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Khakwani MMAK, Ji XY, Khattak S, Sun YC, Yao K, Zhang L. Targeting colorectal cancer at the level of nuclear pore complex. J Adv Res 2025; 70:423-444. [PMID: 38876192 PMCID: PMC11976419 DOI: 10.1016/j.jare.2024.06.009] [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: 03/13/2024] [Revised: 05/23/2024] [Accepted: 06/07/2024] [Indexed: 06/16/2024] Open
Abstract
BACKGROUND Nuclear pore complexes (NPCs) are the architectures entrenched in nuclear envelop of a cell that regulate the nucleo-cytoplasmic transportation of materials, such as proteins and RNAs for proper functioning of a cell. The appropriate localization of proteins and RNAs within the cell is essential for its normal functionality. For such a complex transportation of materials across the NPC, around 60 proteins are involved comprising nucleoporins, karyopherins and RAN system proteins that play a vital role in NPC's structure formation, cargo translocation across NPC, and cargoes' rapid directed transportation respectively. In various cancers, the structure and function of NPC is often exaggerated, following altered expressions of its nucleoporins and karyopherins, affecting other proteins of associated signaling pathways. Some inhibitors of karyopherins at present, have potential to regulate the altered level/expression of these karyopherin molecules. AIM OF REVIEW This review summarizes the data from 1990 to 2023, mainly focusing on recent studies that illustrate the structure and function of NPC, the relationship and mechanisms of nucleoporins and karyopherins with colorectal cancer, as well as therapeutic values, in order to understand the pathology and underlying basis of colorectal cancer associated with NPC. This is the first review to our knowledge elucidating the detailed updated studies targeting colorectal cancer at NPC. The review also aims to target certain karyopherins, Nups and their possible inhibitors and activators molecules as a therapeutic strategy. KEY SCIENTIFIC CONCEPTS OF REVIEW NPC structure provides understanding, how nucleoporins and karyopherins as key molecules are responsible for appropriate nucleocytoplasmic transportation. Many studies provide evidences, describing the role of disrupted nucleoporins and karyopherins not only in CRC but also in other non-hematological and hematological malignancies. At present, some inhibitors of karyopherins have therapeutic potential for CRC, however development of more potent inhibitors may provide more effective therapeutic strategies for CRC in near future.
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Affiliation(s)
- Muhammad Mahtab Aslam Khan Khakwani
- Department of General Surgery, Huaihe Hospital of Henan University, Henan University, Kaifeng 475004, China; Henan International Joint Laboratory for Nuclear Protein Regulation, School of Basic Medicine, Henan University, Kaifeng, Henan 475004, China
| | - Xin-Ying Ji
- Department of Oncology, Huaxian County Hospital, Huaxian, Henan Province 456400, China; Faculty of Basic Medical Subjects, Shu-Qing Medical College of Zhengzhou, Gong-Ming Rd, Mazhai Town, Erqi District, Zhengzhou, Henan 450064, China
| | - Saadullah Khattak
- Department of General Surgery, Huaihe Hospital of Henan University, Henan University, Kaifeng 475004, China
| | - Ying-Chuan Sun
- Department of Internal Oncology (Section I), Xuchang Municipal Central Hospital, Xuchang, Henan 430000, China
| | - Kunhou Yao
- Department of General Surgery, Huaihe Hospital of Henan University, Henan University, Kaifeng 475004, China.
| | - Lei Zhang
- Department of General Surgery, Huaihe Hospital of Henan University, Henan University, Kaifeng 475004, China; Henan International Joint Laboratory for Nuclear Protein Regulation, School of Basic Medicine, Henan University, Kaifeng, Henan 475004, China.
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Fung HYJ, Mittal SR, Niesman AB, Jiou J, Shakya B, Yoshizawa T, Cansizoglu AE, Rout MP, Chook YM. Phosphate-dependent nuclear export via a non-classical NES class recognized by exportin Msn5. Nat Commun 2025; 16:2580. [PMID: 40089503 PMCID: PMC11910620 DOI: 10.1038/s41467-025-57752-3] [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: 07/24/2024] [Accepted: 02/27/2025] [Indexed: 03/17/2025] Open
Abstract
Gene expression in response to environmental stimuli is dependent on nuclear localization of key signaling components, which can be tightly regulated by phosphorylation. This is exemplified by the phosphate-sensing transcription factor Pho4, which requires phosphorylation for nuclear export by the yeast exportin Msn5. Here, we present a high resolution cryogenic-electron microscopy structure showing the phosphorylated 35-residue nuclear export signal of Pho4, which binds the concave surface of Msn5 through two Pho4 phospho-serines that align with two Msn5 basic patches. These findings characterize a mechanism of phosphate-specific recognition mediated by a non-classical signal distinct from that for Exportin-1. Furthermore, the discovery that unliganded Msn5 is autoinhibited explains the positive cooperativity of Pho4/Ran-binding and proposes a mechanism for Pho4's release in the cytoplasm. These findings advance our understanding of the diversity of signals that drive nuclear export and how cargo phosphorylation is crucial in regulating nuclear transport and controlling cellular signaling pathways.
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Affiliation(s)
- Ho Yee Joyce Fung
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US
- Department of Biophysics, UT Southwestern Medical Center, Dallas, TX, 75039, US
| | - Sanraj R Mittal
- Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, NY, 10021, US
| | - Ashley B Niesman
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US
- Department of Biophysics, UT Southwestern Medical Center, Dallas, TX, 75039, US
| | - Jenny Jiou
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US
- The Walter and Eliza Hall Institute of Medical Research, 1G, Royal Parade, Parkville, Victoria, 302, Australia
| | - Binita Shakya
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US
- Department of Clinical, Diagnostic & Therapeutic Sciences, College of Allied Health Professions, University of Nebraska Medical Center, Omaha, NE, 68198, US
| | - Takuya Yoshizawa
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US
- Research Division, Chugai Pharmaceutical Co., Ltd, Kanagawa, Japan
| | - Ahmet E Cansizoglu
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US
- EMD Serono Research & Development Institute, 45A Middlesex Turnpike, Billerica, MA, 01821, US
| | - Michael P Rout
- Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, NY, 10021, US
| | - Yuh Min Chook
- Department of Pharmacology, UT Southwestern Medical Center, Dallas, TX, 75039, US.
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Fung HYJ, Mittal SR, Niesman AB, Jiou J, Shakya B, Yoshizawa T, Cansizoglu AE, Rout MP, Chook YM. Phosphate-dependent nuclear export via a novel NES class recognized by exportin Msn5. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.08.12.607649. [PMID: 39211127 PMCID: PMC11361136 DOI: 10.1101/2024.08.12.607649] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 09/04/2024]
Abstract
Gene expression in response to environmental stimuli is dependent on nuclear localization of key signaling components, which can be tightly regulated by phosphorylation. This is exemplified by the phosphate-sensing transcription factor Pho4, which requires phosphorylation for nuclear export by the yeast exportin Msn5. Unlike the traditional hydrophobic nuclear export signal (NES) utilized by the Exportin-1/XPO1 system, cryogenic-electron microscopy structures reveal that Pho4 presents a novel, phosphorylated 35-residue NES that interacts with the concave surface of Msn5 through two Pho4 phospho-serines that align with two Msn5 basic patches, unveiling a previously unknown mechanism of phosphate-specific recognition. Furthermore, the discovery that unliganded Msn5 is autoinhibited explains the positive cooperativity of Pho4/Ran-binding and proposes a mechanism for Pho4's release in the cytoplasm. These findings advance our understanding of the diversity of signals that drive nuclear export and how cargo phosphorylation is crucial in regulating nuclear transport and controlling cellular signaling pathways.
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Huang X, Huang Y, Qin L, Xiao Q, Wang Q, Wang J, Wang W, Lu X, Wu Y. Maize DDK1 encoding an Importin-4 β protein is essential for seed development and grain filling by mediating nuclear exporting of eIF1A. THE NEW PHYTOLOGIST 2024; 241:2075-2089. [PMID: 38095260 DOI: 10.1111/nph.19475] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/01/2023] [Accepted: 11/27/2023] [Indexed: 02/09/2024]
Abstract
Nuclear-cytoplasmic trafficking is crucial for protein synthesis in eukaryotic cells due to the spatial separation of transcription and translation by the nuclear envelope. However, the mechanism underlying this process remains largely unknown in plants. In this study, we isolated a maize (Zea mays) mutant designated developmentally delayed kernel 1 (ddk1), which exhibits delayed seed development and slower filling. Ddk1 encodes a plant-specific protein known as Importin-4 β, and its mutation results in reduced 80S monosomes and suppressed protein synthesis. Through our investigations, we found that DDK1 interacts with eIF1A proteins in vivo. However, in vitro experiments revealed that this interaction exhibits low affinity in the absence of RanGTP. Additionally, while the eIF1A protein primarily localizes to the cytoplasm in the wild-type, it remains significantly retained within the nuclei of ddk1 mutants. These observations suggest that DDK1 functions as an exportin and collaborates with RanGTP to facilitate the nuclear export of eIF1A, consequently regulating endosperm development at the translational level. Importantly, both DDK1 and eIF1A are conserved among various plant species, implying the preservation of this regulatory module across diverse plants.
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Affiliation(s)
- Xing Huang
- National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology & Ecology, Chinese Academy of Sciences, Shanghai, 200032, China
- University of the Chinese Academy of Sciences, Beijing, 100049, China
| | - Yongcai Huang
- National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology & Ecology, Chinese Academy of Sciences, Shanghai, 200032, China
| | - Li Qin
- Institute of Molecular Breeding for Maize, Qilu Normal University, Jinan, 250200, China
| | - Qiao Xiao
- National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology & Ecology, Chinese Academy of Sciences, Shanghai, 200032, China
- University of the Chinese Academy of Sciences, Beijing, 100049, China
| | - Qiong Wang
- National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology & Ecology, Chinese Academy of Sciences, Shanghai, 200032, China
| | - Jiechen Wang
- National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology & Ecology, Chinese Academy of Sciences, Shanghai, 200032, China
| | - Wenqin Wang
- College of Life Science, Shanghai Normal University, 100 Guilin Road, Shanghai, 200233, China
| | - Xiaoduo Lu
- Institute of Molecular Breeding for Maize, Qilu Normal University, Jinan, 250200, China
| | - Yongrui Wu
- National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology & Ecology, Chinese Academy of Sciences, Shanghai, 200032, China
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Gobert AP, Hawkins CV, Williams KJ, Snyder LA, Barry DP, Asim M, Allaman MM, McNamara KM, Delgado AG, Wang Y, Zhao S, Rose KL, Piazuelo MB, Wilson KT. Hypusination in intestinal epithelial cells protects mice from infectious colitis. Gut Microbes 2024; 16:2438828. [PMID: 39673545 DOI: 10.1080/19490976.2024.2438828] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/03/2024] [Revised: 11/26/2024] [Accepted: 12/02/2024] [Indexed: 12/16/2024] Open
Abstract
Enteropathogenic Escherichia coli (EPEC) is a bacterium that causes attaching/effacing (A/E) lesions and serious diarrheal disease, a major health issue in developing countries. EPEC pathogenicity results from the effect of virulence factors and dysregulation of host responses. Polyamines, including spermidine, play a major role in intestinal homeostasis. Spermidine is the substrate for deoxyhypusine synthase (DHPS), which catalyzes the conjugation of the amino acid hypusine to eukaryotic translation initiation factor 5A (EIF5A); hypusinated EIF5A (EIF5AHyp) binds specific mRNAs and initiates translation. Our aim was to determine the role of hypusination during infection with A/E pathogens. We found that DHPS and EIF5AHyp levels are induced in i) a colonic epithelial cell line and human-derived colon organoids infected with EPEC, and ii) the colon of mice infected with Citrobacter rodentium, the rodent equivalent of EPEC. Specific deletion of Dhps in intestinal epithelial cells worsened clinical, histological, and pro-inflammatory parameters in C. rodentium-infected mice. These animals also exhibited an exacerbated pathogenic transcriptome in their colon. Furthermore, infected mice with specific Dhps deletion exhibited reduced levels of proteins involved in detoxification of tissue-damaging reactive aldehydes and consequently increased electrophile adducts in the colon. Thus, hypusination in intestinal epithelial cells protects from infectious colitis mediated by A/E pathogens.
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Affiliation(s)
- Alain P Gobert
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, TN, USA
- Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Caroline V Hawkins
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Kamery J Williams
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Lydia A Snyder
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Daniel P Barry
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Mohammad Asim
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Margaret M Allaman
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Kara M McNamara
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
- Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Alberto G Delgado
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Yu Wang
- Department of Biostatistics, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Shilin Zhao
- Department of Biostatistics, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Kristie L Rose
- Department of Biochemistry, Mass Spectrometry Research Center, Vanderbilt University School of Medicine, Nashville, TN, USA
| | - M Blanca Piazuelo
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Keith T Wilson
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, TN, USA
- Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, TN, USA
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA
- Veterans Affairs Tennessee Valley Healthcare System, Nashville, TN, USA
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6
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Gonzalez CE, Ben Abdeljelil N, Pearson A. The Disruption of a Nuclear Export Signal in the C-Terminus of the Herpes Simplex Virus 1 Determinant of Pathogenicity UL24 Protein Leads to a Syncytial Plaque Phenotype. Viruses 2023; 15:1971. [PMID: 37766377 PMCID: PMC10535440 DOI: 10.3390/v15091971] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2023] [Revised: 09/17/2023] [Accepted: 09/19/2023] [Indexed: 09/29/2023] Open
Abstract
UL24 of herpes simplex virus 1 (HSV-1) has been shown to be a determinant of pathogenesis in mouse models of infection. The N-terminus of UL24 localizes to the nucleus and drives the redistribution of nucleolin and B23. In contrast, when expressed alone, the C-terminal domain of UL24 accumulates in the Golgi apparatus; its importance during infection is unknown. We generated a series of mammalian expression vectors encoding UL24 with nested deletions in the C-terminal domain. Interestingly, enhanced nuclear staining was observed for several UL24-deleted forms in transient transfection assays. The substitution of a threonine phosphorylation site had no effect on UL24 localization or viral titers in cell culture. In contrast, mutations targeting a predicted nuclear export signal (NES) significantly enhanced nuclear localization, indicating that UL24 is able to shuttle between the nucleus and the cytoplasm. Recombinant viruses that encode UL24-harboring substitutions in the NES led to the accumulation of UL24 in the nucleus. Treatment with the CRM-1-specific inhibitor leptomycin B blocked the nuclear export of UL24 in transfected cells but not in the context of infection. Viruses encoding UL24 with NES mutations resulted in a syncytial phenotype, but viral yield was unaffected. These results are consistent with a role for HSV-1 UL24 in late cytoplasmic events in HSV-1 replication.
