclear pore and drive the establishment of a productive and irreparable infection1,2. The timely release of the viral genome from the capsid-referred to as uncoating-is emerging as a critical parameter for nuclear import, but the triggers and mechanisms that orchestrate these steps are unknown. Here, we identify β-karyopherin Transportin-1 (TRN-1) as a cellular co-factor of HIV-1 infection, which binds to incoming capsids, triggers their uncoating and promotes viral nuclear import. Depletion of TRN-1, which we characterized by mass spectrometry, significantly reduced the early steps of HIV-1 infection in target cells, including primary CD4+ T cells. TRN-1 bound directly to capsid nanotubes and induced dramatic structural damage, indicating that TRN-1 is necessary and sufficient for uncoating in vitro. Glycine 89 on the capsid protein, which is positioned within a nuclear localization signal in the cyclophilin A-binding loop, is critical for engaging the hydrophobic pocket of TRN-1 at position W730. In addition, TRN-1 promotes the efficient nuclear import of both viral DNA and capsid protein. Our study suggests that TRN-1 mediates the timely release of the HIV-1 genome from the capsid protein shell and efficient viral nuclear import.
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44
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Discovery of Novel Integrase Inhibitors Acting outside the Active Site Through High-Throughput Screening. Molecules 2019; 24:molecules24203675. [PMID: 31614773 PMCID: PMC6832134 DOI: 10.3390/molecules24203675] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2019] [Revised: 10/03/2019] [Accepted: 10/09/2019] [Indexed: 02/07/2023] Open
Abstract
Currently, an increasing number of drugs are becoming available to clinics for the treatment of HIV infection. Even if this targeted therapy is highly effective at suppressing viral replication, caregivers are facing growing therapeutic failures in patients, due to resistance with or without treatment adherence concerns. Accordingly, it is important to continue to discover small molecules that have a novel mechanism of inhibition. In this work, HIV integrase inhibitors were selected by high-throughput screening. Chemical structure comparisons enabled the identification of stilbene disulfonic acids as a potential new chemotype. Biochemical characterization of the lead compound stilbenavir (NSC34931) and a few derivatives was performed. Stilbene disulfonic acid derivatives exhibit low to sub-micromolar antiviral activity, and they inhibit integrase through DNA-binding inhibition. They probably bind to the C-terminal domain of integrase, in the cavity normally occupied by the noncleaved strand of the viral DNA substrate. Because of this original mode of action compared to active site strand transfer inhibitors, they do not exhibit cross-resistance to the three main resistance pathways to integrase inhibitors (G140S-Q148H, N155H, and Y143R). Further structure–activity optimization should enable the development of more active and less toxic derivatives with potential clinical relevance.
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Jiang X, Wu G, Zalloum WA, Meuser ME, Dick A, Sun L, Chen CH, Kang D, Jing L, Jia R, Cocklin S, Lee KH, Liu X, Zhan P. Discovery of novel 1,4-disubstituted 1,2,3-triazole phenylalanine derivatives as HIV-1 capsid inhibitors. RSC Adv 2019; 9:28961-28986. [PMID: 32089839 PMCID: PMC7034945 DOI: 10.1039/c9ra05869a] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
The HIV-1 capsid (CA) protein plays crucial roles in both early and late stages of the viral life cycle, which has intrigued researchers to target it to develop anti-HIV drugs. Accordingly, in this research, we report the design, synthesis and biological evaluation of a series of novel phenylalanine derivatives as HIV-1 CA protein inhibitors using the Cu(I)-catalyzed azide and alkyne 1,3-dipolar cycloaddition (CuAAC) reaction. Among this series of inhibitors, compound II-10c displayed a remarkable anti-HIV activity (EC50 = 2.13 μM, CC50 > 35.49 μM). Furthermore, surface plasmon resonance (SPR) binding assays showed that compounds II-10c and PF-74 (lead compound) have similar affinities to HIV-1 CA monomer. Further investigation showed that the weak permeability and water solubility of representative compounds were probably the important factors that restricted their cell-based activity. Preliminary structure-activity relationships (SARs) were inferred based on the activities of these compounds, and their known structure. The most promising new compound was studied with molecular dynamics simulation (MD) to determine the preferred interactions with the drug target. Finally, the activities of members of this series of inhibitors were deeply inspected to find the potential reasons for their anti-HIV-1 activity from various perspectives. This highlights the important factors required to design compounds with improved potency.
