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Ribeiro JC, Rodrigues BC, Bernardino RL, Alves MG, Oliveira PF. The interactome of cystic fibrosis transmembrane conductance regulator and its role in male fertility: A critical review. J Cell Physiol 2024; 239:e31422. [PMID: 39324358 DOI: 10.1002/jcp.31422] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2024] [Revised: 08/06/2024] [Accepted: 08/13/2024] [Indexed: 09/27/2024]
Abstract
The cystic fibrosis transmembrane conductance regulator (CFTR) is a cyclic adenosine monophosphate (cAMP)-regulated chloride and bicarbonate ion channel found in many human cells. Its unique biochemical characteristics and role as a member of the adenosine triphosphate (ATP)-binding cassette transporters superfamily are pivotal for the transport of several substrates across cellular membranes. CFTR is known to interact, physically and functionally, with several other cellular proteins. Hence, its properties are essential for moving various substances across cell membranes and ensuring correct cell functioning. Genetic mutations or environmental factors may disrupt CFTR's function resulting in different possible phenotypes due to gene variations that affect not only CFTR's function, localization, and processing within cells, but also those of its interactors. This has been reported as an underlying cause of various diseases, including cystic fibrosis. The severe clinical implications of cystic fibrosis have driven intense research into the role of CFTR in lung function but its significance to fertility, particularly in men, has been comparatively understudied. However, ongoing and more recent research into CFTR and its interacting proteins in the testis or specific testicular cells is beginning to shed light on this field. Herein, we provide a comprehensive and up-to-date overview of the CFTR, its interactome, and its crucial role in male reproduction, highlighting recent discoveries and advancements in understanding the molecular mechanisms involved. The comprehension of these complex interactions may pave the way for potential therapeutic approaches to improve fertility of men suffering from alterations in the function of CFTR.
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Grants
- This research was funded by "Fundação para a Ciência e a Tecnologia"-FCT to UMIB (UIDB/00215/2020, and UIDP/00215/2020), ITR-Laboratory for Integrative and Translational Research in Population Health (LA/P/0064/2020) and the post-graduation students João C. Ribeiro (UI/BD/150749/2020). The work was co-funded by FEDER through the COMPETE/QREN, FSE/POPH and POCI-COMPETE 2020 (POCI-01-0145-FEDER-007491) funds.
- Pedro F. Oliveira is funded by national funds through FCT-Fundação para a Ciência e a Tecnologia, I.P., under the Scientific Employment Stimulus-Institutional Call-reference CEEC-INST/00026/2018.
- This work also received support and help from FCT/MCTES to LAQV-REQUIMTE (LA/P/0008/202 - DOI 10.54499/LA/P/0008/2020; UIDP/50006/2020 - DOI 10.54499/UIDP/50006/2020; and UIDB/50006/2020 - DOI 10.54499/UIDB/50006/2020) and to iBiMed (UIDB/04501/2020 - DOI 10.54499/UIDB/04501/2020 and UIDP/04501/2020 - DOI 10.54499/UIDP/04501/2020), through national funds
- This research was funded by "Fundação para a Ciência e a Tecnologia"-FCT to UMIB (UIDB/00215/2020, and UIDP/00215/2020), ITR-Laboratory for Integrative and Translational Research in Population Health (LA/P/0064/2020) and the post-graduation students João C. Ribeiro (UI/BD/150749/2020). The work was co-funded by FEDER through the COMPETE/QREN, FSE/POPH and POCI-COMPETE 2020 (POCI-01-0145-FEDER-007491) funds. Pedro F. Oliveira is funded by national funds through FCT-Fundação para a Ciência e a Tecnologia, I.P., under the Scientific Employment Stimulus-Institutional Call-reference CEEC-INST/00026/2018. This work also received support and help from FCT/MCTES to LAQV-REQUIMTE (LA/P/0008/202 - DOI 10.54499/LA/P/0008/2020; UIDP/50006/2020 - DOI 10.54499/UIDP/50006/2020; and UIDB/50006/2020 - DOI 10.54499/UIDB/50006/2020) and to iBiMed (UIDB/04501/2020 - DOI 10.54499/UIDB/04501/2020 and UIDP/04501/2020 - DOI 10.54499/UIDP/04501/2020), through national funds
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Affiliation(s)
- João C Ribeiro
- Unit for Multidisciplinary Research in Biomedicine (UMIB), Institute of Biomedical Sciences Abel Salazar (ICBAS), University of Porto, Porto, Portugal
- Laboratory for Integrative and Translational Research in Population Health (ITR), University of Porto, Porto, Portugal
- LAQV-REQUIMTE and Department of Chemistry, University of Aveiro, Aveiro, Portugal
| | - Bernardo C Rodrigues
- LAQV-REQUIMTE and Department of Chemistry, University of Aveiro, Aveiro, Portugal
| | - Raquel L Bernardino
- Unit for Multidisciplinary Research in Biomedicine (UMIB), Institute of Biomedical Sciences Abel Salazar (ICBAS), University of Porto, Porto, Portugal
- Laboratory for Integrative and Translational Research in Population Health (ITR), University of Porto, Porto, Portugal
| | - Marco G Alves
- Department of Medical Sciences, Institute of Biomedicine (iBiMED), Aveiro, Portugal
| | - Pedro F Oliveira
- LAQV-REQUIMTE and Department of Chemistry, University of Aveiro, Aveiro, Portugal
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Lakli M, Onnée M, Carrez T, Becq F, Falguières T, Fanen P. ABC transporters involved in respiratory and cholestatic diseases: From rare to very rare monogenic diseases. Biochem Pharmacol 2024; 229:116468. [PMID: 39111603 DOI: 10.1016/j.bcp.2024.116468] [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: 02/26/2024] [Revised: 07/16/2024] [Accepted: 08/03/2024] [Indexed: 08/24/2024]
Abstract
ATP-binding cassette (ABC) transporters constitute a 49-member superfamily in humans. These proteins, most of them being transmembrane, allow the active transport of an important variety of substrates across biological membranes, using ATP hydrolysis as an energy source. For an important proportion of these ABC transporters, genetic variations of the loci encoding them have been correlated with rare genetic diseases, including cystic fibrosis and interstitial lung disease (variations in CFTR/ABCC7 and ABCA3) as well as cholestatic liver diseases (variations in ABCB4 and ABCB11). In this review, we first describe these ABC transporters and how their molecular dysfunction may lead to human diseases. Then, we propose a classification of the genetic variants according to their molecular defect (expression, traffic, function and/or stability), which may be considered as a general guideline for all ABC transporters' variants. Finally, we discuss recent progress in the field of targeted pharmacotherapy, which aim to correct specific molecular defects using small molecules. In conclusion, we are opening the path to treatment repurposing for diseases involving similar deficiencies in other ABC transporters.
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Affiliation(s)
- Mounia Lakli
- Inserm, Université Paris-Saclay, Physiopathogenèse et traitement des maladies du foie, UMR_S 1193, Hepatinov, 91400 Orsay, France
| | - Marion Onnée
- Univ Paris Est Creteil, INSERM, IMRB, F-94010, Créteil, France
| | - Thomas Carrez
- Université de Poitiers, Laboratoire Physiopathologie et Régulation des Transports Ioniques, Pôle Biologie Santé, 86000 Poitiers, France; ManRos Therapeutics, Hôtel de Recherche, Centre de Perharidy, 29680, Roscoff, France
| | - Frédéric Becq
- Université de Poitiers, Laboratoire Physiopathologie et Régulation des Transports Ioniques, Pôle Biologie Santé, 86000 Poitiers, France
| | - Thomas Falguières
- Inserm, Université Paris-Saclay, Physiopathogenèse et traitement des maladies du foie, UMR_S 1193, Hepatinov, 91400 Orsay, France
| | - Pascale Fanen
- Univ Paris Est Creteil, INSERM, IMRB, F-94010, Créteil, France; AP-HP, Département de Génétique Médicale, Hôpital Henri Mondor, F-94010, Créteil, France.