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Affiliation(s)
| | | | - Angela Pearson
- Centre Armand-Frappier Santé Biotechnologie, Institut National de la Recherche Scientifique, Laval, QC H7V 1B7, Canada
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7
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Gobert AP, Smith TM, Latour YL, Asim M, Barry DP, Allaman MM, Williams KJ, McNamara KM, Delgado AG, Short SP, Mirmira RG, Rose KL, Schey KL, Zagol-Ikapitte I, Coleman JS, Boutaud O, Zhao S, Piazuelo MB, Washington MK, Coburn LA, Wilson KT. Hypusination Maintains Intestinal Homeostasis and Prevents Colitis and Carcinogenesis by Enhancing Aldehyde Detoxification. Gastroenterology 2023; 165:656-669.e8. [PMID: 37271289 PMCID: PMC10527201 DOI: 10.1053/j.gastro.2023.05.041] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/29/2022] [Revised: 04/19/2023] [Accepted: 05/25/2023] [Indexed: 06/06/2023]
Abstract
BACKGROUND & AIMS The amino acid hypusine, synthesized from the polyamine spermidine by the enzyme deoxyhypusine synthase (DHPS), is essential for the activity of eukaryotic translation initiation factor 5A (EIF5A). The role of hypusinated EIF5A (EIF5AHyp) remains unknown in intestinal homeostasis. Our aim was to investigate EIF5AHyp in the gut epithelium in inflammation and carcinogenesis. METHODS We used human colon tissue messenger RNA samples and publicly available transcriptomic datasets, tissue microarrays, and patient-derived colon organoids. Mice with intestinal epithelial-specific deletion of Dhps were investigated at baseline and in models of colitis and colon carcinogenesis. RESULTS We found that patients with ulcerative colitis and Crohn's disease exhibit reduced colon levels of DHPS messenger RNA and DHPS protein and reduced levels of EIF5AHyp. Similarly, colonic organoids from colitis patients also show down-regulated DHPS expression. Mice with intestinal epithelial-specific deletion of Dhps develop spontaneous colon hyperplasia, epithelial proliferation, crypt distortion, and inflammation. Furthermore, these mice are highly susceptible to experimental colitis and show exacerbated colon tumorigenesis when treated with a carcinogen. Transcriptomic and proteomic analysis on colonic epithelial cells demonstrated that loss of hypusination induces multiple pathways related to cancer and immune response. Moreover, we found that hypusination enhances translation of numerous enzymes involved in aldehyde detoxification, including glutathione S-transferases and aldehyde dehydrogenases. Accordingly, hypusination-deficient mice exhibit increased levels of aldehyde adducts in the colon, and their treatment with a scavenger of electrophiles reduces colitis. CONCLUSIONS Hypusination in intestinal epithelial cells has a key role in the prevention of colitis and colorectal cancer, and enhancement of this pathway via supplementation of spermidine could have a therapeutic impact.
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Affiliation(s)
- Alain P Gobert
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee; Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee.
| | - Thaddeus M Smith
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Yvonne L Latour
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Mohammad Asim
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Daniel P Barry
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Margaret M Allaman
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Kamery J Williams
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Kara M McNamara
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Alberto G Delgado
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Sarah P Short
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Raghavendra G Mirmira
- Kovler Diabetes Center, Department of Medicine, The University of Chicago, Chicago, Illinois
| | - Kristie L Rose
- Department of Biochemistry, Mass Spectrometry Research Center, Vanderbilt University School of Medicine, Nashville, Tennessee
| | - Kevin L Schey
- Department of Biochemistry, Mass Spectrometry Research Center, Vanderbilt University School of Medicine, Nashville, Tennessee
| | - Irene Zagol-Ikapitte
- Warren Center for Neuroscience Drug Discovery, Vanderbilt University, Nashville, Tennessee; Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee
| | - Jeremy S Coleman
- Warren Center for Neuroscience Drug Discovery, Vanderbilt University, Nashville, Tennessee; Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee
| | - Olivier Boutaud
- Warren Center for Neuroscience Drug Discovery, Vanderbilt University, Nashville, Tennessee; Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee
| | - Shilin Zhao
- Department of Biostatistics, Vanderbilt University Medical Center, Nashville, Tennessee
| | - M Blanca Piazuelo
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee
| | - M Kay Washington
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee; Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee
| | - Lori A Coburn
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee; Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee; Veterans Affairs Tennessee Valley Healthcare System, Nashville, Tennessee
| | - Keith T Wilson
- Division of Gastroenterology, Hepatology, and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee; Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee; Program in Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee; Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee; Veterans Affairs Tennessee Valley Healthcare System, Nashville, Tennessee.
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Wu YY, Wu GQ, Cai NL, Xu YM, Lau ATY. Comparison of Human Eukaryotic Translation Initiation Factors 5A1 and 5AL1: Identification of Amino Acid Residues Important for EIF5A1 Lysine 50 Hypusination and Its Protein Stability. Int J Mol Sci 2023; 24:ijms24076067. [PMID: 37047039 PMCID: PMC10093921 DOI: 10.3390/ijms24076067] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2022] [Revised: 03/07/2023] [Accepted: 03/09/2023] [Indexed: 04/14/2023] Open
Abstract
The human eukaryotic translation initiation factor 5A (EIF5A) family consists of three members, namely EIF5A1, EIF5A2, and EIF5AL1. Recent studies have shown that the expression of EIF5As is related to many human diseases, such as diabetes, viral infection, central nervous system injury, and cancer. Among them, EIF5A1 plays different functions in various cancers, possibly as a tumor-suppressor or oncogene, while EIF5A2 promotes the occurrence and development of cancer. Yet, the biological function of EIF5AL1 is not being studied so far. Interestingly, although there are only three amino acid (at residues 36, 45, and 109) differences between EIF5A1 and EIF5AL1, we demonstrate that only EIF5A1 can be hypusinated while EIF5AL1 cannot, and EIF5AL1 has a tumor-suppressor-like function by inhibiting cell proliferation and migration. We also show that EIF5AL1 protein turnover is mediated through the proteasomal pathway, and EIF5AL1 protein turnover is much faster than that of EIF5A1, which may explain their differential protein expression level in cells. By engineering single and double mutations on these three amino acids, we pinpoint which of these amino acids are critical for hypusination and protein stability. The data of this work should fill in the gaps in EIF5As research and pave the way for future studies on EIF5AL1.
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Affiliation(s)
- Yu-Yao Wu
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou 515041, China
| | - Gao-Qi Wu
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou 515041, China
| | - Na-Li Cai
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou 515041, China
| | - Yan-Ming Xu
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou 515041, China
| | - Andy T Y Lau
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou 515041, China
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9
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Chen L, Wang Y, Lin J, Song Z, Wang Q, Zhao W, Wang Y, Xiu X, Deng Y, Li X, Li Q, Wang X, Li J, Liu X, Liu K, Zhou J, Li K, Liu Y, Liao S, Deng Q, Xu C, Sun Q, Wu S, Zhang K, Guan MX, Zhou T, Sun F, Cai X, Huang C, Shan G. Exportin 4 depletion leads to nuclear accumulation of a subset of circular RNAs. Nat Commun 2022; 13:5769. [PMID: 36182935 PMCID: PMC9526749 DOI: 10.1038/s41467-022-33356-z] [Citation(s) in RCA: 42] [Impact Index Per Article: 14.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2022] [Accepted: 09/09/2022] [Indexed: 12/19/2022] Open
Abstract
Numerous RNAs are exported from the nucleus, abnormalities of which lead to cellular complications and diseases. How thousands of circular RNAs (circRNAs) are exported from the nucleus remains elusive. Here, we provide lines of evidence to demonstrate a link between the conserved Exportin 4 (XPO4) and nuclear export of a subset of circRNAs in metazoans. Exonic circRNAs (ecircRNAs) with higher expression levels, larger length, and lower GC content are more sensitive to XPO4 deficiency. Cellular insufficiency of XPO4 leads to nuclear circRNA accumulation, circRNA:DNA (ciR-loop) formation, linear RNA:DNA (liR-loop) buildup, and DNA damage. DDX39 known to modulate circRNA export can resolve ciR-loop, and splicing factors involved in the biogenesis of circRNAs can also affect the levels of ciR-loop. Testis and brain are two organs with high abundance of circRNAs, and insufficient XPO4 levels are detrimental, as Xpo4 heterozygous mice display male infertility and neural phenotypes. Increased levels of ciR-loop, R-loop, and DNA damage along with decreased cell numbers are observed in testis and hippocampus of Xpo4 heterozygotes. This study sheds light on the understandings of mechanism of circRNA export and reveals the significance of efficient nuclear export of circRNAs in cellular physiology. This study identifies the evolutionarily conserved Exportin 4 as an essential regulator in the nuclear export of circRNAs. Defective circRNA export results in R-loop formation and DNA damage in cells, as well as testis and neurological defects in mice.
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Affiliation(s)
- Liang Chen
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Yucong Wang
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Jiamei Lin
- School of Life Sciences, Chongqing University, Chongqing, 401331, China.,Center of Plant Functional Genomics, Institute of Advanced Interdisciplinary Studies, Chongqing University, Chongqing, 401331, China
| | - Zhenxing Song
- School of Life Sciences, Chongqing University, Chongqing, 401331, China.,Center of Plant Functional Genomics, Institute of Advanced Interdisciplinary Studies, Chongqing University, Chongqing, 401331, China
| | - Qinwei Wang
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Wenfang Zhao
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Yan Wang
- School of Optometry and Ophthalmology and the Eye Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, 325003, China.,State Key Laboratory of Optometry, Ophthalmology, and Visual Science, 270 Xueyuan Road, Wenzhou, Zhejiang, 325003, China
| | - Xiaoyu Xiu
- School of Optometry and Ophthalmology and the Eye Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, 325003, China.,State Key Laboratory of Optometry, Ophthalmology, and Visual Science, 270 Xueyuan Road, Wenzhou, Zhejiang, 325003, China
| | - Yuqi Deng
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Xiuzhi Li
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Qiqi Li
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Xiaolin Wang
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Jingxin Li
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Xu Liu
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Kunpeng Liu
- Center for Plant Biology, School of Life Sciences, Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China
| | - Jincong Zhou
- Center for Plant Biology, School of Life Sciences, Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China
| | - Kuan Li
- Center for Plant Biology, School of Life Sciences, Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China
| | - Yuchan Liu
- MOE Key Laboratory for Membraneless Organelles and Cellular Dynamics, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Shanhui Liao
- Division of Life Science and Medicine, CAS Key Laboratory of Structural Biology, University of Science and Technology of China, Hefei, 230027, China
| | - Qin Deng
- Analytical and Testing Center, Chongqing University, Chongqing, 400030, China
| | - Chao Xu
- Division of Life Science and Medicine, CAS Key Laboratory of Structural Biology, University of Science and Technology of China, Hefei, 230027, China
| | - Qianwen Sun
- Center for Plant Biology, School of Life Sciences, Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China
| | - Shengzhou Wu
- School of Optometry and Ophthalmology and the Eye Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, 325003, China.,State Key Laboratory of Optometry, Ophthalmology, and Visual Science, 270 Xueyuan Road, Wenzhou, Zhejiang, 325003, China
| | - Kaiming Zhang
- MOE Key Laboratory for Membraneless Organelles and Cellular Dynamics, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, 230027, China
| | - Min-Xin Guan
- The Children's Hospital, Zhejiang University School of Medicine and National Clinical Research Center for Child Health, Zhejiang Provincial Key Lab of Genetic and Developmental Disorder, Institute of Genetics, Zhejiang University School of Medicine, Hangzhou, 310016, China
| | - Tianhua Zhou
- Department of Cell Biology and Department of Gastroenterology, Sir Run Run Shaw Hospital, School of Medicine, Cancer Center, Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, China
| | - Fei Sun
- Sir Run-Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, 310016, China
| | - Xiujun Cai
- Sir Run-Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, 310016, China
| | - Chuan Huang
- School of Life Sciences, Chongqing University, Chongqing, 401331, China. .,Center of Plant Functional Genomics, Institute of Advanced Interdisciplinary Studies, Chongqing University, Chongqing, 401331, China.
| | - Ge Shan
- Department of Clinical Laboratory, The First Affiliated Hospital of USTC, the CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei, 230027, China. .,Department of Pulmonary and Critical Care Medicine, Regional Medical Center for National Institute of Respiratory Diseases, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, 310016, China.
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10
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Regulating Phase Transition in Neurodegenerative Diseases by Nuclear Import Receptors. BIOLOGY 2022; 11:biology11071009. [PMID: 36101390 PMCID: PMC9311884 DOI: 10.3390/biology11071009] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/15/2022] [Revised: 06/12/2022] [Accepted: 06/16/2022] [Indexed: 11/17/2022]
Abstract
RNA-binding proteins (RBPs) with a low-complexity prion-like domain (PLD) can undergo aberrant phase transitions and have been implicated in neurodegenerative diseases such as ALS and FTD. Several nuclear RBPs mislocalize to cytoplasmic inclusions in disease conditions. Impairment in nucleocytoplasmic transport is another major event observed in ageing and in neurodegenerative disorders. Nuclear import receptors (NIRs) regulate the nucleocytoplasmic transport of different RBPs bearing a nuclear localization signal by restoring their nuclear localization. NIRs can also specifically dissolve or prevent the aggregation and liquid–liquid phase separation of wild-type or disease-linked mutant RBPs, due to their chaperoning activity. This review focuses on the LLPS of intrinsically disordered proteins and the role of NIRs in regulating LLPS in neurodegeneration. This review also discusses the implication of NIRs as therapeutic agents in neurogenerative diseases.
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11
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Barba-Aliaga M, Alepuz P. Role of eIF5A in Mitochondrial Function. Int J Mol Sci 2022; 23:1284. [PMID: 35163207 PMCID: PMC8835957 DOI: 10.3390/ijms23031284] [Citation(s) in RCA: 31] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2021] [Revised: 01/19/2022] [Accepted: 01/21/2022] [Indexed: 12/17/2022] Open
Abstract
The eukaryotic translation initiation factor 5A (eIF5A) is an evolutionarily conserved protein that binds ribosomes to facilitate the translation of peptide motifs with consecutive prolines or combinations of prolines with glycine and charged amino acids. It has also been linked to other molecular functions and cellular processes, such as nuclear mRNA export and mRNA decay, proliferation, differentiation, autophagy, and apoptosis. The growing interest in eIF5A relates to its association with the pathogenesis of several diseases, including cancer, viral infection, and diabetes. It has also been proposed as an anti-aging factor: its levels decay in aged cells, whereas increasing levels of active eIF5A result in the rejuvenation of the immune and vascular systems and improved brain cognition. Recent data have linked the role of eIF5A in some pathologies with its function in maintaining healthy mitochondria. The eukaryotic translation initiation factor 5A is upregulated under respiratory metabolism and its deficiency reduces oxygen consumption, ATP production, and the levels of several mitochondrial metabolic enzymes, as well as altering mitochondria dynamics. However, although all the accumulated data strongly link eIF5A to mitochondrial function, the precise molecular role and mechanisms involved are still unknown. In this review, we discuss the findings linking eIF5A and mitochondria, speculate about its role in regulating mitochondrial homeostasis, and highlight its potential as a target in diseases related to energy metabolism.