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Affiliation(s)
- Xiangyi Jiang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Gaochan Wu
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Waleed A Zalloum
- Department of Pharmacy, Faculty of Health Science, /Department of pharmacy, American University of Madaba, P.O Box 2882, Amman, 11821, Jordan
| | - Megan E Meuser
- Department of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, USA
| | - Alexej Dick
- Department of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, USA
| | - Lin Sun
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Chin-Ho Chen
- Duke University Medical Center, Box 2926, Surgical Oncology Research Facility, Durham, NC, 27710, USA
| | - Dongwei Kang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Lanlan Jing
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Ruifang Jia
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Simon Cocklin
- Department of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, USA
| | - Kuo-Hsiung Lee
- Natural Products Research Laboratories, Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC, 27599, USA
| | - Xinyong Liu
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
| | - Peng Zhan
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Ji'nan, Shandong, PR China
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Hejnar J, Ruml T. The Current View of Retroviruses as Seen from the Shoulders of a Giant. Viruses 2019; 11:v11090828. [PMID: 31491994 PMCID: PMC6784152 DOI: 10.3390/v11090828] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2019] [Accepted: 09/03/2019] [Indexed: 11/16/2022] Open
Abstract
It has now been more than two years since we said our last goodbye to Jan Svoboda (14 [...].
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Affiliation(s)
- Jiří Hejnar
- Department of Viral and Cellular Genetics, Institute of Molecular Genetics of the Czech Academy of Sciences, Videnska 1083, CZ-14220 Prague, Czech Republic.
| | - Tomáš Ruml
- Department of Biochemistry and Microbiology, University of Chemistry and Technology, CZ-166 28 Prague, Czech Republic.
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47
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Engelman AN. Multifaceted HIV integrase functionalities and therapeutic strategies for their inhibition. J Biol Chem 2019; 294:15137-15157. [PMID: 31467082 DOI: 10.1074/jbc.rev119.006901] [Citation(s) in RCA: 54] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Antiretroviral inhibitors that are used to manage HIV infection/AIDS predominantly target three enzymes required for virus replication: reverse transcriptase, protease, and integrase. Although integrase inhibitors were the last among this group to be approved for treating people living with HIV, they have since risen to the forefront of treatment options. Integrase strand transfer inhibitors (INSTIs) are now recommended components of frontline and drug-switch antiretroviral therapy formulations. Integrase catalyzes two successive magnesium-dependent polynucleotidyl transferase reactions, 3' processing and strand transfer, and INSTIs tightly bind the divalent metal ions and viral DNA end after 3' processing, displacing from the integrase active site the DNA 3'-hydroxyl group that is required for strand transfer activity. Although second-generation INSTIs present higher barriers to the development of viral drug resistance than first-generation compounds, the mechanisms underlying these superior barrier profiles are incompletely understood. A separate class of HIV-1 integrase inhibitors, the allosteric integrase inhibitors (ALLINIs), engage integrase distal from the enzyme active site, namely at the binding site for the cellular cofactor lens epithelium-derived growth factor (LEDGF)/p75 that helps to guide integration into host genes. ALLINIs inhibit HIV-1 replication by inducing integrase hypermultimerization, which precludes integrase binding to genomic RNA and perturbs the morphogenesis of new viral particles. Although not yet approved for human use, ALLINIs provide important probes that can be used to investigate the link between HIV-1 integrase and viral particle morphogenesis. Herein, I review the mechanisms of retroviral integration as well as the promises and challenges of using integrase inhibitors for HIV/AIDS management.
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Affiliation(s)
- Alan N Engelman
- Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts 02215 Department of Medicine, Harvard Medical School, Boston, Massachusetts 02115
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48
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Patterson K, Shavarebi F, Magnan C, Chang I, Qi X, Baldi P, Bilanchone V, Sandmeyer SB. Local features determine Ty3 targeting frequency at RNA polymerase III transcription start sites. Genome Res 2019; 29:1298-1309. [PMID: 31249062 PMCID: PMC6673722 DOI: 10.1101/gr.240861.118] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/10/2018] [Accepted: 06/12/2019] [Indexed: 12/27/2022]
Abstract
Retroelement integration into host genomes affects chromosome structure and function. A goal of a considerable number of investigations is to elucidate features influencing insertion site selection. The Saccharomyces cerevisiae Ty3 retrotransposon inserts proximal to the transcription start sites (TSS) of genes transcribed by RNA polymerase III (RNAP3). In this study, differential patterns of insertion were profiled genome-wide using a random barcode-tagged Ty3. Saturation transposition showed that tRNA genes (tDNAs) are targeted at widely different frequencies even within isoacceptor families. Ectopic expression of Ty3 integrase (IN) showed that it localized to targets independent of other Ty3 proteins and cDNA. IN, RNAP3, and transcription factor Brf1 were enriched at tDNA targets with high frequencies of transposition. To examine potential effects of cis-acting DNA features on transposition, targeting was tested on high-copy plasmids with restricted amounts of 5′ flanking sequence plus tDNA. Relative activity of targets was reconstituted in these constructions. Weighting of genomic insertions according to frequency identified an A/T-rich sequence followed by C as the dominant site of strand transfer. This site lies immediately adjacent to the adenines previously implicated in the RNAP3 TSS motif (CAA). In silico DNA structural analysis upstream of this motif showed that targets with elevated DNA curvature coincide with reduced integration. We propose that integration mediated by the Ty3 intasome complex (IN and cDNA) is subject to inputs from a combination of host factor occupancy and insertion site architecture, and that this results in the wide range of Ty3 targeting frequencies.