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Breimann S, Kamp F, Steiner H, Frishman D. AAontology: An Ontology of Amino Acid Scales for Interpretable Machine Learning. J Mol Biol 2024; 436:168717. [PMID: 39053689 DOI: 10.1016/j.jmb.2024.168717] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2024] [Revised: 07/15/2024] [Accepted: 07/19/2024] [Indexed: 07/27/2024]
Abstract
Amino acid scales are crucial for protein prediction tasks, many of them being curated in the AAindex database. Despite various clustering attempts to organize them and to better understand their relationships, these approaches lack the fine-grained classification necessary for satisfactory interpretability in many protein prediction problems. To address this issue, we developed AAontology-a two-level classification for 586 amino acid scales (mainly from AAindex) together with an in-depth analysis of their relations-using bag-of-word-based classification, clustering, and manual refinement over multiple iterations. AAontology organizes physicochemical scales into 8 categories and 67 subcategories, enhancing the interpretability of scale-based machine learning methods in protein bioinformatics. Thereby it enables researchers to gain a deeper biological insight. We anticipate that AAontology will be a building block to link amino acid properties with protein function and dysfunctions as well as aid informed decision-making in mutation analysis or protein drug design.
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Affiliation(s)
- Stephan Breimann
- Department of Bioinformatics, School of Life Sciences, Technical University of Munich, Freising, Germany; Ludwig-Maximilians-University Munich, Biomedical Center, Division of Metabolic Biochemistry, Munich, Germany; German Center for Neurodegenerative Diseases (DZNE), Munich, Germany
| | - Frits Kamp
- Ludwig-Maximilians-University Munich, Biomedical Center, Division of Metabolic Biochemistry, Munich, Germany
| | - Harald Steiner
- Ludwig-Maximilians-University Munich, Biomedical Center, Division of Metabolic Biochemistry, Munich, Germany; German Center for Neurodegenerative Diseases (DZNE), Munich, Germany
| | - Dmitrij Frishman
- Department of Bioinformatics, School of Life Sciences, Technical University of Munich, Freising, Germany.
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Dastoor P, Muiler C, Garrison A, Egan M, Carlos Dos Reis D, Santos A, Ameen NA. Localization and function of humanized F508del-CFTR in mouse intestine following activation of serum glucocorticoid kinase 1 and Trikafta. Eur J Pharmacol 2024; 978:176771. [PMID: 38925289 DOI: 10.1016/j.ejphar.2024.176771] [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: 03/04/2024] [Revised: 06/21/2024] [Accepted: 06/23/2024] [Indexed: 06/28/2024]
Abstract
The CFTR modulator Trikafta has markedly improved lung disease for Cystic Fibrosis (CF) patients carrying the common delta F508 (F508del-CFTR) CFTR mutation. F508del-CFTR results in an apical trafficking defect and loss of function in CFTR-expressing epithelial cells. However, Trikafta has not resulted in improved gastrointestinal function in CF patients. A humanized mouse model of F508del-CFTR was recently generated to evaluate CFTR modulators and other compounds to treat human F508del-CFTR CF intestinal disease. Short-term (4 h) treatment of rats with Dexamethasone (Dex) potently activates serum glucocorticoid kinase 1 (SGK1) and increases CFTR apical traffic and ion transport in the native intestine. This study examined CFTR localization and ion transport in intestinal segments from humanized F508del-CFTR mice following treatment with Dex in the presence/absence of Trikafta. Dex treatment improved apical CFTR localization and function but was inconsistent along intestinal segments. Combined treatment with Dex and Trikafta was superior to Dex alone but inconsistently improved CFTR localization and function. These data suggest further optimization of humanized CF mouse models will be necessary to test the efficacy of compounds to treat human CF intestinal disease.
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Affiliation(s)
- Parinaz Dastoor
- Department of Pediatrics/Gastroenterology and Hepatology, Yale School of Medicine, New Haven, CT, USA.
| | - Caroline Muiler
- Department of Pediatrics/Gastroenterology and Hepatology, Yale School of Medicine, New Haven, CT, USA
| | - Alannah Garrison
- Department of Pediatric Pulmonary Medicine, Yale School of Medicine, New Haven, CT, USA
| | - Marie Egan
- Department of Pediatric Pulmonary Medicine, Yale School of Medicine, New Haven, CT, USA; Department of Molecular Physiology, Yale School of Medicine, New Haven, CT, USA
| | - Diego Carlos Dos Reis
- Department of Pediatrics/Gastroenterology and Hepatology, Yale School of Medicine, New Haven, CT, USA.
| | - Anderson Santos
- Department of Genetics, Yale School of Medicine, New Haven, CT, USA
| | - Nadia A Ameen
- Department of Pediatrics/Gastroenterology and Hepatology, Yale School of Medicine, New Haven, CT, USA; Department of Molecular Physiology, Yale School of Medicine, New Haven, CT, USA.
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Abubakar-Waziri H, Edwards DA, Bhatta DB, Hull JH, Rudd M, Small P, Chung KF. Inhaled alkaline hypertonic divalent salts reduce refractory chronic cough frequency. ERJ Open Res 2024; 10:00241-2024. [PMID: 39377090 PMCID: PMC11456969 DOI: 10.1183/23120541.00241-2024] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2024] [Accepted: 05/13/2024] [Indexed: 10/09/2024] Open
Abstract
Background Treatment of chronic cough remains a challenge. We hypothesised that inhaled alkaline hypertonic divalent salts (alkaline HDS) might provide relief for refractory chronic cough by laryngeal and tracheal hydration. Methods We conducted an exploratory, single-blinded, nasal saline-controlled study in 12 refractory chronic cough patients to examine cough suppression efficacy of an alkaline HDS composition (SC001) at pH 8 or pH 9 administered by nasal inhalation. As control, we used nasal saline with the same hand-held pump spray aerosol device. Each subject was monitored continuously using a digital cough monitor watch for 1 week of baseline, 1 week of control treatment and 1 week of active treatment. Results Baseline daily cough rates ranged from 4 to 34 coughs·h-1 with mean visual analogue score 65±17 pre- and post-baseline testing. Control-adjusted efficacy of cough rate reduction ranged from 15% (p=0.015) (from Day 1) to 23% (p=0.002) (from Day 3). Control-adjusted efficacy was highest with SC001 pH 9 (n=5), ranging from 25% (p=0.03) (from Day 1) to 35% (p=0.02) (from Day 3), and lowest for SC001 pH 8 (n=7), ranging from 9% (p=0.08) (from Day 1) to 16% (p=0.02) (from Day 3). Hourly cough counts and visual analogue score correlated for baseline (r=0.254, p=0.02) and control (r=0.299, p=0.007) monitoring weeks. Treatment improved this correlation (r=0.434, p=0.00006). No adverse events were reported. Conclusions Alkaline (pH 9) HDS aerosol is a promising treatment for refractory chronic cough and should be further evaluated.
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Affiliation(s)
- Hisham Abubakar-Waziri
- National Heart and Lung Institute, Imperial College London, and Royal Brompton and Harefield Hospital, London, UK
| | - David A. Edwards
- Center for Nanomedicine, Johns Hopkins University Medical School, Baltimore, MD, USA
- John Paulson School of Engineering and Applied Sciences, Harvard University, MA, USA
- Sensory Cloud Inc., Boston, MA, USA
| | | | - James H. Hull
- National Heart and Lung Institute, Imperial College London, and Royal Brompton and Harefield Hospital, London, UK
| | - Matthew Rudd
- Hyfe Inc., Wilmington, DE, USA
- Department of Mathematics and Computer Science, The University of the South, Seewanee, TN, USA
| | - Peter Small
- Hyfe Inc., Wilmington, DE, USA
- Department of Global Health, University Washington, Seattle, WA, USA
| | - Kian Fan Chung
- National Heart and Lung Institute, Imperial College London, and Royal Brompton and Harefield Hospital, London, UK
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Thakur S, Ankita, Dash S, Verma R, Kaur C, Kumar R, Mazumder A, Singh G. Understanding CFTR Functionality: A Comprehensive Review of Tests and Modulator Therapy in Cystic Fibrosis. Cell Biochem Biophys 2024; 82:15-34. [PMID: 38048024 DOI: 10.1007/s12013-023-01200-w] [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: 10/11/2023] [Accepted: 11/13/2023] [Indexed: 12/05/2023]
Abstract
Cystic fibrosis is a genetic disorder inherited in an autosomal recessive manner. It is caused by a mutation in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene on chromosome 7, which leads to abnormal regulation of chloride and bicarbonate ions in cells that line organs like the lungs and pancreas. The CFTR protein plays a crucial role in regulating chloride ion flow, and its absence or malfunction causes the production of thick mucus that affects several organs. There are more than 2000 identified mutations that are classified into seven categories based on their dysfunction mechanisms. In this article, we have conducted a thorough examination and consolidation of the diverse array of tests essential for the quantification of CFTR functionality. Furthermore, we have engaged in a comprehensive discourse regarding the recent advancements in CFTR modulator therapy, a pivotal approach utilized for the management of cystic fibrosis, alongside its concomitant relevance in evaluating CFTR functionality.