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Affiliation(s)
- Marina Barba-Aliaga
- Instituto de Biotecnología y Biomedicina (Biotecmed), Universitat de València, 46100 València, Spain
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universitat de València, 46100 València, Spain
| | - Paula Alepuz
- Instituto de Biotecnología y Biomedicina (Biotecmed), Universitat de València, 46100 València, Spain
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universitat de València, 46100 València, Spain
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12
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Wing CE, Fung HYJ, Chook YM. Karyopherin-mediated nucleocytoplasmic transport. Nat Rev Mol Cell Biol 2022; 23:307-328. [PMID: 35058649 PMCID: PMC10101760 DOI: 10.1038/s41580-021-00446-7] [Citation(s) in RCA: 159] [Impact Index Per Article: 53.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/09/2021] [Indexed: 12/25/2022]
Abstract
Efficient and regulated nucleocytoplasmic trafficking of macromolecules to the correct subcellular compartment is critical for proper functions of the eukaryotic cell. The majority of the macromolecular traffic across the nuclear pores is mediated by the Karyopherin-β (or Kap) family of nuclear transport receptors. Work over more than two decades has shed considerable light on how the different Kap family members bring their respective cargoes into the nucleus or the cytoplasm in efficient and highly regulated manners. In this Review, we overview the main features and established functions of Kap family members, describe how Kaps recognize their cargoes and discuss the different ways in which these Kap-cargo interactions can be regulated, highlighting new findings and open questions. We also describe current knowledge of the import and export of the components of three large gene expression machines - the core replisome, RNA polymerase II and the ribosome - pointing out the questions that persist about how such large macromolecular complexes are trafficked to serve their function in a designated subcellular location.
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13
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Tauc M, Cougnon M, Carcy R, Melis N, Hauet T, Pellerin L, Blondeau N, Pisani DF. The eukaryotic initiation factor 5A (eIF5A1), the molecule, mechanisms and recent insights into the pathophysiological roles. Cell Biosci 2021; 11:219. [PMID: 34952646 PMCID: PMC8705083 DOI: 10.1186/s13578-021-00733-y] [Citation(s) in RCA: 27] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2021] [Accepted: 12/14/2021] [Indexed: 11/29/2022] Open
Abstract
Since the demonstration of its involvement in cell proliferation, the eukaryotic initiation factor 5A (eIF5A) has been studied principally in relation to the development and progression of cancers in which the isoform A2 is mainly expressed. However, an increasing number of studies report that the isoform A1, which is ubiquitously expressed in normal cells, exhibits novel molecular features that reveal its new relationships between cellular functions and organ homeostasis. At a first glance, eIF5A can be regarded, among other things, as a factor implicated in the initiation of translation. Nevertheless, at least three specificities: (1) its extreme conservation between species, including plants, throughout evolution, (2) its very special and unique post-translational modification through the activating-hypusination process, and finally (3) its close relationship with the polyamine pathway, suggest that the role of eIF5A in living beings remains to be uncovered. In fact, and beyond its involvement in facilitating the translation of proteins containing polyproline residues, eIF5A is implicated in various physiological processes including ischemic tolerance, metabolic adaptation, aging, development, and immune cell differentiation. These newly discovered physiological properties open up huge opportunities in the clinic for pathologies such as, for example, the ones in which the oxygen supply is disrupted. In this latter case, organ transplantation, myocardial infarction or stroke are concerned, and the current literature defines eIF5A as a new drug target with a high level of potential benefit for patients with these diseases or injuries. Moreover, the recent use of genomic and transcriptomic association along with metadata studies also revealed the implication of eIF5A in genetic diseases. Thus, this review provides an overview of eIF5A from its molecular mechanism of action to its physiological roles and the clinical possibilities that have been recently reported in the literature.
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Affiliation(s)
- Michel Tauc
- LP2M, CNRS, Université Côte d'Azur, Nice, France. .,Laboratories of Excellence Ion Channel Science and Therapeutics, Nice, France. .,Laboratoire de Physiomédecine Moléculaire, UMR7370, Faculté de Médecine, CNRS, Université Côte d'Azur, 28 Avenue de Valombrose, 06107, Nice Cedex, France.
| | - Marc Cougnon
- LP2M, CNRS, Université Côte d'Azur, Nice, France.,Laboratories of Excellence Ion Channel Science and Therapeutics, Nice, France
| | - Romain Carcy
- Service de Réanimation Polyvalente et Service de Réanimation des Urgences Vitales, CHU Nice, Hôpital Pasteur 2, Nice, France
| | - Nicolas Melis
- Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, Bethesda, MD, 20892, USA
| | - Thierry Hauet
- INSERM, IRTOMIT, CHU de Poitiers, Université de Poitiers, La Milétrie, Poitiers, France
| | - Luc Pellerin
- INSERM, IRTOMIT, CHU de Poitiers, Université de Poitiers, La Milétrie, Poitiers, France
| | - Nicolas Blondeau
- Laboratories of Excellence Ion Channel Science and Therapeutics, Nice, France.,IPMC, CNRS, Université Côte d'Azur, Valbonne, France
| | - Didier F Pisani
- LP2M, CNRS, Université Côte d'Azur, Nice, France.,Laboratories of Excellence Ion Channel Science and Therapeutics, Nice, France
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14
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Pasha T, Zatorska A, Sharipov D, Rogelj B, Hortobágyi T, Hirth F. Karyopherin abnormalities in neurodegenerative proteinopathies. Brain 2021; 144:2915-2932. [PMID: 34019093 PMCID: PMC8194669 DOI: 10.1093/brain/awab201] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2020] [Revised: 04/08/2021] [Accepted: 05/11/2021] [Indexed: 11/12/2022] Open
Abstract
Neurodegenerative proteinopathies are characterized by progressive cell loss that is preceded by the mislocalization and aberrant accumulation of proteins prone to aggregation. Despite their different physiological functions, disease-related proteins like tau, α-synuclein, TAR DNA binding protein-43, fused in sarcoma and mutant huntingtin, all share low complexity regions that can mediate their liquid-liquid phase transitions. The proteins' phase transitions can range from native monomers to soluble oligomers, liquid droplets and further to irreversible, often-mislocalized aggregates that characterize the stages and severity of neurodegenerative diseases. Recent advances into the underlying pathogenic mechanisms have associated mislocalization and aberrant accumulation of disease-related proteins with defective nucleocytoplasmic transport and its mediators called karyopherins. These studies identify karyopherin abnormalities in amyotrophic lateral sclerosis, frontotemporal dementia, Alzheimer's disease, and synucleinopathies including Parkinson's disease and dementia with Lewy bodies, that range from altered expression levels to the subcellular mislocalization and aggregation of karyopherin α and β proteins. The reported findings reveal that in addition to their classical function in nuclear import and export, karyopherins can also act as chaperones by shielding aggregation-prone proteins against misfolding, accumulation and irreversible phase-transition into insoluble aggregates. Karyopherin abnormalities can, therefore, be both the cause and consequence of protein mislocalization and aggregate formation in degenerative proteinopathies. The resulting vicious feedback cycle of karyopherin pathology and proteinopathy identifies karyopherin abnormalities as a common denominator of onset and progression of neurodegenerative disease. Pharmacological targeting of karyopherins, already in clinical trials as therapeutic intervention targeting cancers such as glioblastoma and viral infections like COVID-19, may therefore represent a promising new avenue for disease-modifying treatments in neurodegenerative proteinopathies.
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Affiliation(s)
- Terouz Pasha
- King’s College London, Institute of Psychiatry, Psychology and Neuroscience, Maurice Wohl Clinical Neuroscience Institute, Department of Basic and Clinical Neuroscience, Institute of Psychiatry, Psychology and Neuroscience, London SE5 9RT, UK
| | - Anna Zatorska
- King’s College London, Institute of Psychiatry, Psychology and Neuroscience, Maurice Wohl Clinical Neuroscience Institute, Department of Basic and Clinical Neuroscience, Institute of Psychiatry, Psychology and Neuroscience, London SE5 9RT, UK
| | - Daulet Sharipov
- King’s College London, Institute of Psychiatry, Psychology and Neuroscience, Maurice Wohl Clinical Neuroscience Institute, Department of Basic and Clinical Neuroscience, Institute of Psychiatry, Psychology and Neuroscience, London SE5 9RT, UK
| | - Boris Rogelj
- Jozef Stefan Institute, Department of Biotechnology, 1000 Ljubljana, Slovenia
- University of Ljubljana, Faculty of Chemistry and Chemical Technology, 1000 Ljubljana, Slovenia
| | - Tibor Hortobágyi
- ELKH-DE Cerebrovascular and Neurodegenerative Research Group, Department of Neurology, University of Debrecen, 4032 Debrecen, Hungary
- King's College London, Department of Old Age Psychiatry, Institute of Psychiatry, Psychology and Neuroscience, London SE5 8AF, UK
| | - Frank Hirth
- King’s College London, Institute of Psychiatry, Psychology and Neuroscience, Maurice Wohl Clinical Neuroscience Institute, Department of Basic and Clinical Neuroscience, Institute of Psychiatry, Psychology and Neuroscience, London SE5 9RT, UK
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15
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Kimura M, Imai K, Morinaka Y, Hosono-Sakuma Y, Horton P, Imamoto N. Distinct mutations in importin-β family nucleocytoplasmic transport receptors transportin-SR and importin-13 affect specific cargo binding. Sci Rep 2021; 11:15649. [PMID: 34341383 PMCID: PMC8329185 DOI: 10.1038/s41598-021-94948-1] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2020] [Accepted: 07/20/2021] [Indexed: 01/25/2023] Open
Abstract
Importin-(Imp)β family nucleocytoplasmic transport receptors (NTRs) are supposed to bind to their cargoes through interaction between a confined interface on an NTR and a nuclear localization or export signal (NLS/NES) on a cargo. Although consensus NLS/NES sequence motifs have been defined for cargoes of some NTRs, many experimentally identified cargoes of those NTRs lack those motifs, and consensus NLSs/NESs have been reported for only a few NTRs. Crystal structures of NTR-cargo complexes have exemplified 3D structure-dependent binding of cargoes lacking a consensus NLS/NES to different sites on an NTR. Since only a limited number of NTR-cargo interactions have been studied, whether most cargoes lacking a consensus NLS/NES bind to the same confined interface or to various sites on an NTR is still unclear. Addressing this issue, we generated four mutants of transportin-(Trn)SR, of which many cargoes lack a consensus NLS, and eight mutants of Imp13, where no consensus NLS has been defined, and we analyzed their binding to as many as 40 cargo candidates that we previously identified by a nuclear import reaction-based method. The cargoes bind differently to the NTR mutants, suggesting that positions on an NTR contribute differently to the binding of respective cargoes.
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Affiliation(s)
- Makoto Kimura
- Cellular Dynamics Laboratory, RIKEN Cluster for Pioneering Research, Wako, Saitama, Japan.
| | - Kenichiro Imai
- Cellular and Molecular Biotechnology Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Tokyo, Japan.
- Molecular Profiling Research Center for Drug Discovery, National Institute of Advanced Industrial Science and Technology (AIST), Tokyo, Japan.
| | - Yuriko Morinaka
- Cellular Dynamics Laboratory, RIKEN Cluster for Pioneering Research, Wako, Saitama, Japan
| | - Yoshiko Hosono-Sakuma
- Cellular Dynamics Laboratory, RIKEN Cluster for Pioneering Research, Wako, Saitama, Japan
| | - Paul Horton
- Department of Computer Science and Information Engineering, National Cheng Kung University, Tainan City, Taiwan
| | - Naoko Imamoto
- Cellular Dynamics Laboratory, RIKEN Cluster for Pioneering Research, Wako, Saitama, Japan.
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16
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Affiliation(s)
- Shao-Jung Hsu
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC
- Faculty of Medicine, National Yang Ming Chiao Tung University, Hsinchu, Taiwan, ROC
| | - Hui-Chun Huang
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC
- Faculty of Medicine, National Yang Ming Chiao Tung University, Hsinchu, Taiwan, ROC
- Division of General Medicine, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC
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17
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Unique evolutionary trajectories of breast cancers with distinct genomic and spatial heterogeneity. Sci Rep 2021; 11:10571. [PMID: 34011996 PMCID: PMC8134446 DOI: 10.1038/s41598-021-90170-1] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2021] [Accepted: 04/30/2021] [Indexed: 11/08/2022] Open
Abstract
Breast cancers exhibit intratumoral heterogeneity associated with disease progression and therapeutic resistance. To define the sources and the extent of heterogeneity, we performed an in-depth analysis of the genomic architecture of three chemoradiation-naïve breast cancers with well-defined clinical features including variable ER, PR, ERBB2 receptor expression and two distinct pathogenic BRCA2mut genotypes. The latter included a germ line carrier and a patient with a somatic variant. In each case we combined DNA content-based flow cytometry with whole exome sequencing and genome wide copy number variant (CNV) analysis of distinct populations sorted from multiple (4–18) mapped biopsies within the tumors and involved lymph nodes. Interrogating flow-sorted tumor populations from each biopsy provided an objective method to distinguish fixed and variable genomic lesions in each tumor. Notably we show that tumors exploit CNVs to fix mutations and deletions in distinct populations throughout each tumor. The identification of fixed genomic lesions that are shared or unique within each tumor, has broad implications for the study of tumor heterogeneity including the presence of tumor markers and therapeutic targets, and of candidate neoepitopes in breast and other solid tumors that can advance more effective treatment and clinical management of patients with disease.