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Affiliation(s)
- Kurt Patterson
- Department of Biological Chemistry, School of Medicine, University of California, Irvine, Irvine, California 92697, USA
| | - Farbod Shavarebi
- Department of Biological Chemistry, School of Medicine, University of California, Irvine, Irvine, California 92697, USA
| | - Christophe Magnan
- School of Information and Computer Sciences, University of California, Irvine, Irvine, California 92697, USA
| | - Ivan Chang
- Department of Biological Chemistry, School of Medicine, University of California, Irvine, Irvine, California 92697, USA
| | - Xiaojie Qi
- Department of Biological Chemistry, School of Medicine, University of California, Irvine, Irvine, California 92697, USA
| | - Pierre Baldi
- School of Information and Computer Sciences, University of California, Irvine, Irvine, California 92697, USA
| | - Virginia Bilanchone
- Department of Biological Chemistry, School of Medicine, University of California, Irvine, Irvine, California 92697, USA
| | - Suzanne B Sandmeyer
- Department of Biological Chemistry, School of Medicine, University of California, Irvine, Irvine, California 92697, USA
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49
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Balasubramaniam M, Pandhare J, Dash C. Immune Control of HIV. JOURNAL OF LIFE SCIENCES (WESTLAKE VILLAGE, CALIF.) 2019; 1:4-37. [PMID: 31468033 PMCID: PMC6714987] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Subscribe] [Scholar Register] [Indexed: 06/10/2023]
Abstract
The human immunodeficiency virus (HIV) infection of the immune cells expressing the cluster of differentiation 4 cell surface glycoprotein (CD4+ cells) causes progressive decline of the immune system and leads to the acquired immunodeficiency syndrome (AIDS). The ongoing global HIV/AIDS pandemic has already claimed over 35 million lives. Even after 37 years into the epidemic, neither a cure is available for the 37 million people living with HIV (PLHIV) nor is a vaccine discovered to avert the millions of new HIV infections that continue to occur each year. If left untreated, HIV infection typically progresses to AIDS and, ultimately, causes death in a majority of PLHIV. The recommended combination antiretroviral therapy (cART) suppresses virus replication and viremia, prevents or delays progression to AIDS, reduces transmission rates, and lowers HIV-associated mortality and morbidity. However, because cART does not eliminate HIV, and an enduring pool of infected resting memory CD4+ T cells (latent HIV reservoir) is established early on, any interruption to cART leads to a relapse of viremia and disease progression. Hence, strict adherence to a life-long cART regimen is mandatory for managing HIV infection in PLHIV. The HIV-1-specific cytotoxic T cells expressing the CD8 glycoprotein (CD8+ CTL) limit the virus replication in vivo by recognizing the viral antigens presented by human leukocyte antigen (HLA) class I molecules on the infected cell surface and killing those cells. Nevertheless, CTLs fail to durably control HIV-1 replication and disease progression in the absence of cART. Intriguingly, <1% of cART-naive HIV-infected individuals called elite controllers/HIV controllers (HCs) exhibit the core features that define a HIV-1 "functional cure" outcome in the absence of cART: durable viral suppression to below the limit of detection, long-term non-progression to AIDS, and absence of viral transmission. Robust HIV-1-specific CTL responses and prevalence of protective HLA alleles associated with enduring HIV-1 control have been linked to the HC phenotype. An understanding of the molecular mechanisms underlying the CTL-mediated suppression of HIV-1 replication and disease progression in HCs carrying specific protective HLA alleles may yield promising insights towards advancing the research on HIV cure and prophylactic HIV vaccine.