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Affiliation(s)
- Shorya Thakur
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India
| | - Ankita
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India
| | - Shubham Dash
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India
| | - Rupali Verma
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India
| | - Charanjit Kaur
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India
| | - Rajesh Kumar
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India
| | - Avijit Mazumder
- Noida Institute of Engineering and Technology (Pharmacy Institute), Greater Noida, UP, India
| | - Gurvinder Singh
- School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Panjab, India.
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El Makhzen N, Daimi H, Bouguenouch L, Abriel H. The burden of cystic fibrosis in North Africa. Front Genet 2024; 14:1295008. [PMID: 38269366 PMCID: PMC10806102 DOI: 10.3389/fgene.2023.1295008] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2023] [Accepted: 12/26/2023] [Indexed: 01/26/2024] Open
Abstract
Background: Over 200 pathogenic variants in the cystic fibrosis transmembrane conductance regulator (CFTR) gene are associated with cystic fibrosis (CF)-the most prevalent autosomal recessive disease globally, the p.Phe508del variant being the most commonly observed. Main text: Recent epidemiological studies suggest a higher global prevalence of CF than previously thought. Nevertheless, comprehensive CF data remains extremely scarce among African populations, contributing to a significant information gap within the African healthcare system. Consequently, the underestimation of CF among children from African populations is likely. The goal of this article is to review the pathogenesis of CF and its prevalence in the countries of North Africa. Conclusion: The prevalence of CF in North African countries is likely underestimated due to the complexity of the disease and the lack of a timely, proper clinical and genetic investigation that allows the early identification of CF patients and thus facilitates therapeutic recommendations. Therefore, specific genetic and epidemiological studies on African individuals showing CF symptoms should be conducted to enhance the diagnostic yield of CF in Africa.
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Affiliation(s)
- Nada El Makhzen
- Ion Channels and Channelopathies Laboratory, Institute for Biochemistry and Molecular Medicine, University of Bern, Bern, Switzerland
| | - Houria Daimi
- Laboratory of Human Genome and Multifactorial Diseases (LR12ES07), Faculty of Pharmacy, University of Monastir, Monastir, Tunisia
- Department of Biology, Faculty of Sciences, University of Gabes, Gabès, Tunisia
| | - Laila Bouguenouch
- Laboratory of Medical Genetics and Oncogenetics, University Hospital Hassan II, Sidi Mohamed Ben Abdellah University, Fez, Morocco
| | - Hugues Abriel
- Ion Channels and Channelopathies Laboratory, Institute for Biochemistry and Molecular Medicine, University of Bern, Bern, Switzerland
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Novischi SYP, Karoly-Lakatos A, Chok K, Bonifer C, Becker-Baldus J, Glaubitz C. Probing the allosteric NBD-TMD crosstalk in the ABC transporter MsbA by solid-state NMR. Commun Biol 2024; 7:43. [PMID: 38182790 PMCID: PMC10770068 DOI: 10.1038/s42003-023-05617-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2023] [Accepted: 11/21/2023] [Indexed: 01/07/2024] Open
Abstract
The ABC transporter MsbA plays a critical role in Gram-negative bacteria in the regulation of the outer membrane by translocating core-LPS across the inner membrane. Additionally, a broad substrate specificity for lipophilic drugs has been shown. The allosteric interplay between substrate binding in the transmembrane domains and ATP binding and turnover in the nucleotide-binding domains must be mediated via the NBD/TMD interface. Previous studies suggested the involvement of two intracellular loops called coupling helix 1 and 2 (CH1, CH2). Here, we demonstrate by solid-state NMR spectroscopy that substantial chemical shift changes within both CH1 and CH2 occur upon substrate binding, in the ATP hydrolysis transition state, and upon inhibitor binding. CH2 is domain-swapped within the MsbA structure, and it is noteworthy that substrate binding induces a larger response in CH2 compared to CH1. Our data demonstrate that CH1 and CH2 undergo structural changes as part of the TMD-NBD cross-talk.
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Affiliation(s)
- S Y Phoebe Novischi
- Institute for Biophysical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max von Laue Str. 9, 60438, Frankfurt, Germany
| | - Andrea Karoly-Lakatos
- Institute for Biophysical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max von Laue Str. 9, 60438, Frankfurt, Germany
| | - Kerby Chok
- Institute for Biophysical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max von Laue Str. 9, 60438, Frankfurt, Germany
| | - Christian Bonifer
- Institute for Biophysical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max von Laue Str. 9, 60438, Frankfurt, Germany
| | - Johanna Becker-Baldus
- Institute for Biophysical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max von Laue Str. 9, 60438, Frankfurt, Germany
| | - Clemens Glaubitz
- Institute for Biophysical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max von Laue Str. 9, 60438, Frankfurt, Germany.
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Yeh HI, Sutcliffe KJ, Sheppard DN, Hwang TC. CFTR Modulators: From Mechanism to Targeted Therapeutics. Handb Exp Pharmacol 2024; 283:219-247. [PMID: 35972584 DOI: 10.1007/164_2022_597] [Citation(s) in RCA: 4] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/15/2023]
Abstract
People with cystic fibrosis (CF) suffer from a multi-organ disorder caused by loss-of-function variants in the gene encoding the epithelial anion channel cystic fibrosis transmembrane conductance regulator (CFTR). Tremendous progress has been made in both basic and clinical sciences over the past three decades since the identification of the CFTR gene. Over 90% of people with CF now have access to therapies targeting dysfunctional CFTR. This success was made possible by numerous studies in the field that incrementally paved the way for the development of small molecules known as CFTR modulators. The advent of CFTR modulators transformed this life-threatening illness into a treatable disease by directly binding to the CFTR protein and correcting defects induced by pathogenic variants. In this chapter, we trace the trajectory of structural and functional studies that brought CF therapies from bench to bedside, with an emphasis on mechanistic understanding of CFTR modulators.
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Affiliation(s)
- Han-I Yeh
- Department of Pharmacology, National Yang Ming Chiao Tung University, Taipei City, Taiwan
- Department of Medical Pharmacology and Physiology, University of Missouri, Columbia, MO, USA
- Dalton Cardiovascular Research Center, University of Missouri, Columbia, MO, USA
| | - Katy J Sutcliffe
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, UK
| | - David N Sheppard
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, UK
| | - Tzyh-Chang Hwang
- Department of Pharmacology, National Yang Ming Chiao Tung University, Taipei City, Taiwan.
- Department of Medical Pharmacology and Physiology, University of Missouri, Columbia, MO, USA.
- Dalton Cardiovascular Research Center, University of Missouri, Columbia, MO, USA.
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Baharara H, Kesharwani P, Johnston TP, Sahebkar A. Therapeutic potential of phytochemicals for cystic fibrosis. Biofactors 2023; 49:984-1009. [PMID: 37191383 DOI: 10.1002/biof.1960] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/14/2022] [Accepted: 05/01/2023] [Indexed: 05/17/2023]
Abstract
The aim of this review was to review and discuss various phytochemicals that exhibit beneficial effects on mutated membrane channels, and hence, improve transmembrane conductance. These therapeutic phytochemicals may have the potential to decrease mortality and morbidity of CF patients. Four databases were searched using keywords. Relevant studies were identified, and related articles were separated. Google Scholar, as well as gray literature (i.e., information that is not produced by commercial publishers), were also checked for related articles to locate/identify additional studies. The relevant databases were searched a second time to ensure that recent studies were included. In conclusion, while curcumin, genistein, and resveratrol have demonstrated effectiveness in this regard, it should be emphasized that coumarins, quercetin, and other herbal medicines also have beneficial effects on transporter function, transmembrane conductivity, and overall channel activity. Additional in vitro and in vivo studies should be conducted on mutant CFTR to unequivocally define the mechanism by which phytochemicals alter transmembrane channel function/activity, since the results of the studies evaluated in this review have a high degree of heterogenicity and discrepancy. Finally, continued research be undertaken to clearly define the mechanism(s) of action and the therapeutic effects that therapeutic phytochemicals have on the symptoms observed in CF patients in an effort to reduce mortality and morbidity.