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18
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Ding B, Sepehrimanesh M. Nucleocytoplasmic Transport: Regulatory Mechanisms and the Implications in Neurodegeneration. Int J Mol Sci 2021; 22:4165. [PMID: 33920577 PMCID: PMC8072611 DOI: 10.3390/ijms22084165] [Citation(s) in RCA: 38] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2021] [Revised: 04/13/2021] [Accepted: 04/15/2021] [Indexed: 12/12/2022] Open
Abstract
Nucleocytoplasmic transport (NCT) across the nuclear envelope is precisely regulated in eukaryotic cells, and it plays critical roles in maintenance of cellular homeostasis. Accumulating evidence has demonstrated that dysregulations of NCT are implicated in aging and age-related neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), Alzheimer's disease (AD), and Huntington disease (HD). This is an emerging research field. The molecular mechanisms underlying impaired NCT and the pathogenesis leading to neurodegeneration are not clear. In this review, we comprehensively described the components of NCT machinery, including nuclear envelope (NE), nuclear pore complex (NPC), importins and exportins, RanGTPase and its regulators, and the regulatory mechanisms of nuclear transport of both protein and transcript cargos. Additionally, we discussed the possible molecular mechanisms of impaired NCT underlying aging and neurodegenerative diseases, such as ALS/FTD, HD, and AD.
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Affiliation(s)
- Baojin Ding
- Department of Biology, University of Louisiana at Lafayette, 410 East Saint Mary Boulevard, Lafayette, LA 70503, USA;
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19
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Kalita J, Kapinos LE, Lim RYH. On the asymmetric partitioning of nucleocytoplasmic transport - recent insights and open questions. J Cell Sci 2021; 134:239102. [PMID: 33912945 DOI: 10.1242/jcs.240382] [Citation(s) in RCA: 23] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Macromolecular cargoes are asymmetrically partitioned in the nucleus or cytoplasm by nucleocytoplasmic transport (NCT). At the center of this activity lies the nuclear pore complex (NPC), through which soluble factors circulate to orchestrate NCT. These include cargo-carrying importin and exportin receptors from the β-karyopherin (Kapβ) family and the small GTPase Ran, which switches between guanosine triphosphate (GTP)- and guanosine diphosphate (GDP)-bound forms to regulate cargo delivery and compartmentalization. Ongoing efforts have shed considerable light on how these soluble factors traverse the NPC permeability barrier to sustain NCT. However, this does not explain how importins and exportins are partitioned in the cytoplasm and nucleus, respectively, nor how a steep RanGTP-RanGDP gradient is maintained across the nuclear envelope. In this Review, we peel away the multiple layers of control that regulate NCT and juxtapose unresolved features against known aspects of NPC function. Finally, we discuss how NPCs might function synergistically with Kapβs, cargoes and Ran to establish the asymmetry of NCT.
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Affiliation(s)
- Joanna Kalita
- Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel CH4056, Switzerland
| | - Larisa E Kapinos
- Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel CH4056, Switzerland
| | - Roderick Y H Lim
- Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel CH4056, Switzerland
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20
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Mehmood R, Jibiki K, Shibazaki N, Yasuhara N. Molecular profiling of nucleocytoplasmic transport factor genes in breast cancer. Heliyon 2021; 7:e06039. [PMID: 33553736 PMCID: PMC7851789 DOI: 10.1016/j.heliyon.2021.e06039] [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: 09/27/2020] [Revised: 12/14/2020] [Accepted: 01/14/2021] [Indexed: 11/24/2022] Open
Abstract
Transport of functional molecules across the nuclear membrane of a eukaryotic cell is regulated by a dedicated set of transporter proteins that carry molecules into the nucleus or out of the nucleus to the cytoplasm for homeostasis of the cell. One of the categories of cargo molecules these transporters carry are the molecules for cell cycle regulation. Therefore, their role is critical in terms of cancer development. Any misregulation of the transport factors would means aberrant abundance of cell cycle regulators and might have consequences in cell cycle progression. While earlier studies have focussed on individual transport related molecules, a collective overview of how these molecules may be dysregulated in breast cancer is lacking. Using genomic and transcriptomic datasets from TCGA (The Cancer Genome Atlas) and microarray platforms, we carried out bioinformatic analysis and provide a genetic and molecular profile of all the molecules directly related to nucleocytoplasmic shuttling of proteins and RNAs. Interestingly, we identified that many of these molecules are either mutated or have dysregulated expression in breast cancer. Strikingly, some of the molecules, namely, KPNA2, KPNA3, KPNA5, IPO8, TNPO1, XPOT, XPO7 and CSE1L were correlated with poor patient survival. This study provides a comprehensive genetic and molecular landscape of nucleocytoplasmic factors in breast cancer and points to the important roles of various nucleocytoplasmic factors in cancer progression. This data might have implications in prognosis and therapeutic targeting in breast cancer.
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Affiliation(s)
- Rashid Mehmood
- Department of Life Sciences, College of Science and General Studies, Alfaisal University, Riyadh, Kingdom of Saudi Arabia
| | - Kazuya Jibiki
- Graduate School of Integrated Basic Sciences, Nihon University, Setagaya-ku, Tokyo, Japan
| | - Noriko Shibazaki
- Graduate School of Integrated Basic Sciences, Nihon University, Setagaya-ku, Tokyo, Japan
| | - Noriko Yasuhara
- Graduate School of Integrated Basic Sciences, Nihon University, Setagaya-ku, Tokyo, Japan
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21
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Jeelani G, Nozaki T. Eukaryotic translation initiation factor 5A and its posttranslational modifications play an important role in proliferation and potentially in differentiation of the human enteric protozoan parasite Entamoeba histolytica. PLoS Pathog 2021; 17:e1008909. [PMID: 33592076 PMCID: PMC7909649 DOI: 10.1371/journal.ppat.1008909] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2020] [Revised: 02/26/2021] [Accepted: 01/19/2021] [Indexed: 11/19/2022] Open
Abstract
The eukaryotic translation initiation factor 5A (eIF5A) is a highly conserved protein and is essential in all eukaryotes. However, the specific roles of eIF5A in translation and in other biological processes remain elusive. In the present study, we described the role of eIF5A, its posttranslational modifications (PTM), and the biosynthetic pathway needed for the PTM in Entamoeba histolytica, the protozoan parasite responsible for amoebic dysentery and liver abscess in humans. E. histolytica encodes two isotypes of eIF5A and two isotypes of enzymes, deoxyhypusine synthase (DHS), responsible for their PTM. Both of the two eIF5A isotypes are functional, whereas only one DHS (EhDHS1, but not EhDHS2), is catalytically active. The DHS activity increased ~2000-fold when EhDHS1 was co-expressed with EhDHS2 in Escherichia coli, suggesting that the formation of a heteromeric complex is needed for full enzymatic activity. Both EhDHS1 and 2 genes were required for in vitro growth of E. histolytica trophozoites, indicated by small antisense RNA-mediated gene silencing. In trophozoites, only eIF5A2, but not eIF5A1, gene was actively transcribed. Gene silencing of eIF5A2 caused compensatory induction of expression of eIF5A1 gene, suggesting interchangeable role of the two eIF5A isotypes and also reinforcing the importance of eIF5As for parasite proliferation and survival. Furthermore, using a sibling species, Entamoeba invadens, we found that eIF5A1 gene was upregulated during excystation, while eIF5A2 was downregulated, suggesting that eIF5A1 gene plays an important role during differentiation. Taken together, these results have underscored the essentiality of eIF5A and DHS, for proliferation and potentially in the differentiation of this parasite, and suggest that the hypusination associated pathway represents a novel rational target for drug development against amebiasis.
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Affiliation(s)
- Ghulam Jeelani
- Department of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, Japan
| | - Tomoyoshi Nozaki
- Department of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, Japan
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22
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Wu GQ, Xu YM, Lau ATY. Recent insights into eukaryotic translation initiation factors 5A1 and 5A2 and their roles in human health and disease. Cancer Cell Int 2020; 20:142. [PMID: 32368188 PMCID: PMC7191727 DOI: 10.1186/s12935-020-01226-7] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2020] [Accepted: 04/20/2020] [Indexed: 02/05/2023] Open
Abstract
The eukaryotic translation initiation factor 5A1 (eIF5A1) and its homolog eIF5A2 are the only two human proteins containing the unique post-translational modification-hypusination, which is essential for the function of these two proteins. eIF5A1 was initially identified as a translation initiation factor by promoting the first peptide bond formation of protein during translation; however, recent results suggest that eIF5A1 also functions as a translation elongation factor. It has been shown that eIF5A1 is implicated in certain human diseases, including diabetes, several human cancer types, viral infections and diseases of neural system. Meanwhile, eIF5A2 is overexpressed in many cancers, and plays an important role in the development and progression of cancers. As multiple roles of these two factors were observed among these studies, therefore, it remains unclear whether they act as oncogene or tumor suppressor. In this review, the recent literature of eIF5As and their roles in human diseases, especially in human cancers, will be discussed.
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Affiliation(s)
- Gao-Qi Wu
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou, Guangdong 515041 People’s Republic of China
| | - Yan-Ming Xu
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou, Guangdong 515041 People’s Republic of China
| | - Andy T. Y. Lau
- Laboratory of Cancer Biology and Epigenetics, Department of Cell Biology and Genetics, Shantou University Medical College, Shantou, Guangdong 515041 People’s Republic of China
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23
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Ernst S, Müller-Newen G. Nucleocytoplasmic Shuttling of STATs. A Target for Intervention? Cancers (Basel) 2019; 11:cancers11111815. [PMID: 31752278 PMCID: PMC6895884 DOI: 10.3390/cancers11111815] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2019] [Revised: 11/08/2019] [Accepted: 11/13/2019] [Indexed: 12/12/2022] Open
Abstract
Signal transducer and activator of transcription (STAT) proteins are transcription factors that in the latent state are located predominantly in the cytoplasm. Activation of STATs through phosphorylation of a single tyrosine residue results in nuclear translocation. The requirement of tyrosine phosphorylation for nuclear accumulation is shared by all STAT family members but mechanisms of nuclear translocation vary between different STATs. These differences offer opportunities for specific intervention. To achieve this, the molecular mechanisms of nucleocytoplasmic shuttling of STATs need to be understood in more detail. In this review we will give an overview on the various aspects of nucleocytoplasmic shuttling of latent and activated STATs with a special focus on STAT3 and STAT5. Potential targets for cancer treatment will be identified and discussed.
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Affiliation(s)
- Sabrina Ernst
- Institute of Biochemistry and Molecular Biology, RWTH Aachen University, 52074 Aachen, Germany;
- Confocal Microscopy Facility, Interdisciplinary Center for Clinical Research IZKF, RWTH Aachen University, 52074 Aachen, Germany
| | - Gerhard Müller-Newen
- Institute of Biochemistry and Molecular Biology, RWTH Aachen University, 52074 Aachen, Germany;
- Correspondence:
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Single-Cell Transcriptomic Analyses of Cell Fate Transitions during Human Cardiac Reprogramming. Cell Stem Cell 2019; 25:149-164.e9. [PMID: 31230860 DOI: 10.1016/j.stem.2019.05.020] [Citation(s) in RCA: 101] [Impact Index Per Article: 16.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2017] [Revised: 03/21/2019] [Accepted: 05/22/2019] [Indexed: 12/11/2022]
Abstract
Direct cellular reprogramming provides a powerful platform to study cell plasticity and dissect mechanisms underlying cell fate determination. Here, we report a single-cell transcriptomic study of human cardiac (hiCM) reprogramming that utilizes an analysis pipeline incorporating current data normalization methods, multiple trajectory prediction algorithms, and a cell fate index calculation we developed to measure reprogramming progression. These analyses revealed hiCM reprogramming-specific features and a decision point at which cells either embark on reprogramming or regress toward their original fibroblast state. In combination with functional screening, we found that immune-response-associated DNA methylation is required for hiCM induction and validated several downstream targets of reprogramming factors as necessary for productive hiCM reprograming. Collectively, this single-cell transcriptomics study provides detailed datasets that reveal molecular features underlying hiCM determination and rigorous analytical pipelines for predicting cell fate conversion.
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25
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Meng QB, Peng JJ, Qu ZW, Zhu XM, Wen Z, Kang WM. Eukaryotic initiation factor 5A2 and human digestive system neoplasms. World J Gastrointest Oncol 2019; 11:449-458. [PMID: 31236196 PMCID: PMC6580320 DOI: 10.4251/wjgo.v11.i6.449] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/31/2018] [Revised: 04/17/2019] [Accepted: 05/04/2019] [Indexed: 02/05/2023] Open
Abstract
Eukaryotic initiation factor 5A2 (eIF5A2), as one of the two isoforms in the family, is reported to be a novel oncogenic protein that is involved in multiple aspects of many types of human cancer. Overexpression or gene amplification of EIF5A2 has been demonstrated in many cancers. Accumulated evidence shows that eIF5A2 initiates tumor formation, enhances cancer cell growth, increases cancer cell metastasis, and promotes treatment resistance through multiple means, including inducing epithelial–mesenchymal transition, cytoskeletal rearrangement, angiogenesis, and metabolic reprogramming. Expression of eIF5A2 in cancer correlates with poor survival, advanced disease stage, as well as metastasis, suggesting that eIF5A2 function is crucial for tumor development and maintenance but not for normal tissue homeostasis. All these studies suggest that eIF5A2 is a useful biomarker in the prediction of cancer prognosis and serves as an anticancer molecular target. This review focuses on the expression, subcellular localization, post-translational modifications, and regulatory networks of eIF5A2, as well as its biochemical functions and evolving clinical applications in cancer, especially in human digestive system neoplasms.
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Affiliation(s)
- Qing-Bin Meng
- Department of Gastrointestinal Surgery, the First Hospital of Wuhan City, Wuhan 430022, Hubei Province, China
| | - Jing-Jing Peng
- Department of Gastroenterology, General Hospital of the Yangtze River Shipping, Wuhan 430015, Hubei Province, China
| | - Zi-Wei Qu
- Department of Gastrointestinal Surgery, the First Hospital of Wuhan City, Wuhan 430022, Hubei Province, China
| | | | - Zhang Wen
- Department of Hepato-Biliary-Pancreatic Surgery and Liver Transplantation, the First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China
| | - Wei-Ming Kang
- Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China
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26
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Vera Rodriguez A, Frey S, Görlich D. Engineered SUMO/protease system identifies Pdr6 as a bidirectional nuclear transport receptor. J Cell Biol 2019; 218:2006-2020. [PMID: 31023724 PMCID: PMC6548132 DOI: 10.1083/jcb.201812091] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2018] [Revised: 03/26/2019] [Accepted: 03/28/2019] [Indexed: 01/07/2023] Open
Abstract
Cleavage of affinity tags by specific proteases can be exploited for highly selective affinity chromatography. The SUMO/SENP1 system is the most efficient for such application but fails in eukaryotic expression because it cross-reacts with endogenous proteases. Using a novel selection system, we have evolved the SUMOEu/SENP1Eu pair to orthogonality with the yeast and animal enzymes. SUMOEu fusions therefore remain stable in eukaryotic cells. Likewise, overexpressing a SENP1Eu protease is nontoxic in yeast. We have used the SUMOEu system in an affinity-capture-proteolytic-release approach to identify interactors of the yeast importin Pdr6/Kap122. This revealed not only further nuclear import substrates such as Ubc9, but also Pil1, Lsp1, eIF5A, and eEF2 as RanGTP-dependent binders and thus as export cargoes. We confirmed that Pdr6 functions as an exportin in vivo and depletes eIF5A and eEF2 from cell nuclei. Thus, Pdr6 is a bidirectional nuclear transport receptor (i.e., a biportin) that shuttles distinct sets of cargoes in opposite directions.