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Affiliation(s)
- Muthukumar Balasubramaniam
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, TN – 37208. USA
- Department of Biochemistry and Cancer Biology, Meharry Medical College, Nashville, TN – 37208. USA
| | - Jui Pandhare
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, TN – 37208. USA
- School of Graduate Studies and Research, Meharry Medical College, Nashville, TN – 37208. USA
| | - Chandravanu Dash
- Center for AIDS Health Disparities Research, Meharry Medical College, Nashville, TN – 37208. USA
- Department of Biochemistry and Cancer Biology, Meharry Medical College, Nashville, TN – 37208. USA
- School of Graduate Studies and Research, Meharry Medical College, Nashville, TN – 37208. USA
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50
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Bejarano DA, Peng K, Laketa V, Börner K, Jost KL, Lucic B, Glass B, Lusic M, Müller B, Kräusslich HG. HIV-1 nuclear import in macrophages is regulated by CPSF6-capsid interactions at the nuclear pore complex. eLife 2019; 8:41800. [PMID: 30672737 PMCID: PMC6400501 DOI: 10.7554/elife.41800] [Citation(s) in RCA: 139] [Impact Index Per Article: 23.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2018] [Accepted: 01/16/2019] [Indexed: 12/19/2022] Open
Abstract
Nuclear entry of HIV-1 replication complexes through intact nuclear pore complexes is critical for successful infection. The host protein cleavage-and-polyadenylation-specificity-factor-6 (CPSF6) has been implicated in different stages of early HIV-1 replication. Applying quantitative microscopy of HIV-1 reverse-transcription and pre-integration-complexes (RTC/PIC), we show that CPSF6 is strongly recruited to nuclear replication complexes but absent from cytoplasmic RTC/PIC in primary human macrophages. Depletion of CPSF6 or lack of CPSF6 binding led to accumulation of HIV-1 subviral complexes at the nuclear envelope of macrophages and reduced infectivity. Two-color stimulated-emission-depletion microscopy indicated that under these circumstances HIV-1 complexes are retained inside the nuclear pore and undergo CA-multimer dependent CPSF6 clustering adjacent to the nuclear basket. We propose that nuclear entry of HIV-1 subviral complexes in macrophages is mediated by consecutive binding of Nup153 and CPSF6 to the hexameric CA lattice. Viruses are miniscule parasites that hijack the resources of a cell to make more of themselves. For many, this involves getting inside the nucleus, the fortress that protects the cell’s genetic information. To do so, viruses need to first find a way through a double-layered membrane called the nuclear envelope, which only opens up when a cell divides. Yet, the human immunodeficiency virus type 1 (HIV-1) can infect cells that no longer divide, and in which the nucleus’ walls never come down. The virus cores then head for the nuclear pores, heavily guarded holes in the nuclear envelope that allow the cell's own molecules to go in and out of the nucleus. But HIV-1 is too big to fit through, as its genetic information is encased in a capsid, a coat made of a complex assembly of proteins. However, research shows that these capsid proteins can bind to host proteins at the pore or even inside the nucleus. For example, the capsid protein can recognize the pore protein Nup153, or the nuclear protein CPSF6. These interactions could help the virus make its way in, but how these events unfold is still unclear. To explore this, Bejarano, Peng et al. attached fluorescent labels to HIV-1 and watched as it infected non-dividing cells. Rather than completely get rid of their capsid before they crossed the pores, the virus particles hung on to a large part of their lattice. This remaining coat then attached to CPSF6; when this protein was missing or could not bind to capsid proteins, the viral complexes got stuck in the nuclear pores. This suggests that the capsid lattice could first interact with Nup153 inside the pores: then, CPSF6 would take over, knocking Nup153 away and pulling HIV-1 into the nucleus. Armed with this knowledge, virologists and drug developers could try to block HIV-1 from entering the cell’s nucleus; they could also start to dissect how drugs that target the HIV-1 capsid work. Ultimately, HIV-1 may serve as a model to unravel how large objects can pass the nuclear pore, which may help us understand how molecules are constantly trafficked in and out of the nucleus.
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Affiliation(s)
| | - Ke Peng
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany
| | - Vibor Laketa
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany.,German Center for Infection Research, Heidelberg, Germany
| | - Kathleen Börner
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany.,German Center for Infection Research, Heidelberg, Germany
| | - K Laurence Jost
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany.,German Center for Infection Research, Heidelberg, Germany
| | - Bojana Lucic
- German Center for Infection Research, Heidelberg, Germany.,Department of Infectious Diseases, Integrative Virology, University of Heidelberg, Heidelberg, Germany
| | - Bärbel Glass
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany
| | - Marina Lusic
- German Center for Infection Research, Heidelberg, Germany.,Department of Infectious Diseases, Integrative Virology, University of Heidelberg, Heidelberg, Germany
| | - Barbara Müller
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany
| | - Hans-Georg Kräusslich
- Department of Infectious Diseases Virology, University of Heidelberg, Heidelberg, Germany
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