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Affiliation(s)
- Hamed Baharara
- Department of Clinical Pharmacy, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Prashant Kesharwani
- Department of Pharmaceutics, School of Pharmaceutical Education and Research, Jamia Hamdard, New Delhi, India
- Center for Transdisciplinary Research, Department of Pharmacology, Saveetha Dental College, Saveetha Institute of Medical and Technical Science, Chennai, India
| | - Thomas P Johnston
- Division of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Missouri-Kansas City, Kansas City, Missouri, USA
| | - AmirHossein Sahebkar
- Biotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran
- Applied Biomedical Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
- Department of Biotechnology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran
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Rumpf M, Pautz S, Drebes B, Herberg FW, Müller HAJ. Microtubule-Associated Serine/Threonine (MAST) Kinases in Development and Disease. Int J Mol Sci 2023; 24:11913. [PMID: 37569286 PMCID: PMC10419289 DOI: 10.3390/ijms241511913] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2023] [Revised: 07/17/2023] [Accepted: 07/19/2023] [Indexed: 08/13/2023] Open
Abstract
Microtubule-Associated Serine/Threonine (MAST) kinases represent an evolutionary conserved branch of the AGC protein kinase superfamily in the kinome. Since the discovery of the founding member, MAST2, in 1993, three additional family members have been identified in mammals and found to be broadly expressed across various tissues, including the brain, heart, lung, liver, intestine and kidney. The study of MAST kinases is highly relevant for unraveling the molecular basis of a wide range of different human diseases, including breast and liver cancer, myeloma, inflammatory bowel disease, cystic fibrosis and various neuronal disorders. Despite several reports on potential substrates and binding partners of MAST kinases, the molecular mechanisms that would explain their involvement in human diseases remain rather obscure. This review will summarize data on the structure, biochemistry and cell and molecular biology of MAST kinases in the context of biomedical research as well as organismal model systems in order to provide a current profile of this field.
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Affiliation(s)
- Marie Rumpf
- Department of Developmental Genetics, Institute of Biology, University of Kassel, 34321 Kassel, Germany; (M.R.)
| | - Sabine Pautz
- Department of Biochemistry, Institute of Biology, University of Kassel, 34321 Kassel, Germany
| | - Benedikt Drebes
- Department of Developmental Genetics, Institute of Biology, University of Kassel, 34321 Kassel, Germany; (M.R.)
| | - Friedrich W. Herberg
- Department of Biochemistry, Institute of Biology, University of Kassel, 34321 Kassel, Germany
| | - Hans-Arno J. Müller
- Department of Developmental Genetics, Institute of Biology, University of Kassel, 34321 Kassel, Germany; (M.R.)
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12
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Barros P, Matos AM, Matos P, Jordan P. YES1 Kinase Mediates the Membrane Removal of Rescued F508del-CFTR in Airway Cells by Promoting MAPK Pathway Activation via SHC1. Biomolecules 2023; 13:949. [PMID: 37371529 DOI: 10.3390/biom13060949] [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: 05/04/2023] [Revised: 05/26/2023] [Accepted: 06/01/2023] [Indexed: 06/29/2023] Open
Abstract
Recent developments in CFTR modulator drugs have had a significant transformational effect on the treatment of individuals with Cystic Fibrosis (CF) who carry the most frequent F508del-CFTR mutation in at least one allele. However, the clinical effects of these revolutionary drugs remain limited by their inability to fully restore the plasma membrane (PM) stability of the rescued mutant channels. Here, we shed new light on the molecular mechanisms behind the reduced half-life of rescued F508del-CFTR at the PM of airway cells. We describe that YES1 protein kinase is enriched in F508del-CFTR protein PM complexes, and that its interaction with rescued channels is mediated and dependent on the adaptor protein YAP1. Moreover, we show that interference with this complex, either by depletion of one of these components or inhibiting YES1 activity, is sufficient to significantly improve the abundance and stability of modulator-rescued F508del-CFTR at the surface of airway cells. In addition, we found that this effect was mediated by a decreased phosphorylation of the scaffold protein SHC1, a key regulator of MAPK pathway activity. In fact, we showed that depletion of SHC1 or inhibition of MAPK pathway signaling was sufficient to improve rescued F508del-CFTR surface levels, whereas an ectopic increase in pathway activation downstream of SHC1, through the use of a constitutively active H-RAS protein, abrogated the stabilizing effect of YES1 inhibition on rescued F508del-CFTR. Taken together, our findings not only provide new mechanistic insights into the regulation of modulator-rescued F508del-CFTR membrane stability, but also open exciting new avenues to be further explored in CF research and treatment.
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Affiliation(s)
- Patrícia Barros
- Departamento de Genética Humana, Instituto Nacional de Saúde Doutor Ricardo Jorge, 1649-016 Lisboa, Portugal
- BioISI-Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
| | - Ana M Matos
- Departamento de Genética Humana, Instituto Nacional de Saúde Doutor Ricardo Jorge, 1649-016 Lisboa, Portugal
- BioISI-Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
| | - Paulo Matos
- Departamento de Genética Humana, Instituto Nacional de Saúde Doutor Ricardo Jorge, 1649-016 Lisboa, Portugal
- BioISI-Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
| | - Peter Jordan
- Departamento de Genética Humana, Instituto Nacional de Saúde Doutor Ricardo Jorge, 1649-016 Lisboa, Portugal
- BioISI-Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
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13
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Yang J, Huang T, Zhang J, Bai G, Wang W, Yao J, Chen Z, Tu C. Sulphur dioxide and fluoride co-exposure cause enamel damage by disrupting the Cl -/HCO 3- ion transport. J Trace Elem Med Biol 2023; 77:127131. [PMID: 36630759 DOI: 10.1016/j.jtemb.2023.127131] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Revised: 11/21/2022] [Accepted: 01/04/2023] [Indexed: 01/07/2023]
Abstract
OBJECTIVE Although there is growing evidence linking the exposure to sulphur dioxide (SO2) and fluoride to human diseases, there is little data on the co-exposure of SO2 and fluoride. Moreover, literature on SO2 and fluoride co-exposure to enamel damage is insufficient. In this work, we concentrate on the concurrent environmental issues of excessive SO2 and fluoride in several coal-consuming regions. METHOD To identify the toxicity of SO2 and fluoride exposure either separately or together, we used both ICR mice and LS8 cells, and factorial design was employed to assess the type of potential combined action. RESULT In this study, co-exposure to SO2 and fluoride exacerbated enamel damage, resulting in more severe enamel defects of incisor and the damage occurred earlier. Cl-/HCO3- exchanger expression is increased by SO2 and fluoride in mouse incisor. Consistent with in vivo results, co-exposure of SO2 and fluoride decreased pHi and increased [Cl-]i level by increasing the expression of the Cl-/HCO3- exchanger in LS8 cells. Furthermore, SO2 and F may increase merlin protein expression, and merlin deficiency causes AE2 expression to decrease in vitro. CONCLUSION Overall, these results indicate that co-exposure to SO2 and fluoride may result in more toxicity both in vitro and in vivo than a single exposure to SO2 and fluoride, suggesting that residents in areas contaminated with SO2 and fluoride may be more likely to suffer enamel damage.