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Affiliation(s)
- Arturo Vera Rodriguez
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Steffen Frey
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Dirk Görlich
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
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27
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Aksu M, Trakhanov S, Vera Rodriguez A, Görlich D. Structural basis for the nuclear import and export functions of the biportin Pdr6/Kap122. J Cell Biol 2019; 218:1839-1852. [PMID: 31023722 PMCID: PMC6548137 DOI: 10.1083/jcb.201812093] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2018] [Revised: 03/26/2019] [Accepted: 03/28/2019] [Indexed: 12/26/2022] Open
Abstract
Importins ferry proteins into nuclei while exportins carry cargoes to the cytoplasm. In the accompanying paper in this issue (Vera Rodriguez et al. 2019. J. Cell Biol. https://doi.org/10.1083/jcb.201812091), we discovered that Pdr6 is a biportin that imports, e.g., the SUMO E2 ligase Ubc9 while depleting the translation factor eIF5A from the nuclear compartment. In this paper, we report the structures of key transport intermediates, namely, of the Ubc9•Pdr6 import complex, of the RanGTP•Pdr6 heterodimer, and of the trimeric RanGTP•Pdr6•eIF5A export complex. These revealed nonlinear transport signals, chaperone-like interactions, and how the RanGTPase system drives Pdr6 to transport Ubc9 and eIF5A in opposite directions. The structures also provide unexpected insights into the evolution of transport selectivity. Specifically, they show that recognition of Ubc9 by Pdr6 differs fundamentally from that of the human Ubc9-importer Importin 13. Likewise, Pdr6 recognizes eIF5A in a nonhomologous manner compared with the mammalian eIF5A-exporter Exportin 4. This suggests that the import of Ubc9 and active nuclear exclusion of eIF5A evolved in different eukaryotic lineages more than once and independently from each other.
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Affiliation(s)
- Metin Aksu
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Sergei Trakhanov
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Arturo Vera Rodriguez
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Dirk Görlich
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
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28
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de Proença ARG, Pereira KD, Meneguello L, Tamborlin L, Luchessi AD. Insulin action on protein synthesis and its association with eIF5A expression and hypusination. Mol Biol Rep 2019; 46:587-596. [PMID: 30519811 DOI: 10.1007/s11033-018-4512-1] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2018] [Accepted: 11/19/2018] [Indexed: 12/20/2022]
Abstract
The hormone insulin plays a central role in the metabolism of carbohydrates, lipids, and proteins. In relation to protein metabolism, insulin stimulates amino acid uptake and activates protein synthesis in responsive cells by modulation of signal transduction pathways, such as associated to Akt/PkB, mTOR, S6Ks, 4E-BP1, and several translation initiation/elongation factors. In this context, there is no information on direct cellular treatment with insulin and effects on eukaryotic translation initiation factor 5A (eIF5A) regulation. The eIF5A protein contains an exclusive amino acid residue denominated hypusine, which is essential for its activity and synthesized by posttranslational modification of a specific lysine residue using spermidine as substrate. The eIF5A protein is involved in cellular proliferation and differentiation processes, as observed for satellite cells derived from rat muscles, revealing that eIF5A has an important role in muscle regeneration. The aim of this study was to determine whether eIF5A expression and hypusination are influenced by direct treatment of insulin on L6 myoblast cells. We observed that insulin increased the content of eIF5A transcripts. This effect occurred in cells treated or depleted of fetal bovine serum, revealing a positive insulin effect independent of other serum components. In addition, it was observed that hypusination follows the maintenance of eIF5A protein content in the serum depleted cells and treated with insulin. These results demonstrate that eIF5A is modulated by insulin, contributing the protein synthesis machinery control, as observed by puromycin incorporation in nascent proteins.
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Affiliation(s)
| | - Karina Danielle Pereira
- Laboratory of Biotechnology, School of Applied Sciences, University of Campinas (UNICAMP), Limeira, São Paulo, Brazil
- Institute of Biosciences, São Paulo State University (UNESP), Rio Claro, São Paulo, Brazil
| | - Leticia Meneguello
- Laboratory of Biotechnology, School of Applied Sciences, University of Campinas (UNICAMP), Limeira, São Paulo, Brazil
- Institute of Biosciences, São Paulo State University (UNESP), Rio Claro, São Paulo, Brazil
| | - Leticia Tamborlin
- Laboratory of Biotechnology, School of Applied Sciences, University of Campinas (UNICAMP), Limeira, São Paulo, Brazil
- Institute of Biosciences, São Paulo State University (UNESP), Rio Claro, São Paulo, Brazil
| | - Augusto Ducati Luchessi
- Laboratory of Biotechnology, School of Applied Sciences, University of Campinas (UNICAMP), Limeira, São Paulo, Brazil.
- Institute of Biosciences, São Paulo State University (UNESP), Rio Claro, São Paulo, Brazil.
- Laboratório de Biotecnologia, Faculdade de Ciências Aplicadas, Universidade Estadual de Campinas, Rua Pedro Zaccaria, 1300, Limeira, São Paulo, 13484-350, Brazil.
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29
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Aksu M, Pleiner T, Karaca S, Kappert C, Dehne HJ, Seibel K, Urlaub H, Bohnsack MT, Görlich D. Xpo7 is a broad-spectrum exportin and a nuclear import receptor. J Cell Biol 2018; 217:2329-2340. [PMID: 29748336 PMCID: PMC6028547 DOI: 10.1083/jcb.201712013] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2017] [Revised: 04/09/2018] [Accepted: 04/24/2018] [Indexed: 11/28/2022] Open
Abstract
Exportins bind cargo molecules in a RanGTP-dependent manner inside nuclei and transport them through nuclear pores to the cytoplasm. CRM1/Xpo1 is the best-characterized exportin because specific inhibitors such as leptomycin B allow straightforward cargo validations in vivo. The analysis of other exportins lagged far behind, foremost because no such inhibitors had been available for them. In this study, we explored the cargo spectrum of exportin 7/Xpo7 in depth and identified not only ∼200 potential export cargoes but also, surprisingly, ∼30 nuclear import substrates. Moreover, we developed anti-Xpo7 nanobodies that acutely block Xpo7 function when transfected into cultured cells. The inhibition is pathway specific, mislocalizes export cargoes of Xpo7 to the nucleus and import substrates to the cytoplasm, and allowed validation of numerous tested cargo candidates. This establishes Xpo7 as a broad-spectrum bidirectional transporter and paves the way for a much deeper analysis of exportin and importin function in the future.
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Affiliation(s)
- Metin Aksu
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Tino Pleiner
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Samir Karaca
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Christin Kappert
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Heinz-Jürgen Dehne
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Katharina Seibel
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Henning Urlaub
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
- Institute for Clinical Chemistry, University Medical Center Göttingen, Göttingen, Germany
| | - Markus T Bohnsack
- Institute for Molecular Biology, University Medical Center Göttingen, Göttingen, Germany
| | - Dirk Görlich
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
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30
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Baade I, Spillner C, Schmitt K, Valerius O, Kehlenbach RH. Extensive Identification and In-depth Validation of Importin 13 Cargoes. Mol Cell Proteomics 2018; 17:1337-1353. [PMID: 29666159 PMCID: PMC6030721 DOI: 10.1074/mcp.ra118.000623] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2018] [Revised: 04/09/2018] [Indexed: 11/06/2022] Open
Abstract
Importin 13 is a member of the importin β family of transport receptors. Unlike most family members, importin 13 mediates both, nuclear protein import and export. To search for novel importin 13 cargoes, we used stable isotope labeling of amino acids in cell culture (SILAC) and mass spectrometry. Using stringent criteria, we identified 255 importin 13 substrates, including the known cargoes Ubc9, Mago and eIF1A, and validate many of them as transport cargoes by extensive biochemical and cell biological characterization. Several novel cargoes can also be transported by the export receptor CRM1, demonstrating a clear redundancy in receptor choice. Using importin 13 mutants, we show that many of the novel substrates contact regions on the transport receptor that are not used by Ubc9, Mago or eIF1A. Together, this study significantly expands the repertoire of importin 13 cargoes and sets the basis for a more detailed characterization of this extremely versatile transport receptor.
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Affiliation(s)
- Imke Baade
- From the ‡Department of Molecular Biology, Faculty of Medicine, Göttingen Center of Biosciences (GZMB), Georg-August-University Göttingen, Humboldtallee 23, 37073 Göttingen, Germany
| | - Christiane Spillner
- From the ‡Department of Molecular Biology, Faculty of Medicine, Göttingen Center of Biosciences (GZMB), Georg-August-University Göttingen, Humboldtallee 23, 37073 Göttingen, Germany
| | - Kerstin Schmitt
- §Department of Molecular Microbiology and Genetics, Institute of Microbiology and Genetics, GZMB, Georg-August-University Göttingen, Grisebachstr 8, 37077 Göttingen, Germany
| | - Oliver Valerius
- §Department of Molecular Microbiology and Genetics, Institute of Microbiology and Genetics, GZMB, Georg-August-University Göttingen, Grisebachstr 8, 37077 Göttingen, Germany
| | - Ralph H Kehlenbach
- From the ‡Department of Molecular Biology, Faculty of Medicine, Göttingen Center of Biosciences (GZMB), Georg-August-University Göttingen, Humboldtallee 23, 37073 Göttingen, Germany;
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31
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Hoque M, Park JY, Chang YJ, Luchessi AD, Cambiaghi TD, Shamanna R, Hanauske-Abel HM, Holland B, Pe'ery T, Tian B, Mathews MB. Regulation of gene expression by translation factor eIF5A: Hypusine-modified eIF5A enhances nonsense-mediated mRNA decay in human cells. ACTA ACUST UNITED AC 2017; 5:e1366294. [PMID: 29034140 DOI: 10.1080/21690731.2017.1366294] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2017] [Revised: 07/28/2017] [Accepted: 08/07/2017] [Indexed: 10/19/2022]
Abstract
Nonsense-mediated mRNA decay (NMD) couples protein synthesis to mRNA turnover. It eliminates defective transcripts and controls the abundance of certain normal mRNAs. Our study establishes a connection between NMD and the translation factor eIF5A (eukaryotic initiation factor 5A) in human cells. eIF5A modulates the synthesis of groups of proteins (the eIF5A regulon), and undergoes a distinctive two-step post-translational modification (hypusination) catalyzed by deoxyhypusine synthase and deoxyhypusine hydroxylase. We show that expression of NMD-susceptible constructs was increased by depletion of the major eIF5A isoform, eIF5A1. NMD was also attenuated when hypusination was inhibited by RNA interference with either of the two eIF5A modifying enzymes, or by treatment with the drugs ciclopirox or deferiprone which inhibit deoxyhypusine hydroxylase. Transcriptome analysis by RNA-Seq identified human genes whose expression is coordinately regulated by eIF5A1, its modifying enzymes, and the pivotal NMD factor, Upf1. Transcripts encoding components of the translation system were highly represented, including some encoding ribosomal proteins controlled by alternative splicing coupled to NMD (AS-NMD). Our findings extend and strengthen the association of eIF5A with NMD, previously inferred in yeast, and show that hypusination is important for this function of human eIF5A. In addition, they advance drug-mediated NMD suppression as a therapeutic opportunity for nonsense-associated diseases. We propose that regulation of mRNA stability contributes to eIF5A's role in selective gene expression.
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Affiliation(s)
- Mainul Hoque
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Ji Yeon Park
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Yun-Juan Chang
- Department of Microbiology, Biochemistry & Molecular Genetics, Rutgers New Jersey Medical School, Newark, NJ, USA.,Office of Advanced Research Computing, Rutgers University, Newark, NJ, USA
| | - Augusto D Luchessi
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA.,Laboratory of Biotechnology, School of Applied Sciences, University of Campinas, Limeira, São Paulo, Brazil
| | - Tavane D Cambiaghi
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Raghavendra Shamanna
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Hartmut M Hanauske-Abel
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Bart Holland
- Department of Medicine, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Tsafi Pe'ery
- Department of Medicine, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Bin Tian
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA
| | - Michael B Mathews
- Department of Biochemistry & Molecular Biology, Rutgers New Jersey Medical School, Newark, NJ, USA.,Department of Medicine, Rutgers New Jersey Medical School, Newark, NJ, USA
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32
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Zhang J, Roggero VR, Allison LA. Nuclear Import and Export of the Thyroid Hormone Receptor. VITAMINS AND HORMONES 2017; 106:45-66. [PMID: 29407444 DOI: 10.1016/bs.vh.2017.04.002] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
The thyroid hormone receptors, TRα1 and TRβ1, are members of the nuclear receptor superfamily that forms one of the most abundant classes of transcription factors in multicellular organisms. Although primarily localized to the nucleus, TRα1 and TRβ1 shuttle rapidly between the nucleus and cytoplasm. The fine balance between nuclear import and export of TRs has emerged as a critical control point for modulating thyroid hormone-responsive gene expression. Mutagenesis studies have defined two nuclear localization signal (NLS) motifs that direct nuclear import of TRα1: NLS-1 in the hinge domain and NLS-2 in the N-terminal A/B domain. Three nuclear export signal (NES) motifs reside in the ligand-binding domain. A combined approach of shRNA-mediated knockdown and coimmunoprecipitation assays revealed that nuclear entry of TRα1 is facilitated by importin 7, likely through interactions with NLS-2, and importin β1 and the adapter importin α1 interacting with both NLS-1 and NLS-2. Interestingly, TRβ1 lacks NLS-2 and nuclear import depends solely on the importin α1/β1 heterodimer. Heterokaryon and fluorescence recovery after photobleaching shuttling assays identified multiple exportins that play a role in nuclear export of TRα1, including CRM1 (exportin 1), and exportins 4, 5, and 7. Even single amino acid changes in TRs dramatically alter their intracellular distribution patterns. We conclude that mutations within NLS and NES motifs affect nuclear shuttling activity, and propose that TR mislocalization contributes to the development of some types of cancer and Resistance to Thyroid Hormone syndrome.