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Affiliation(s)
- Junlin Yang
- School of Public Health, Guizhou Medical University, Guian New Region, China; The Affiliated Hospital of Guizhou Medical University, Guiyang, China
| | - Tongtong Huang
- School of Public Health, Guizhou Medical University, Guian New Region, China
| | - Jianghui Zhang
- School of Public Health, Guizhou Medical University, Guian New Region, China
| | - Guohui Bai
- Key Laboratory of Oral Disease Research, School of Stomatology, Zunyi Medical University, Zunyi, China
| | - Wentai Wang
- School of Public Health, Guizhou Medical University, Guian New Region, China
| | - Jie Yao
- School of Public Health, Guizhou Medical University, Guian New Region, China
| | - Zheng Chen
- School of Public Health, Guizhou Medical University, Guian New Region, China
| | - Chenglong Tu
- Key Laboratory of Environmental Pollution Monitoring and Disease Control, Ministry of Education, Guizhou Medical University, Guian New Region, China; Toxicity Testing Center of Guizhou Medical University, Guiyang, China.
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14
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Tahti EF, Blount JM, Jackson SN, Gao M, Gill NP, Smith SN, Pederson NJ, Rumph SN, Struyvenberg SA, Mackley IGP, Madden DR, Amacher JF. Additive energetic contributions of multiple peptide positions determine the relative promiscuity of viral and human sequences for PDZ domain targets. Protein Sci 2023; 32:e4611. [PMID: 36851847 PMCID: PMC10022582 DOI: 10.1002/pro.4611] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2022] [Revised: 02/13/2023] [Accepted: 02/23/2023] [Indexed: 03/01/2023]
Abstract
Protein-protein interactions that involve recognition of short peptides are critical in cellular processes. Protein-peptide interaction surface areas are relatively small and shallow, and there are often overlapping specificities in families of peptide-binding domains. Therefore, dissecting selectivity determinants can be challenging. PDZ domains are a family of peptide-binding domains located in several intracellular signaling and trafficking pathways. These domains are also directly targeted by pathogens, and a hallmark of many oncogenic viral proteins is a PDZ-binding motif. However, amidst sequences that target PDZ domains, there is a wide spectrum in relative promiscuity. For example, the viral HPV16 E6 oncoprotein recognizes over double the number of PDZ domain-containing proteins as the cystic fibrosis transmembrane conductance regulator (CFTR) in the cell, despite similar PDZ targeting-sequences and identical motif residues. Here, we determine binding affinities for PDZ domains known to bind either HPV16 E6 alone or both CFTR and HPV16 E6, using peptides matching WT and hybrid sequences. We also use energy minimization to model PDZ-peptide complexes and use sequence analyses to investigate this difference. We find that while the majority of single mutations had marginal effects on overall affinity, the additive effect on the free energy of binding accurately describes the selectivity observed. Taken together, our results describe how complex and differing PDZ interactomes can be programmed in the cell.
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Affiliation(s)
- Elise F. Tahti
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | - Jadon M. Blount
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | - Sophie N. Jackson
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | - Melody Gao
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | - Nicholas P. Gill
- Department of BiochemistryGeisel School of Medicine at DartmouthHanoverNew HampshireUSA
| | - Sarah N. Smith
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | - Nick J. Pederson
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | | | | | - Iain G. P. Mackley
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
| | - Dean R. Madden
- Department of BiochemistryGeisel School of Medicine at DartmouthHanoverNew HampshireUSA
| | - Jeanine F. Amacher
- Department of ChemistryWestern Washington UniversityBellinghamWashingtonUSA
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15
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Tahti EF, Blount JM, Jackson SN, Gao M, Gill NP, Smith SN, Pederson NJ, Rumph SN, Struyvenberg SA, Mackley IGP, Madden DR, Amacher JF. Additive energetic contributions of multiple peptide positions determine the relative promiscuity of viral and human sequences for PDZ domain targets. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2022.12.31.522388. [PMID: 36711692 PMCID: PMC9881875 DOI: 10.1101/2022.12.31.522388] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 06/18/2023]
Abstract
Protein-protein interactions that include recognition of short sequences of amino acids, or peptides, are critical in cellular processes. Protein-peptide interaction surface areas are relatively small and shallow, and there are often overlapping specificities in families of peptide-binding domains. Therefore, dissecting selectivity determinants can be challenging. PDZ domains are an example of a peptide-binding domain located in several intracellular signaling and trafficking pathways, which form interactions critical for the regulation of receptor endocytic trafficking, tight junction formation, organization of supramolecular complexes in neurons, and other biological systems. These domains are also directly targeted by pathogens, and a hallmark of many oncogenic viral proteins is a PDZ-binding motif. However, amidst sequences that target PDZ domains, there is a wide spectrum in relative promiscuity. For example, the viral HPV16 E6 oncoprotein recognizes over double the number of PDZ domain-containing proteins as the cystic fibrosis transmembrane conductance regulator (CFTR) in the cell, despite similar PDZ targeting-sequences and identical motif residues. Here, we determine binding affinities for PDZ domains known to bind either HPV16 E6 alone or both CFTR and HPV16 E6, using peptides matching WT and hybrid sequences. We also use energy minimization to model PDZ-peptide complexes and use sequence analyses to investigate this difference. We find that while the majority of single mutations had a marginal effect on overall affinity, the additive effect on the free energy of binding accurately describes the selectivity observed. Taken together, our results describe how complex and differing PDZ interactomes can be programmed in the cell.
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Affiliation(s)
- Elise F. Tahti
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Jadon M. Blount
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Sophie N. Jackson
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Melody Gao
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Nicholas P. Gill
- Department of Biochemistry, Geisel School of Medicine at Dartmouth, Hanover, NH, USA
| | - Sarah N. Smith
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Nick J. Pederson
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Simone N. Rumph
- Department of Biochemistry, Bowdoin College, Brunswick, ME, USA
| | | | - Iain G. P. Mackley
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
| | - Dean R. Madden
- Department of Biochemistry, Geisel School of Medicine at Dartmouth, Hanover, NH, USA
| | - Jeanine F. Amacher
- Department of Chemistry, Western Washington University, Bellingham, WA, USA
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Caverly LJ, Riquelme SA, Hisert KB. The Impact of Highly Effective Modulator Therapy on Cystic Fibrosis Microbiology and Inflammation. Clin Chest Med 2022; 43:647-665. [PMID: 36344072 PMCID: PMC10224747 DOI: 10.1016/j.ccm.2022.06.007] [Citation(s) in RCA: 22] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022]
Abstract
Highly effective cystic fibrosis (CF) transmembrane conductance regulator (CFTR) modulator therapy (HEMT) corrects the underlying molecular defect causing CF disease. HEMT decreases symptom burden and improves clinical metrics and quality of life for most people with CF (PwCF) and eligible cftr mutations. Improvements in measures of pulmonary health suggest that restoration of function of defective CFTR anion channels by HEMT not only enhances airway mucociliary clearance, but also reduces chronic pulmonary infection and inflammation. This article reviews the evidence for how HEMT influences the dynamic and interdependent processes of infection and inflammation in the CF airway, and what questions remain unanswered.
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Affiliation(s)
- Lindsay J Caverly
- Department of Pediatrics, University of Michigan Medical School, L2221 UH South, 1500 East Medical Center Drive, Ann Arbor, MI 48109-5212, USA
| | - Sebastián A Riquelme
- Department of Pediatrics, College of Physicians and Surgeons, Columbia University, Columbia University Medical Center, 650West 168th Street, New York, NY 10032, USA
| | - Katherine B Hisert
- Department of Medicine, National Jewish Health, Smith A550, 1400 Jackson Street, Denver, CO 80205, USA.
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17
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Amaral MD. Using the genome to correct the ion transport defect in cystic fibrosis. J Physiol 2022; 601:1573-1582. [PMID: 36068724 DOI: 10.1113/jp282308] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2022] [Accepted: 08/31/2022] [Indexed: 11/08/2022] Open
Abstract
KEY POINTS Human genome information can help finding drugs for human diseases. 'Omics' allow unbiased identification of novel drug targets. High-throughput (HT) approaches provide a global view on disease mechanisms. As a monogenic disease CF has led the way in multiple 'Omic' studies. 'Multi-omics' integration will generate maximal biological significance. ABSTRACT Today Biomedicine faces one of its greatest challenges, i.e. treating diseases through their causative dysfunctional processes and not just their symptoms. However, we still miss a global view of mechanisms and pathways involved in pathophysiology of most diseases. In fact, disease mechanisms and pathways can be achieved by holistic studies provided by 'Omic' approaches. Cystic Fibrosis (CF), caused by mutations in the CF transmembrane conductance regulator (CFTR) gene which encodes an anion channel, is paradigmatic for monogenic disorders, namely channelopathies. A high number of 'omics studies' have focussed on CF, namely several cell-based high-throughput (HT) approaches were developed and applied towards a global mechanistic characterization of CF pathophysiology and the identification of novel and 'unbiased' drug targets. Notwithstanding, it is likely that, through the integration of all these 'layers' of large datasets into comprehensive disease maps that biological significance can be extracted so that the enormous potential of these approaches to identifying dysfunctional mechanisms and novel drugs may become a reality. Abstract figure legend Schematic overview of the 3 main approaches to discovery of new drugs/drug targets. This article is protected by copyright. All rights reserved.
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Affiliation(s)
- Margarida D Amaral
- BioISI - Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisboa, Campo Grande-C8 bdg, Lisboa, 1749-016, Portugal
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18
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Jaganathan D, Bruscia EM, Kopp BT. Emerging Concepts in Defective Macrophage Phagocytosis in Cystic Fibrosis. Int J Mol Sci 2022; 23:7750. [PMID: 35887098 PMCID: PMC9319215 DOI: 10.3390/ijms23147750] [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: 06/10/2022] [Revised: 07/09/2022] [Accepted: 07/11/2022] [Indexed: 01/18/2023] Open
Abstract
Cystic fibrosis (CF) is caused by mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Chronic inflammation and decline in lung function are major reasons for morbidity in CF. Mutant CFTR expressed in phagocytic cells such as macrophages contributes to persistent infection, inflammation, and lung disease in CF. Macrophages play a central role in innate immunity by eliminating pathogenic microbes by a process called phagocytosis. Phagocytosis is required for tissue homeostasis, balancing inflammation, and crosstalk with the adaptive immune system for antigen presentation. This review focused on (1) current understandings of the signaling underlying phagocytic mechanisms; (2) existing evidence for phagocytic dysregulation in CF; and (3) the emerging role of CFTR modulators in influencing CF phagocytic function. Alterations in CF macrophages from receptor initiation to phagosome formation are linked to disease progression in CF. A deeper understanding of macrophages in the context of CFTR and phagocytosis proteins at each step of phagosome formation might contribute to the new therapeutic development of dysregulated innate immunity in CF. Therefore, the review also indicates future areas of research in the context of CFTR and macrophages.
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Affiliation(s)
- Devi Jaganathan
- Center for Microbial Pathogenesis, The Abigail Wexner Research Institute at Nationwide Children’s Hospital, Columbus, OH 43205, USA;
| | - Emanuela M. Bruscia
- Department of Pediatrics, Yale University School of Medicine, New Haven, CT 06510, USA;
| | - Benjamin T. Kopp
- Center for Microbial Pathogenesis, The Abigail Wexner Research Institute at Nationwide Children’s Hospital, Columbus, OH 43205, USA;
- Division of Pulmonary Medicine, Nationwide Children’s Hospital, 700 Children’s Drive, Columbus, OH 43205, USA
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19
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Network-Based Methods for Approaching Human Pathologies from a Phenotypic Point of View. Genes (Basel) 2022; 13:genes13061081. [PMID: 35741843 PMCID: PMC9222217 DOI: 10.3390/genes13061081] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2022] [Revised: 06/10/2022] [Accepted: 06/14/2022] [Indexed: 01/27/2023] Open
Abstract
Network and systemic approaches to studying human pathologies are helping us to gain insight into the molecular mechanisms of and potential therapeutic interventions for human diseases, especially for complex diseases where large numbers of genes are involved. The complex human pathological landscape is traditionally partitioned into discrete “diseases”; however, that partition is sometimes problematic, as diseases are highly heterogeneous and can differ greatly from one patient to another. Moreover, for many pathological states, the set of symptoms (phenotypes) manifested by the patient is not enough to diagnose a particular disease. On the contrary, phenotypes, by definition, are directly observable and can be closer to the molecular basis of the pathology. These clinical phenotypes are also important for personalised medicine, as they can help stratify patients and design personalised interventions. For these reasons, network and systemic approaches to pathologies are gradually incorporating phenotypic information. This review covers the current landscape of phenotype-centred network approaches to study different aspects of human diseases.
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20
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Molecular mechanisms of Cystic Fibrosis - how mutations lead to misfunction and guide therapy. Biosci Rep 2022; 42:231430. [PMID: 35707985 PMCID: PMC9251585 DOI: 10.1042/bsr20212006] [Citation(s) in RCA: 19] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/03/2022] [Revised: 06/03/2022] [Accepted: 06/13/2022] [Indexed: 11/17/2022] Open
Abstract
Cystic fibrosis, the most common autosomal recessive disorder in Caucasians, is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which encodes a cAMP-activated chloride and bicarbonate channel that regulates ion and water transport in secretory epithelia. Although all mutations lead to the lack or reduction in channel function, the mechanisms through which this occurs are diverse – ranging from lack of full-length mRNA, reduced mRNA levels, impaired folding and trafficking, targeting to degradation, decreased gating or conductance, and reduced protein levels to decreased half-life at the plasma membrane. Here, we review the different molecular mechanisms that cause cystic fibrosis and detail how these differences identify theratypes that can inform the use of directed therapies aiming at correcting the basic defect. In summary, we travel through CFTR life cycle from the gene to function, identifying what can go wrong and what can be targeted in terms of the different types of therapeutic approaches.
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21
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Simonin JL, Luscher A, Losa D, Badaoui M, van Delden C, Köhler T, Chanson M. Surface Hydration Protects Cystic Fibrosis Airways from Infection by Restoring Junctional Networks. Cells 2022; 11:cells11091587. [PMID: 35563895 PMCID: PMC9105190 DOI: 10.3390/cells11091587] [Citation(s) in RCA: 3] [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: 03/17/2022] [Revised: 04/20/2022] [Accepted: 05/06/2022] [Indexed: 02/01/2023] Open
Abstract
Defective hydration of airway surface mucosa is associated with recurrent lung infection in cystic fibrosis (CF), a disease caused by CF transmembrane conductance regulator (CFTR) gene mutations. Whether the composition and/or presence of an airway surface liquid (ASL) is sufficient to prevent infection remains unclear. The susceptibility to infection of polarized wild type and CFTR knockdown (CFTR-KD) airway epithelial cells was determined in the presence or absence of a healthy ASL or physiological saline. CFTR-KD epithelia exhibited strong ASL volume reduction, enhanced susceptibility to infection, and reduced junctional integrity. Interestingly, the presence of an apical physiological saline alleviated disruption of the airway epithelial barrier by stimulating essential junctional protein expression. Thus, rehydrated CFTR-KD cells were protected from infection despite normally intense bacterial growth. This study indicates that an epithelial integrity gatekeeper is modulated by the presence of an apical liquid volume, irrespective of the liquid's composition and of expression of a functional CFTR.
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Affiliation(s)
- Juliette L. Simonin
- Department of Cell Physiology & Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (J.L.S.); (D.L.); (M.B.)
| | - Alexandre Luscher
- Department of Microbiology and Molecular Medicine, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (A.L.); (C.v.D.); (T.K.)
| | - Davide Losa
- Department of Cell Physiology & Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (J.L.S.); (D.L.); (M.B.)
| | - Mehdi Badaoui
- Department of Cell Physiology & Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (J.L.S.); (D.L.); (M.B.)
| | - Christian van Delden
- Department of Microbiology and Molecular Medicine, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (A.L.); (C.v.D.); (T.K.)
- Department of Medicine Specialties, Division of Infectious Diseases, Geneva University Hospitals, 1211 Geneva, Switzerland
| | - Thilo Köhler
- Department of Microbiology and Molecular Medicine, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (A.L.); (C.v.D.); (T.K.)
| | - Marc Chanson
- Department of Cell Physiology & Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland; (J.L.S.); (D.L.); (M.B.)