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Affiliation(s)
- Jibo Zhang
- College of William and Mary, Williamsburg, VA, United States
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The elusive MAESTRO gene: Its human reproductive tissue-specific expression pattern. PLoS One 2017; 12:e0174873. [PMID: 28406912 PMCID: PMC5391009 DOI: 10.1371/journal.pone.0174873] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/18/2016] [Accepted: 03/16/2017] [Indexed: 12/16/2022] Open
Abstract
The encoded transcript of the Maestro—Male-specific Transcription in the developing Reproductive Organs (MRO) gene exhibits sexual dimorphic expression during murine gonadal development. The gene has no homology to any known gene and its expression pattern, protein function or structure are still unknown. Previously, studying gene expression in human ovarian cumulus cells, we found increased expression of MRO in lean-type Polycystic Ovarian Syndrome (PCOS) subjects, as compared to controls. In this study, we examined the MRO splice variants and protein expression pattern in various human tissues and cells. We found a differential expression pattern of the MRO 5’-UTR region in luteinized granulosa-cumulus cells and in testicular tissues as compared to non-gonadal tissues. Our study also shows a punctate nuclear expression pattern and disperse cytoplasmic expression pattern of the MRO protein in human granulosa-cumulus cells and in testicular germ cells, which was later validated by western blotting. The tentative and unique features of the protein hampered our efforts to gain more insight about this elusive protein. A better understanding of the tissue-specific MRO isoforms expression patterns and the unique structure of the protein may provide important insights into the function of this gene and possibly to the pathophysiology of PCOS.
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Mu J, Zhang Y, Hu Y, Hu X, Zhou Y, Zhao H, Pei R, Wu C, Chen J, Zhao H, Yang K, van Oers MM, Chen X, Wang Y. Autographa californica Multiple Nucleopolyhedrovirus Ac34 Protein Retains Cellular Actin-Related Protein 2/3 Complex in the Nucleus by Subversion of CRM1-Dependent Nuclear Export. PLoS Pathog 2016; 12:e1005994. [PMID: 27802336 PMCID: PMC5089780 DOI: 10.1371/journal.ppat.1005994] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2016] [Accepted: 10/12/2016] [Indexed: 12/21/2022] Open
Abstract
Actin, nucleation-promoting factors (NPFs), and the actin-related protein 2/3 complex (Arp2/3) are key elements of the cellular actin polymerization machinery. With nuclear actin polymerization implicated in ever-expanding biological processes and the discovery of the nuclear import mechanisms of actin and NPFs, determining Arp2/3 nucleo-cytoplasmic shuttling mechanism is important for understanding the function of nuclear actin. A unique feature of alphabaculovirus infection of insect cells is the robust nuclear accumulation of Arp2/3, which induces actin polymerization in the nucleus to assist in virus replication. We found that Ac34, a viral late gene product encoded by the alphabaculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV), is involved in Arp2/3 nuclear accumulation during virus infection. Further assays revealed that the subcellular distribution of Arp2/3 under steady-state conditions is controlled by chromosomal maintenance 1 (CRM1)-dependent nuclear export. Upon AcMNPV infection, Ac34 inhibits CRM1 pathway and leads to Arp2/3 retention in the nucleus.
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Affiliation(s)
- Jingfang Mu
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Yongli Zhang
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Yangyang Hu
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Xue Hu
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - Yuan Zhou
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - He Zhao
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - Rongjuan Pei
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - Chunchen Wu
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - Jizheng Chen
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - Han Zhao
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Kai Yang
- State Key Laboratory of Biocontrol, Sun Yat-sen University, Guangzhou, China
| | | | - Xinwen Chen
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
| | - Yun Wang
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
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Cáceres CJ, Angulo J, Contreras N, Pino K, Vera-Otarola J, López-Lastra M. Targeting deoxyhypusine hydroxylase activity impairs cap-independent translation initiation driven by the 5'untranslated region of the HIV-1, HTLV-1, and MMTV mRNAs. Antiviral Res 2016; 134:192-206. [PMID: 27633452 DOI: 10.1016/j.antiviral.2016.09.006] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2016] [Accepted: 09/12/2016] [Indexed: 12/14/2022]
Abstract
Replication of the human immunodeficiency virus type 1 (HIV-1) is dependent on eIF5A hypusination. Hypusine is formed post-translationally on the eIF5A precursor by two consecutive enzymatic steps; a reversible reaction involving the enzyme deoxyhypusine synthase (DHS) and an irreversible step involving the enzyme deoxyhypusine hydroxylase (DOHH). In this study we explored the effect of inhibiting DOHH activity and therefore eIF5A hypusination, on HIV-1 gene expression. Results show that the expression of proteins from an HIV-1 molecular clone is reduced when DOHH activity is inhibited by Deferiprone (DFP) or Ciclopirox (CPX). Next we evaluated the requirement of DOHH activity for internal ribosome entry site (IRES)-mediated translation initiation driven by the 5'untranslated region (5'UTR) of the full length HIV-1 mRNA. Results show that HIV-1 IRES activity relies on DOHH protein concentration and enzymatic activity. Similar results were obtained for IRES-dependent translation initiation mediated by 5'UTR of the human T-cell lymphotropic virus type 1 (HTLV-1) and the mouse mammary tumor virus (MMTV) mRNAs. Interestingly, activity of the poliovirus IRES, was less sensitive to the targeting of DOHH suggesting that not all viral IRESs are equally dependent on the cellular concentration or the activity of DOHH. In summary we present evidence indicating that the cellular concentration of DOHH and its enzymatic activity play a role in HIV-1, HTLV-1 and MMTV IRES-mediated translation initiation.
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Affiliation(s)
- C Joaquín Cáceres
- Laboratorio de Virología Molecular, Instituto Milenio de Inmunología e Inmunoterapia, Departamento de Enfermedades Infecciosas e Inmunología Pediátrica, Escuela de Medicina, Pontificia Universidad Católica de Chile, Marcoleta 391, Santiago, Chile
| | - Jenniffer Angulo
- Laboratorio de Virología Molecular, Instituto Milenio de Inmunología e Inmunoterapia, Departamento de Enfermedades Infecciosas e Inmunología Pediátrica, Escuela de Medicina, Pontificia Universidad Católica de Chile, Marcoleta 391, Santiago, Chile
| | - Nataly Contreras
- Laboratorio de Virología Molecular, Instituto Milenio de Inmunología e Inmunoterapia, Departamento de Enfermedades Infecciosas e Inmunología Pediátrica, Escuela de Medicina, Pontificia Universidad Católica de Chile, Marcoleta 391, Santiago, Chile
| | - Karla Pino
- Laboratorio de Virología Molecular, Instituto Milenio de Inmunología e Inmunoterapia, Departamento de Enfermedades Infecciosas e Inmunología Pediátrica, Escuela de Medicina, Pontificia Universidad Católica de Chile, Marcoleta 391, Santiago, Chile
| | - Jorge Vera-Otarola
- Laboratorio de Virología Molecular, Instituto Milenio de Inmunología e Inmunoterapia, Departamento de Enfermedades Infecciosas e Inmunología Pediátrica, Escuela de Medicina, Pontificia Universidad Católica de Chile, Marcoleta 391, Santiago, Chile
| | - Marcelo López-Lastra
- Laboratorio de Virología Molecular, Instituto Milenio de Inmunología e Inmunoterapia, Departamento de Enfermedades Infecciosas e Inmunología Pediátrica, Escuela de Medicina, Pontificia Universidad Católica de Chile, Marcoleta 391, Santiago, Chile.
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Khosravi S, Martinka M, Zhou Y, Ong CJ. Prognostic significance of the expression of nuclear eukaryotic translation initiation factor 5A2 in human melanoma. Oncol Lett 2016; 12:3089-3100. [PMID: 27899968 PMCID: PMC5103909 DOI: 10.3892/ol.2016.5057] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2015] [Accepted: 04/29/2016] [Indexed: 12/18/2022] Open
Abstract
Eukaryotic translation initiation factor 5A2 (EIF5A2) expression is upregulated in various cancers. The present authors previously demonstrated that cytoplasmic EIF5A2 expression increases with melanoma progression and inversely correlates with patient survival. Other studies have suggested that nuclear EIF5A2 may also play a role in oncogenesis. The present study used immunohistochemistry and tissue microarray with a large number of melanocytic lesions (n=459) and demonstrated that nuclear EIF5A2 expression was significantly upregulated between common acquired nevi, dysplastic nevi and primary melanomas, and between primary melanomas and metastatic melanomas. Nuclear EIF5A2 expression was inversely associated with overall and disease-specific 5-year survival rate for all (P<0.001) and primary (P=0.014 and P=0.015, respectively) melanoma patients. Nuclear EIF5A2 expression was directly associated with melanoma thickness (P=0.036) and American Joint Committee on Cancer staging (P<0.001), which suggests the possible role of nuclear EIF5A2 in melanoma cell invasion. Subsequently, the present study investigated the association between the expression of nuclear EIF5A2 and matrix metalloproteinase-2 (MMP-2), which is an important factor for promoting cancer cell invasion. Nuclear EIF5A2 and a strong MMP-2 expression were directly associated, and their concurrent expression was significantly associated with a poorer overall and disease-specific 5-year survival rate for all and primary melanoma patients. Nuclear and cytoplasmic EIF5A2 expression were also demonstrated to be significantly associated, and simultaneous expression of the two forms of EIF5A2 was significantly associated with poor overall and disease-specific 5-year survival rates for all and primary melanoma patients. Multivariate Cox regression analysis revealed that nuclear EIF5A2 expression alone and in combination with cytoplasmic EIF5A2 expression was an adverse independent prognostic factor for all and primary melanoma patients. In conclusion, the present study for the first time, to the best of our knowledge, demonstrated that nuclear EIF5A2 expression is an independent prognostic marker in melanoma, and revealed its role in melanoma progression and patient survival. Therefore, nuclear EIF5A2 may have the potential to serve as a therapeutic marker for melanoma.
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Affiliation(s)
- Shahram Khosravi
- Department of Dermatology and Skin Science, Vancouver Coastal Health Research Institute, University of British Columbia, Vancouver, BC V5Z 1L8, Canada; Department of Surgery, Vancouver Coastal Health Research Institute, University of British Columbia, Vancouver, BC V5Z 1L8, Canada
| | - Magdalena Martinka
- Department of Pathology, Vancouver Coastal Health Research Institute, University of British Columbia, Vancouver, BC V5Z 1L8, Canada
| | - Youwen Zhou
- Department of Dermatology and Skin Science, Vancouver Coastal Health Research Institute, University of British Columbia, Vancouver, BC V5Z 1L8, Canada
| | - Christopher J Ong
- Department of Surgery, Vancouver Coastal Health Research Institute, University of British Columbia, Vancouver, BC V5Z 1L8, Canada
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Zoller H, Tilg H. Nonalcoholic fatty liver disease and hepatocellular carcinoma. Metabolism 2016; 65:1151-60. [PMID: 26907206 DOI: 10.1016/j.metabol.2016.01.010] [Citation(s) in RCA: 132] [Impact Index Per Article: 14.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/21/2015] [Revised: 01/19/2016] [Accepted: 01/20/2016] [Indexed: 02/08/2023]
Abstract
The fastest growing cause of cancer-related death is hepatocellular carcinoma (HCC), which is at least partly attributable to the rising prevalence of non-alcoholic fatty liver disease. Non-alcoholic fatty liver disease (NAFLD) encompasses a broad spectrum of conditions, ranging from non-progressive bland steatosis to malignant transformation into hepatocellular cancer. The estimated annual HCC incidence in the progressive form of NAFLD - non-alcoholic steatohepatitis (NASH) - is about 0.3%. The risk of HCC development is higher in men and increases with age, more advanced fibrosis, progressive obesity, insulin resistance and diabetes mellitus. Studies on the molecular mechanism of HCC development in NAFLD have shown that hepatocarcinogenesis is associated with complex changes at the immunometabolic interface. In line with these clinical risk factors, administration of a choline-deficient high-fat diet to mice over a prolonged period results in spontaneous HCC development in a high percentage of animals. The role of altered insulin signaling in tumorigenesis is further supported by the observation that components of the insulin-signaling cascade are frequently mutated in hepatocellular cancer cells. These changes further enhance insulin-mediated growth and cell division of hepatocytes. Furthermore, studies investigating nuclear factor kappa B (NF-κB) signaling and HCC development allowed dissection of the complex links between inflammation and carcinogenesis. To conclude, NAFLD reflects an important risk factor for HCC, develops also in non-cirrhotic livers and is a prototypic cancer involving inflammatory and metabolic pathways. STRENGTHS/WEAKNESSES AND SUMMARY OF THE TRANSLATIONAL POTENTIAL OF THE MESSAGES IN THE PAPER: The systematic review summarizes findings from unbiased clinical and translational studies on hepatocellular cancer in non-alcoholic fatty liver disease. This provides a concise overview on the epidemiology, risk factors and molecular pathogenesis of the NAFL-NASH-HCC sequence. One limitation in the field is that few HCC studies stratify patients by underlying etiology, although the etiology of the underlying liver disease is an important co-determinant of clinical disease course and molecular pathogenesis. Molecular profiling of NAFL and associated HCC holds great translational potential for individualized surveillance, prevention and therapy.
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Affiliation(s)
- Heinz Zoller
- Department of Medicine II, Medical University of Innsbruck, Anichstrasse 35, A-6020 Innsbruck, Austria
| | - Herbert Tilg
- Department of Medicine I, Medical University of Innsbruck, Anichstrasse 35, A-6020 Innsbruck, Austria.
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38
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Mahipal A, Malafa M. Importins and exportins as therapeutic targets in cancer. Pharmacol Ther 2016; 164:135-43. [PMID: 27113410 DOI: 10.1016/j.pharmthera.2016.03.020] [Citation(s) in RCA: 121] [Impact Index Per Article: 13.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2015] [Accepted: 03/08/2016] [Indexed: 01/01/2023]
Abstract
The nuclear transport proteins, importins and exportins (karyopherin-β proteins), may play an important role in cancer by transporting key mediators of oncogenesis across the nuclear membrane in cancer cells. During nucleocytoplasmic transport of tumor suppressor proteins and cell cycle regulators during the processing of these proteins, aberrant cellular growth signaling and inactivation of apoptosis can occur, both critical to growth and development of tumors. Karyopherin-β proteins bind to these cargo proteins and RanGTP for active transport across the nuclear membrane through the nuclear pore complex. Importins and exportins are overexpressed in multiple tumors including melanoma, pancreatic, breast, colon, gastric, prostate, esophageal, lung cancer, and lymphomas. Furthermore, some of the karyopherin-β proteins such as exportin-1 have been implicated in drug resistance in cancer. Importin and exportin inhibitors are being considered as therapeutic targets against cancer and have shown preclinical anticancer activity. Moreover, synergistic activity has been observed with various chemotherapeutic and targeted agents. However, clinical development of the exportin-1 inhibitor leptomycin B was stopped due to adverse events, including vomiting, anorexia, and dehydration. Selinexor, a selective nuclear export inhibitor, is being tested in multiple clinical trials both as a single agent and in combination with chemotherapy. Selinexor has demonstrated clinical activity in multiple cancers, especially acute myelogenous leukemia and multiple myeloma. The roles of other importin and exportin inhibitors still need to be investigated clinically. Targeting the key mediators of nucleocytoplasmic transport in cancer cells represents a novel strategy in cancer intervention with the potential to significantly affect outcomes.