- Correspondence: ; Tel./Fax: +41-22-37-95-206
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22
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Abstract
Cystic fibrosis (CF), the most common genetic disease among the Caucasian population, is caused by mutations in the gene encoding for the CF transmembrane conductance regulator (CFTR), a chloride epithelial channel whose dysfunction results in severe airway obstruction and inflammation, eventually leading to respiratory failure. The discovery of the CFTR gene in 1989 provided new insights into the basic genetic defect of CF and allowed the study of potential therapies targeting the aberrant protein. In recent years, the approval of “CFTR modulators”, the first molecules designed to selectively target the underlying molecular defects caused by specific CF-causing mutations, marked the beginning of a new era in CF treatment. These drugs have been demonstrated to significantly improve lung function and ameliorate the quality of life of many patients, especially those bearing the most common CFTR mutatant F508del. However, a substantial portion of CF subjects, accounting for ~20% of the European CF population, carry rare CFTR mutations and are still not eligible for CFTR modulator therapy, partly due to our limited understanding of the molecular defects associated with these genetic alterations. Thus, the implementation of models to study the phenotype of these rare CFTR mutations and their response to currently approved drugs, as well as to compounds under research and clinical development, is of key importance. The purpose of this review is to summarize the current knowledge on the potential of CFTR modulators in rescuing the function of rare CF-causing CFTR variants, focusing on both investigational and clinically approved molecules.
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23
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Brusa I, Sondo E, Falchi F, Pedemonte N, Roberti M, Cavalli A. Proteostasis Regulators in Cystic Fibrosis: Current Development and Future Perspectives. J Med Chem 2022; 65:5212-5243. [PMID: 35377645 PMCID: PMC9014417 DOI: 10.1021/acs.jmedchem.1c01897] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
In cystic fibrosis (CF), the deletion of phenylalanine 508 (F508del) in the CF transmembrane conductance regulator (CFTR) leads to misfolding and premature degradation of the mutant protein. These defects can be targeted with pharmacological agents named potentiators and correctors. During the past years, several efforts have been devoted to develop and approve new effective molecules. However, their clinical use remains limited, as they fail to fully restore F508del-CFTR biological function. Indeed, the search for CFTR correctors with different and additive mechanisms has recently increased. Among them, drugs that modulate the CFTR proteostasis environment are particularly attractive to enhance therapy effectiveness further. This Perspective focuses on reviewing the recent progress in discovering CFTR proteostasis regulators, mainly describing the design, chemical structure, and structure-activity relationships. The opportunities, challenges, and future directions in this emerging and promising field of research are discussed, as well.
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Affiliation(s)
- Irene Brusa
- Department of Pharmacy and Biotechnology, University of Bologna, 40126 Bologna, Italy.,Computational & Chemical Biology, Istituto Italiano di Tecnologia, 16163 Genova, Italy
| | - Elvira Sondo
- UOC Genetica Medica, IRCCS Istituto Giannina Gaslini, 16147 Genova, Italy
| | | | | | - Marinella Roberti
- Department of Pharmacy and Biotechnology, University of Bologna, 40126 Bologna, Italy
| | - Andrea Cavalli
- Department of Pharmacy and Biotechnology, University of Bologna, 40126 Bologna, Italy.,Computational & Chemical Biology, Istituto Italiano di Tecnologia, 16163 Genova, Italy
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Amaral MD. Precision medicine for rare diseases: The times they are A-Changin'. Curr Opin Pharmacol 2022; 63:102201. [DOI: 10.1016/j.coph.2022.102201] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/27/2021] [Revised: 01/26/2022] [Accepted: 01/31/2022] [Indexed: 12/30/2022]
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Kim YJ, Sivetz N, Layne J, Voss DM, Yang L, Zhang Q, Krainer AR. Exon-skipping antisense oligonucleotides for cystic fibrosis therapy. Proc Natl Acad Sci U S A 2022; 119:e2114858118. [PMID: 35017301 PMCID: PMC8784140 DOI: 10.1073/pnas.2114858118] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2021] [Accepted: 11/16/2021] [Indexed: 11/25/2022] Open
Abstract
Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene cause cystic fibrosis (CF), and the CFTR-W1282X nonsense mutation causes a severe form of CF. Although Trikafta and other CFTR-modulation therapies benefit most CF patients, targeted therapy for patients with the W1282X mutation is lacking. The CFTR-W1282X protein has residual activity but is expressed at a very low level due to nonsense-mediated messenger RNA (mRNA) decay (NMD). NMD-suppression therapy and read-through therapy are actively being researched for CFTR nonsense mutants. NMD suppression could increase the mutant CFTR mRNA, and read-through therapies may increase the levels of full-length CFTR protein. However, these approaches have limitations and potential side effects: because the NMD machinery also regulates the expression of many normal mRNAs, broad inhibition of the pathway is not desirable, and read-through drugs are inefficient partly because the mutant mRNA template is subject to NMD. To bypass these issues, we pursued an exon-skipping antisense oligonucleotide (ASO) strategy to achieve gene-specific NMD evasion. A cocktail of two splice-site-targeting ASOs induced the expression of CFTR mRNA without the premature-termination-codon-containing exon 23 (CFTR-Δex23), which is an in-frame exon. Treatment of human bronchial epithelial cells with this cocktail of ASOs that target the splice sites flanking exon 23 results in efficient skipping of exon 23 and an increase in CFTR-Δex23 protein. The splice-switching ASO cocktail increases the CFTR-mediated chloride current in human bronchial epithelial cells. Our results set the stage for developing an allele-specific therapy for CF caused by the W1282X mutation.
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Affiliation(s)
- Young Jin Kim
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
- Graduate Program in Genetics, Stony Brook University, Stony Brook, NY 11794
- Medical Scientist Training Program, Stony Brook University School of Medicine, Stony Brook, NY 11794
| | - Nicole Sivetz
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
| | - Jessica Layne
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
| | - Dillon M Voss
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
- Graduate Program in Genetics, Stony Brook University, Stony Brook, NY 11794
- Medical Scientist Training Program, Stony Brook University School of Medicine, Stony Brook, NY 11794
| | - Lucia Yang
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
- Graduate Program in Genetics, Stony Brook University, Stony Brook, NY 11794
- Medical Scientist Training Program, Stony Brook University School of Medicine, Stony Brook, NY 11794
| | - Qian Zhang
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
- Graduate Program in Molecular and Cell Biology, Stony Brook University, Stony Brook, NY 11794
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Yavarian J, Zebardast A, Latifi T. The role of severe acute respiratory syndrome coronavirus 2 viroporins in inflammation. ADVANCES IN HUMAN BIOLOGY 2022. [DOI: 10.4103/aihb.aihb_108_21] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022] Open
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Butnariu LI, Țarcă E, Cojocaru E, Rusu C, Moisă ȘM, Leon Constantin MM, Gorduza EV, Trandafir LM. Genetic Modifying Factors of Cystic Fibrosis Phenotype: A Challenge for Modern Medicine. J Clin Med 2021; 10:5821. [PMID: 34945117 PMCID: PMC8707808 DOI: 10.3390/jcm10245821] [Citation(s) in RCA: 23] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2021] [Revised: 12/06/2021] [Accepted: 12/07/2021] [Indexed: 12/13/2022] Open
Abstract
Cystic fibrosis (CF) is a monogenic autosomal recessive disease caused by cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations. CF is characterized by a high phenotypic variability present even in patients with the same genotype. This is due to the intervention of modifier genes that interact with both the CFTR gene and environmental factors. The purpose of this review is to highlight the role of non-CFTR genetic factors (modifier genes) that contribute to phenotypic variability in CF. We analyzed literature data starting with candidate gene studies and continuing with extensive studies, such as genome-wide association studies (GWAS) and whole exome sequencing (WES). The results of both types of studies revealed that the number of modifier genes in CF patients is impressive. Their identification offers a new perspective on the pathophysiological mechanisms of the disease, paving the way for the understanding of other genetic disorders. In conclusion, in the future, genetic analysis, such as GWAS and WES, should be performed routinely. A challenge for future research is to integrate their results in the process of developing new classes of drugs, with a goal to improve the prognosis, increase life expectancy, and enhance quality of life among CF patients.