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Affiliation(s)
- Amit Mahipal
- Department of Oncology, Mayo Clinic, Rochester, MN 55905, United States
| | - Mokenge Malafa
- Department of Gastrointestinal Oncology, H. Lee Moffitt Cancer Center & Research Institute, Tampa, FL 33612, United States.
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39
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Aksu M, Trakhanov S, Görlich D. Structure of the exportin Xpo4 in complex with RanGTP and the hypusine-containing translation factor eIF5A. Nat Commun 2016; 7:11952. [PMID: 27306458 PMCID: PMC4912631 DOI: 10.1038/ncomms11952] [Citation(s) in RCA: 36] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2016] [Accepted: 05/16/2016] [Indexed: 12/28/2022] Open
Abstract
Xpo4 is a bidirectional nuclear transport receptor that mediates nuclear export of eIF5A and Smad3 as well as import of Sox2 and SRY. How Xpo4 recognizes such a variety of cargoes is as yet unknown. Here we present the crystal structure of the RanGTP·Xpo4·eIF5A export complex at 3.2 Å resolution. Xpo4 has a similar structure as CRM1, but the NES-binding site is occluded, and a new interaction site evolved that recognizes both globular domains of eIF5A. eIF5A contains hypusine, a unique amino acid with two positive charges, which is essential for cell viability and eIF5A function in translation. The hypusine docks into a deep, acidic pocket of Xpo4 and is thus a critical element of eIF5A's complex export signature. This further suggests that Xpo4 recognizes other cargoes differently, and illustrates how Xpo4 suppresses - in a chaperone-like manner - undesired interactions of eIF5A inside nuclei.
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Affiliation(s)
- Metin Aksu
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany
| | - Sergei Trakhanov
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany
| | - Dirk Görlich
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany
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Bhardwaj A, Das S. SIRT6 deacetylates PKM2 to suppress its nuclear localization and oncogenic functions. Proc Natl Acad Sci U S A 2016; 113:E538-47. [PMID: 26787900 PMCID: PMC4747762 DOI: 10.1073/pnas.1520045113] [Citation(s) in RCA: 120] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
SIRT6 (sirtuin 6) is a member of sirtuin family of deacetylases involved in diverse processes including genome stability, metabolic homeostasis, and tumorigenesis. However, the role of SIRT6 deacetylase activity in its tumor-suppressor functions is not well understood. Here we report that SIRT6 binds to and deacetylates nuclear PKM2 (pyruvate kinase M2) at the lysine 433 residue. PKM2 is a glycolytic enzyme with nonmetabolic nuclear oncogenic functions. SIRT6-mediated deacetylation results in PKM2 nuclear export. We further have identified exportin 4 as the specific transporter mediating PKM2 nuclear export. As a result of SIRT6-mediated deacetylation, PKM2 nuclear protein kinase and transcriptional coactivator functions are abolished. Thus, SIRT6 suppresses PKM2 oncogenic functions, resulting in reduced cell proliferation, migration potential, and invasiveness. Furthermore, studies in mouse tumor models demonstrate that PKM2 deacetylation is integral to SIRT6-mediated tumor suppression and inhibition of metastasis. Additionally, reduced SIRT6 levels correlate with elevated nuclear acetylated PKM2 levels in increasing grades of hepatocellular carcinoma. These findings provide key insights into the pivotal role of deacetylase activity in SIRT6 tumor-suppressor functions.
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Affiliation(s)
- Abhishek Bhardwaj
- Molecular Oncology Laboratory, National Institute of Immunology, New Delhi-110067, India
| | - Sanjeev Das
- Molecular Oncology Laboratory, National Institute of Immunology, New Delhi-110067, India
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Schröder M, Kolodzik A, Windshügel B, Krepstakies M, Priyadarshini P, Hartjen P, van Lunzen J, Rarey M, Hauber J, Meier C. Linker-Region Modified Derivatives of the Deoxyhypusine Synthase Inhibitor CNI-1493 Suppress HIV-1 Replication. Arch Pharm (Weinheim) 2016; 349:91-103. [PMID: 26725082 DOI: 10.1002/ardp.201500323] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2015] [Revised: 12/02/2015] [Accepted: 12/04/2015] [Indexed: 11/12/2022]
Abstract
The inhibition of cellular factors that are involved in viral replication may be an important alternative to the commonly used strategy of targeting viral enzymes. The guanylhydrazone CNI-1493, a potent inhibitor of the deoxyhypusine synthase (DHS), prevents the activation of the cellular factor eIF-5A and thereby suppresses HIV replication and a number of other diseases. Here, we report on the design, synthesis and biological evaluation of a series of CNI-1493 analogues. The sebacoyl linker in CNI-1493 was replaced by different alkyl or aryl dicarboxylic acids. Most of the tested derivatives suppress HIV-1 replication efficiently in a dose-dependent manner without showing toxic side effects. The unexpected antiviral activity of the rigid derivatives point to a second binding mode as previously assumed for CNI-1493. Moreover, the chemical stability of CNI-1493 was analysed, showing a successive hydrolysis of the imino bonds. By molecular dynamics simulations, the behaviour of the parent CNI-1493 in solution and its interactions with DHS were investigated.
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Affiliation(s)
- Marcus Schröder
- Institute of Organic Chemistry, Department of Chemistry, Faculty of Sciences, University of Hamburg, Hamburg, Germany
| | - Adrian Kolodzik
- ZBH Center for Bioinformatics, University of Hamburg, Hamburg, Germany
| | - Björn Windshügel
- ZBH Center for Bioinformatics, University of Hamburg, Hamburg, Germany
| | - Marcel Krepstakies
- Heinrich Pette Institute - Leibniz Institute for Experimental Virology, Hamburg, Germany
| | - Poornima Priyadarshini
- Heinrich Pette Institute - Leibniz Institute for Experimental Virology, Hamburg, Germany
| | - Philip Hartjen
- Department of Medicine, Infectious Diseases Unit, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Jan van Lunzen
- Department of Medicine, Infectious Diseases Unit, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Matthias Rarey
- ZBH Center for Bioinformatics, University of Hamburg, Hamburg, Germany
| | - Joachim Hauber
- Heinrich Pette Institute - Leibniz Institute for Experimental Virology, Hamburg, Germany
| | - Chris Meier
- Institute of Organic Chemistry, Department of Chemistry, Faculty of Sciences, University of Hamburg, Hamburg, Germany
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Kırlı K, Karaca S, Dehne HJ, Samwer M, Pan KT, Lenz C, Urlaub H, Görlich D. A deep proteomics perspective on CRM1-mediated nuclear export and nucleocytoplasmic partitioning. eLife 2015; 4:e11466. [PMID: 26673895 PMCID: PMC4764573 DOI: 10.7554/elife.11466] [Citation(s) in RCA: 165] [Impact Index Per Article: 16.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2015] [Accepted: 12/13/2015] [Indexed: 12/23/2022] Open
Abstract
CRM1 is a highly conserved, RanGTPase-driven exportin that carries proteins and RNPs from the nucleus to the cytoplasm. We now explored the cargo-spectrum of CRM1 in depth and identified surprisingly large numbers, namely >700 export substrates from the yeast S. cerevisiae, ≈1000 from Xenopus oocytes and >1050 from human cells. In addition, we quantified the partitioning of ≈5000 unique proteins between nucleus and cytoplasm of Xenopus oocytes. The data suggest new CRM1 functions in spatial control of vesicle coat-assembly, centrosomes, autophagy, peroxisome biogenesis, cytoskeleton, ribosome maturation, translation, mRNA degradation, and more generally in precluding a potentially detrimental action of cytoplasmic pathways within the nuclear interior. There are also numerous new instances where CRM1 appears to act in regulatory circuits. Altogether, our dataset allows unprecedented insights into the nucleocytoplasmic organisation of eukaryotic cells, into the contributions of an exceedingly promiscuous exportin and it provides a new basis for NES prediction.
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Affiliation(s)
- Koray Kırlı
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Samir Karaca
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Heinz Jürgen Dehne
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Matthias Samwer
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Kuan Ting Pan
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
| | - Christof Lenz
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
- Bioanalytics, Institute for Clinical Chemistry, University Medical Center Göttingen, Göttingen, Germany
| | - Henning Urlaub
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
- Bioanalytics, Institute for Clinical Chemistry, University Medical Center Göttingen, Göttingen, Germany
| | - Dirk Görlich
- Department of Cellular Logistics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
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Zain SM, Mohamed Z, Pirmohamed M, Tan HL, Alshawsh MA, Mahadeva S, Chan WK, Mustapha NRN, Mohamed R. Copy number variation in exportin-4 (XPO4) gene and its association with histological severity of non-alcoholic fatty liver disease. Sci Rep 2015; 5:13306. [PMID: 26293807 PMCID: PMC4543956 DOI: 10.1038/srep13306] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2015] [Accepted: 06/26/2015] [Indexed: 12/17/2022] Open
Abstract
A recent genome-wide copy number (CNV) scan identified a 13q12.11 duplication in the exportin-4 (XPO4) gene to be associated with non-alcoholic steatohepatitis (NASH). We sought to confirm the finding in a larger cohort and to assess the serum XPO4 pattern in a broad spectrum of non-alcoholic fatty liver disease (NAFLD) cases. We analysed 249 NAFLD patients and 232 matched controls using TaqMan assay and serum XPO4 was measured. Copy number distribution was as follows: copy number neutral (NAFLD: 53.8%, controls: 68.6%), copy number losses (NAFLD: 13.3%, controls: 12.9%), copy number gains (NAFLD: 32.9%, controls: 18.5%). CNV gain was significantly associated with a greater risk of NAFLD (adjusted OR 2.22, 95% CI 1.42–3.46, P = 0.0004) and NASH (adjusted OR 2.33, 95% CI 1.47–3.68, P = 0.0003). Interestingly, subjects carrying extra copy number showed significantly higher serum ALT and triglyceride (P < 0.05). Serum XPO4 levels progressively declined (P = 0.043) from controls (24.6 ng/mL) to simple steatosis (20.8 ng/mL) to NASH (13.8 ng/mL). In conclusion, XPO4 CNV duplication was associated with histological severity of NAFLD, and accompanied by changes in serum XPO4 levels providing insights into NAFLD pathogenesis, and has the potential for biomarker development.
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Affiliation(s)
- Shamsul Mohd Zain
- The Pharmacogenomics Laboratory, Department of Pharmacology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.,Department of Pharmacology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
| | - Zahurin Mohamed
- The Pharmacogenomics Laboratory, Department of Pharmacology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.,Department of Pharmacology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
| | - Munir Pirmohamed
- The Wolfson Centre for Personalised Medicine, University of Liverpool, Liverpool, United Kingdom
| | - Hwa Li Tan
- The Pharmacogenomics Laboratory, Department of Pharmacology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.,Department of Pharmacology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
| | | | - Sanjiv Mahadeva
- Department of Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
| | - Wah-Kheong Chan
- Department of Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
| | | | - Rosmawati Mohamed
- Department of Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
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44
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Ellis KL, Zhou Y, Beshansky JR, Ainehsazan E, Selker HP, Cupples LA, Huggins GS, Peter I. Genetic modifiers of response to glucose-insulin-potassium (GIK) infusion in acute coronary syndromes and associations with clinical outcomes in the IMMEDIATE trial. THE PHARMACOGENOMICS JOURNAL 2015; 15:488-95. [PMID: 25778467 DOI: 10.1038/tpj.2015.10] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/11/2014] [Revised: 12/17/2014] [Accepted: 01/28/2015] [Indexed: 11/09/2022]
Abstract
Modifiers of response to glucose, insulin and potassium (GIK) infusion may affect clinical outcomes in acute coronary syndromes (ACS). In an Immediate Myocardial Metabolic Enhancement During Initial Assessment And Treatment In Emergency Care (IMMEDIATE) trial's sub-study (n = 318), we explored effects of 132,634 genetic variants on plasma glucose and potassium response to 12-h GIK infusion. Associations between metabolite-associated variants and infarct size (n = 84) were assessed. The 'G' allele of rs12641551, near ACSL1, as well as the 'A' allele of XPO4 rs2585897 were associated with a differential glucose response (P for 2 degrees of freedom test, P2df ⩽ 4.75 × 10(-7)) and infarct size with GIK (P2df < 0.05). Variants within or near TAS1R3, LCA5, DNAH5, PTPRG, MAGI1, PTCSC3, STRADA, AKAP12, ARFGEF2, ADCYAP1, SETX, NDRG4 and ABCB11 modified glucose response, and near CSF1/AHCYL1 potassium response (P2df ⩽ 4.26 × 10(-7)), but not outcomes. Gene variants may modify glucose and potassium response to GIK infusion, contributing to cardiovascular outcomes in ACS.
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Affiliation(s)
- K L Ellis
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Y Zhou
- Department of Biostatistics, Boston University School of Public Health, Boston, MA, USA
| | - J R Beshansky
- Institute for Clinical Research and Health Policy Studies, Tufts Medical Center and Tufts University School of Medicine, Boston, MA, USA.,Regulatory and Clinical Research Management, Regis College, Weston, MA, USA
| | - E Ainehsazan
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - H P Selker
- Regulatory and Clinical Research Management, Regis College, Weston, MA, USA
| | - L A Cupples
- Department of Biostatistics, Boston University School of Public Health, Boston, MA, USA
| | - G S Huggins
- Molecular Cardiology Research Institute Center for Translational Genomics, Tufts Medical Center, Boston, MA, USA
| | - I Peter
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
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45
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Racca JD, Chen YS, Maloy JD, Wickramasinghe N, Phillips NB, Weiss MA. Structure-function relationships in human testis-determining factor SRY: an aromatic buttress underlies the specific DNA-bending surface of a high mobility group (HMG) box. J Biol Chem 2014; 289:32410-29. [PMID: 25258310 DOI: 10.1074/jbc.m114.597526] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Human testis determination is initiated by SRY, a Y-encoded architectural transcription factor. Mutations in SRY cause 46 XY gonadal dysgenesis with female somatic phenotype (Swyer syndrome) and confer a high risk of malignancy (gonadoblastoma). Such mutations cluster in the SRY high mobility group (HMG) box, a conserved motif of specific DNA binding and bending. To explore structure-function relationships, we constructed all possible substitutions at a site of clinical mutation (W70L). Our studies thus focused on a core aromatic residue (position 15 of the consensus HMG box) that is invariant among SRY-related HMG box transcription factors (the SOX family) and conserved as aromatic (Phe or Tyr) among other sequence-specific boxes. In a yeast one-hybrid system sensitive to specific SRY-DNA binding, the variant domains exhibited reduced (Phe and Tyr) or absent activity (the remaining 17 substitutions). Representative nonpolar variants with partial or absent activity (Tyr, Phe, Leu, and Ala in order of decreasing side-chain volume) were chosen for study in vitro and in mammalian cell culture. The clinical mutation (Leu) was found to markedly impair multiple biochemical and cellular activities as respectively probed through the following: (i) in vitro assays of specific DNA binding and protein stability, and (ii) cell culture-based assays of proteosomal degradation, nuclear import, enhancer DNA occupancy, and SRY-dependent transcriptional activation. Surprisingly, however, DNA bending is robust to this or the related Ala substitution that profoundly impairs box stability. Together, our findings demonstrate that the folding, trafficking, and gene-regulatory function of SRY requires an invariant aromatic "buttress" beneath its specific DNA-bending surface.
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Affiliation(s)
- Joseph D Racca
- From the Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106
| | - Yen-Shan Chen
- From the Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106
| | - James D Maloy
- From the Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106
| | - Nalinda Wickramasinghe
- From the Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106
| | - Nelson B Phillips
- From the Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106
| | - Michael A Weiss
- From the Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106
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46
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Sievert H, Pällmann N, Miller KK, Hermans-Borgmeyer I, Venz S, Sendoel A, Preukschas M, Schweizer M, Boettcher S, Janiesch PC, Streichert T, Walther R, Hengartner MO, Manz MG, Brümmendorf TH, Bokemeyer C, Braig M, Hauber J, Duncan KE, Balabanov S. A novel mouse model for inhibition of DOHH-mediated hypusine modification reveals a crucial function in embryonic development, proliferation and oncogenic transformation. Dis Model Mech 2014; 7:963-76. [PMID: 24832488 PMCID: PMC4107325 DOI: 10.1242/dmm.014449] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2013] [Accepted: 05/08/2014] [Indexed: 12/12/2022] Open
Abstract
The central importance of translational control by post-translational modification has spurred major interest in regulatory pathways that control translation. One such pathway uniquely adds hypusine to eukaryotic initiation factor 5A (eIF5A), and thereby affects protein synthesis and, subsequently, cellular proliferation through an unknown mechanism. Using a novel conditional knockout mouse model and a Caenorhabditis elegans knockout model, we found an evolutionarily conserved role for the DOHH-mediated second step of hypusine synthesis in early embryonic development. At the cellular level, we observed reduced proliferation and induction of senescence in 3T3 Dohh-/- cells as well as reduced capability for malignant transformation. Furthermore, mass spectrometry showed that deletion of DOHH results in an unexpected complete loss of hypusine modification. Our results provide new biological insight into the physiological roles of the second step of the hypusination of eIF5A. Moreover, the conditional mouse model presented here provides a powerful tool for manipulating hypusine modification in a temporal and spatial manner, to analyse both how this unique modification normally functions in vivo as well as how it contributes to different pathological conditions.
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Affiliation(s)
- Henning Sievert
- Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, Hubertus Wald-Tumor Zentrum, University Hospital Eppendorf, 20246 Hamburg, Germany
| | - Nora Pällmann
- Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, Hubertus Wald-Tumor Zentrum, University Hospital Eppendorf, 20246 Hamburg, Germany. Heinrich Pette Institute, Leibniz Institute for Experimental Virology, 20251 Hamburg, Germany
| | - Katharine K Miller
- Center for Molecular Neurobiology (ZMNH), University Medical Center Hamburg-Eppendorf (UKE), 20251 Hamburg, Germany
| | - Irm Hermans-Borgmeyer
- Center for Molecular Neurobiology (ZMNH), University Medical Center Hamburg-Eppendorf (UKE), 20251 Hamburg, Germany
| | - Simone Venz
- Department of Medical Biochemistry and Molecular Biology, University of Greifswald, 17475 Greifswald, Germany
| | - Ataman Sendoel
- Institute of Molecular Life Sciences, University of Zurich, 8057 Zurich, Switzerland. Division of Hematology, University Hospital Zurich, 8091 Zurich, Switzerland
| | - Michael Preukschas
- Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, Hubertus Wald-Tumor Zentrum, University Hospital Eppendorf, 20246 Hamburg, Germany
| | - Michaela Schweizer
- Center for Molecular Neurobiology (ZMNH), University Medical Center Hamburg-Eppendorf (UKE), 20251 Hamburg, Germany
| | - Steffen Boettcher
- Division of Hematology, University Hospital Zurich, 8091 Zurich, Switzerland
| | - P Christoph Janiesch
- Center for Molecular Neurobiology (ZMNH), University Medical Center Hamburg-Eppendorf (UKE), 20251 Hamburg, Germany
| | - Thomas Streichert
- Department of Clinical Chemistry, University Hospital of Cologne, 50924 Cologne, Germany
| | - Reinhard Walther
- Department of Medical Biochemistry and Molecular Biology, University of Greifswald, 17475 Greifswald, Germany
| | - Michael O Hengartner
- Institute of Molecular Life Sciences, University of Zurich, 8057 Zurich, Switzerland
| | - Markus G Manz
- Division of Hematology, University Hospital Zurich, 8091 Zurich, Switzerland
| | - Tim H Brümmendorf
- Clinic for Internal Medicine IV, Hematology and Oncology, University Hospital of the RWTH Aachen, 52074 Aachen, Germany
| | - Carsten Bokemeyer
- Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, Hubertus Wald-Tumor Zentrum, University Hospital Eppendorf, 20246 Hamburg, Germany
| | - Melanie Braig
- Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, Hubertus Wald-Tumor Zentrum, University Hospital Eppendorf, 20246 Hamburg, Germany
| | - Joachim Hauber
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, 20251 Hamburg, Germany
| | - Kent E Duncan
- Center for Molecular Neurobiology (ZMNH), University Medical Center Hamburg-Eppendorf (UKE), 20251 Hamburg, Germany
| | - Stefan Balabanov
- Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, Hubertus Wald-Tumor Zentrum, University Hospital Eppendorf, 20246 Hamburg, Germany. Division of Hematology, University Hospital Zurich, 8091 Zurich, Switzerland.
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47
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Mittal N, Morada M, Tripathi P, Gowri VS, Mandal S, Quirch A, Park MH, Yarlett N, Madhubala R. Cryptosporidium parvum has an active hypusine biosynthesis pathway. Mol Biochem Parasitol 2014; 195:14-22. [PMID: 24893338 PMCID: PMC4176827 DOI: 10.1016/j.molbiopara.2014.05.005] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/14/2013] [Revised: 05/21/2014] [Accepted: 05/25/2014] [Indexed: 11/25/2022]
Abstract
The protozoan parasite Cryptosporidium parvum causes severe enteric infection and diarrheal disease with substantial morbidity and mortality in untreated AIDS patients and children in developing or resource-limited countries. No fully effective treatment is available. Hypusination of eIF5A is an important post-translational modification essential for cell proliferation. This modification occurs in a two step process catalyzed by deoxyhypusine synthase (DHS) followed by deoxyhypusine hydroxylase. An ORF of 1086bp was identified in the C. parvum (Cp) genome which encodes for a putative polypeptide of 362 amino acids. The recombinant CpDHS protein was purified to homogeneity and used to probe the enzyme's mechanism, structure, and inhibition profile in a series of kinetic experiments. Sequence analysis and structural modeling of CpDHS were performed to probe differences with respect to the DHS of other species. Unlike Leishmania, Trypanosomes and Entamoeba, Cryptosporidium contains only a single gene for DHS. Phylogenetic analysis shows that CpDHS is more closely related to apicomplexan DHS than kinetoplastid DHS. Important residues that are essential for the functioning of the enzyme including NAD(+) binding residues, spermidine binding residues and the active site lysine are conserved between CpDHS and human DHS. N(1)-guanyl-1,7-diaminoheptane (GC7), a potent inhibitor of DHS caused an effective inhibition of infection and growth of C. parvum in HCT-8 cells.
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Affiliation(s)
- Nimisha Mittal
- School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India
| | - Marie Morada
- Haskins Laboratories, and the Department of Chemistry and Physical Sciences, Pace University, USA
| | - Pankaj Tripathi
- School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India
| | - V S Gowri
- School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India
| | - Swati Mandal
- Oral and Pharyngeal Cancer Branch, NIDCR, National Institute of Health, Bethesda, MD 20892-4340, USA
| | - Alison Quirch
- Haskins Laboratories, and the Department of Chemistry and Physical Sciences, Pace University, USA
| | - Myung Hee Park
- Oral and Pharyngeal Cancer Branch, NIDCR, National Institute of Health, Bethesda, MD 20892-4340, USA
| | - Nigel Yarlett
- Haskins Laboratories, and the Department of Chemistry and Physical Sciences, Pace University, USA
| | - Rentala Madhubala
- School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
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48
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Importins and exportins regulating allergic immune responses. Mediators Inflamm 2014; 2014:476357. [PMID: 24733961 PMCID: PMC3964845 DOI: 10.1155/2014/476357] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2013] [Revised: 01/18/2014] [Accepted: 01/21/2014] [Indexed: 12/21/2022] Open
Abstract
Nucleocytoplasmic shuttling of macromolecules is a well-controlled process involving importins and exportins. These karyopherins recognize and bind to receptor-mediated intracellular signals through specific signal sequences that are present on cargo proteins and transport into and out of the nucleus through nuclear pore complexes. Nuclear localization signals (NLS) present on cargo molecules to be imported while nuclear export signals (NES) on the molecules to be exported are recognized by importins and exportins, respectively. The classical NLS are found on many transcription factors and molecules that are involved in the pathogenesis of allergic diseases. In addition, several immune modulators, including corticosteroids and vitamin D, elicit their cellular responses by regulating the expression and activity of importin molecules. In this review article, we provide a comprehensive list of importin and exportin molecules and their specific cargo that shuttled between cytoplasm and the nucleus. We also critically review the role and regulation of specific importin and exportin involved in the transport of activated transcription factors in allergic diseases, the underlying molecular mechanisms, and the potential target sites for developing better therapeutic approaches.
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49
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Murine leukemia virus uses NXF1 for nuclear export of spliced and unspliced viral transcripts. J Virol 2014; 88:4069-82. [PMID: 24478440 DOI: 10.1128/jvi.03584-13] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
UNLABELLED Intron-containing mRNAs are subject to restricted nuclear export in higher eukaryotes. Retroviral replication requires the nucleocytoplasmic transport of both spliced and unspliced RNA transcripts, and RNA export mechanisms of gammaretroviruses are poorly characterized. Here, we report the involvement of the nuclear export receptor NXF1/TAP in the nuclear export of gammaretroviral RNA transcripts. We identified a conserved cis-acting element in the pol gene of gammaretroviruses, including murine leukemia virus (MLV) and xenotropic murine leukemia virus (XMRV), named the CAE (cytoplasmic accumulation element). The CAE enhanced the cytoplasmic accumulation of viral RNA transcripts and the expression of viral proteins without significantly affecting the stability, splicing, or translation efficiency of the transcripts. Insertion of the CAE sequence also facilitated Rev-independent HIV Gag expression. We found that the CAE sequence interacted with NXF1, whereas disruption of NXF1 ablated CAE function. Thus, the CAE sequence mediates the cytoplasmic accumulation of gammaretroviral transcripts in an NXF1-dependent manner. Disruption of NXF1 expression impaired cytoplasmic accumulations of both spliced and unspliced RNA transcripts of XMRV and MLV, resulting in their nuclear retention or degradation. Thus, our results demonstrate that gammaretroviruses use NXF1 for the cytoplasmic accumulation of both spliced and nonspliced viral RNA transcripts. IMPORTANCE Murine leukemia virus (MLV) has been studied as one of the classic models of retrovirology. Although unspliced host messenger RNAs are rarely exported from the nucleus, MLV actively exports unspliced viral RNAs to the cytoplasm. Despite extensive studies, how MLV achieves this difficult task has remained a mystery. Here, we have studied the RNA export mechanism of MLV and found that (i) the genome contains a sequence which supports the efficient nuclear export of viral RNAs, (ii) the cellular factor NXF1 is involved in the nuclear export of both spliced and unspliced viral RNAs, and, finally, (iii) depletion of NXF1 results in nuclear retention or degradation of viral RNAs. Our study provides a novel insight into MLV nuclear export.
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50
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Quintas-Granados LI, López-Camarillo C, Fandiño Armas J, Mendoza Hernandez G, Alvarez-Sánchez ME. Identification of the phosphorylated residues in TveIF5A by mass spectrometry. GENOMICS, PROTEOMICS & BIOINFORMATICS 2013; 11:378-384. [PMID: 24308916 PMCID: PMC4357829 DOI: 10.1016/j.gpb.2013.07.004] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/11/2013] [Revised: 06/27/2013] [Accepted: 07/08/2013] [Indexed: 11/16/2022]
Abstract
The initiation factor eIF5A in Trichomonas vaginalis (TveIF5A) is previously shown to undergo hypusination, phosphorylation and glycosylation. Three different pI isoforms of TveIF5A have been reported. The most acidic isoform (pI 5.2) corresponds to the precursor TveIF5A, whereas the mature TveIF5A appears to be the most basic isoform (pI 5.5). In addition, the intermediary isoform (pI 5.3) is found only under polyamine-depleted conditions and restored with exogenous putrescine. We propose that differences in PI are due to phosphorylation of the TveIF5A isoforms. Here, we have identified phosphorylation sites using mass spectrometry. The mature TveIF5A contains four phosphorylated residues (S3, T55, T78 and T82). Phosphorylation at S3 and T82 is also identified in the intermediary TveIF5A, while no phosphorylated residues are found in the precursor TveIF5A. It has been demonstrated that eIF5A proteins from plants and yeast are phosphorylated by a casein kinase 2 (CK2). Interestingly, a gene encoding a protein highly similar to CK2 (TvCK2) is found in T. vaginalis, which might be involved in the phosphorylation of TveIF5A in T. vaginalis.
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Affiliation(s)
| | - César López-Camarillo
- Genomic Sciences Postgraduate, Autonomous University of Mexico City (UACM), CP 03100 Mexico City, Mexico
| | - Jesús Fandiño Armas
- Genomic Sciences Postgraduate, Autonomous University of Mexico City (UACM), CP 03100 Mexico City, Mexico
| | - Guillermo Mendoza Hernandez
- Department of Biochemistry, Faculty of Medicine, National Autonomous University of Mexico, CP 04510 Mexico City, Mexico
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