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Affiliation(s)
- Lăcrămioara Ionela Butnariu
- Department of Medical Genetics, Faculty of Medicine, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iasi, Romania; (L.I.B.); (C.R.); (E.V.G.)
| | - Elena Țarcă
- Department of Surgery II—Pediatric Surgery, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iaşi, Romania
| | - Elena Cojocaru
- Department of Morphofunctional Sciences I, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iaşi, Romania
| | - Cristina Rusu
- Department of Medical Genetics, Faculty of Medicine, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iasi, Romania; (L.I.B.); (C.R.); (E.V.G.)
| | - Ștefana Maria Moisă
- Department of Mother and Child, Faculty of Medicine, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iasi, Romania; (Ș.M.M.); (L.M.T.)
| | | | - Eusebiu Vlad Gorduza
- Department of Medical Genetics, Faculty of Medicine, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iasi, Romania; (L.I.B.); (C.R.); (E.V.G.)
| | - Laura Mihaela Trandafir
- Department of Mother and Child, Faculty of Medicine, “Grigore T. Popa” University of Medicine and Pharmacy, 700115 Iasi, Romania; (Ș.M.M.); (L.M.T.)
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Farinha CM, Gentzsch M. Revisiting CFTR Interactions: Old Partners and New Players. Int J Mol Sci 2021; 22:13196. [PMID: 34947992 PMCID: PMC8703571 DOI: 10.3390/ijms222413196] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/17/2021] [Revised: 12/02/2021] [Accepted: 12/03/2021] [Indexed: 01/07/2023] Open
Abstract
Remarkable progress in CFTR research has led to the therapeutic development of modulators that rescue the basic defect in cystic fibrosis. There is continuous interest in studying CFTR molecular disease mechanisms as not all cystic fibrosis patients have a therapeutic option available. Addressing the basis of the problem by comprehensively understanding the critical molecular associations of CFTR interactions remains key. With the availability of CFTR modulators, there is interest in comprehending which interactions are critical to rescue CFTR and which are altered by modulators or CFTR mutations. Here, the current knowledge on interactions that govern CFTR folding, processing, and stability is summarized. Furthermore, we describe protein complexes and signal pathways that modulate the CFTR function. Primary epithelial cells display a spatial control of the CFTR interactions and have become a common system for preclinical and personalized medicine studies. Strikingly, the novel roles of CFTR in development and differentiation have been recently uncovered and it has been revealed that specific CFTR gene interactions also play an important role in transcriptional regulation. For a comprehensive understanding of the molecular environment of CFTR, it is important to consider CFTR mutation-dependent interactions as well as factors affecting the CFTR interactome on the cell type, tissue-specific, and transcriptional levels.
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Affiliation(s)
- Carlos M. Farinha
- BioISI—Biosystems and Integrative Sciences Institute, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
| | - Martina Gentzsch
- Marsico Lung Institute and Cystic Fibrosis Research Center, School of Medicine, University of North Carolina, Chapel Hill, NC 27599, USA
- Department of Pediatrics, Division of Pediatric Pulmonology, School of Medicine, University of North Carolina, Chapel Hill, NC 27599, USA
- Department of Cell Biology and Physiology, School of Medicine, University of North Carolina, Chapel Hill, NC 27599, USA
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Liu M, Zaman R, Sawczak V, Periasamy A, Sun F, Zaman K. S-nitrosothiols signaling in cystic fibrosis airways. J Biosci 2021. [DOI: 10.1007/s12038-021-00223-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
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Nardella C, Visconti L, Malagrinò F, Pagano L, Bufano M, Nalli M, Coluccia A, La Regina G, Silvestri R, Gianni S, Toto A. Targeting PDZ domains as potential treatment for viral infections, neurodegeneration and cancer. Biol Direct 2021; 16:15. [PMID: 34641953 PMCID: PMC8506081 DOI: 10.1186/s13062-021-00303-9] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2021] [Accepted: 09/24/2021] [Indexed: 02/08/2023] Open
Abstract
The interaction between proteins is a fundamental event for cellular life that is generally mediated by specialized protein domains or modules. PDZ domains are the largest class of protein-protein interaction modules, involved in several cellular pathways such as signal transduction, cell-cell junctions, cell polarity and adhesion, and protein trafficking. Because of that, dysregulation of PDZ domain function often causes the onset of pathologies, thus making this family of domains an interesting pharmaceutical target. In this review article we provide an overview of the structural and functional features of PDZ domains and their involvement in the cellular and molecular pathways at the basis of different human pathologies. We also discuss some of the strategies that have been developed with the final goal to hijack or inhibit the interaction of PDZ domains with their ligands. Because of the generally low binding selectivity of PDZ domain and the scarce efficiency of small molecules in inhibiting PDZ binding, this task resulted particularly difficult to pursue and still demands increasing experimental efforts in order to become completely feasible and successful in vivo.
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Affiliation(s)
- Caterina Nardella
- Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli" and Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, 00185, Rome, Italy
| | - Lorenzo Visconti
- Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli" and Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, 00185, Rome, Italy
| | - Francesca Malagrinò
- Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli" and Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, 00185, Rome, Italy
| | - Livia Pagano
- Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli" and Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, 00185, Rome, Italy
| | - Marianna Bufano
- Laboratory Affiliated with the Institute Pasteur Italy - Cenci Bolognetti Foundation, Department of Drug Chemistry and Technologies, Sapienza University of Rome, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Marianna Nalli
- Laboratory Affiliated with the Institute Pasteur Italy - Cenci Bolognetti Foundation, Department of Drug Chemistry and Technologies, Sapienza University of Rome, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Antonio Coluccia
- Laboratory Affiliated with the Institute Pasteur Italy - Cenci Bolognetti Foundation, Department of Drug Chemistry and Technologies, Sapienza University of Rome, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Giuseppe La Regina
- Laboratory Affiliated with the Institute Pasteur Italy - Cenci Bolognetti Foundation, Department of Drug Chemistry and Technologies, Sapienza University of Rome, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Romano Silvestri
- Laboratory Affiliated with the Institute Pasteur Italy - Cenci Bolognetti Foundation, Department of Drug Chemistry and Technologies, Sapienza University of Rome, Piazzale Aldo Moro 5, 00185, Rome, Italy.
| | - Stefano Gianni
- Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli" and Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, 00185, Rome, Italy.
| | - Angelo Toto
- Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli" and Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, 00185, Rome, Italy.
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31
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Domain Analysis and Motif Matcher (DAMM): A Program to Predict Selectivity Determinants in Monosiga brevicollis PDZ Domains Using Human PDZ Data. Molecules 2021; 26:molecules26196034. [PMID: 34641578 PMCID: PMC8512817 DOI: 10.3390/molecules26196034] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2021] [Revised: 09/30/2021] [Accepted: 10/01/2021] [Indexed: 11/17/2022] Open
Abstract
Choanoflagellates are single-celled eukaryotes with complex signaling pathways. They are considered the closest non-metazoan ancestors to mammals and other metazoans and form multicellular-like states called rosettes. The choanoflagellate Monosiga brevicollis contains over 150 PDZ domains, an important peptide-binding domain in all three domains of life (Archaea, Bacteria, and Eukarya). Therefore, an understanding of PDZ domain signaling pathways in choanoflagellates may provide insight into the origins of multicellularity. PDZ domains recognize the C-terminus of target proteins and regulate signaling and trafficking pathways, as well as cellular adhesion. Here, we developed a computational software suite, Domain Analysis and Motif Matcher (DAMM), that analyzes peptide-binding cleft sequence identity as compared with human PDZ domains and that can be used in combination with literature searches of known human PDZ-interacting sequences to predict target specificity in choanoflagellate PDZ domains. We used this program, protein biochemistry, fluorescence polarization, and structural analyses to characterize the specificity of A9UPE9_MONBE, a M. brevicollis PDZ domain-containing protein with no homology to any metazoan protein, finding that its PDZ domain is most similar to those of the DLG family. We then identified two endogenous sequences that bind A9UPE9 PDZ with <100 μM affinity, a value commonly considered the threshold for cellular PDZ-peptide interactions. Taken together, this approach can be used to predict cellular targets of previously uncharacterized PDZ domains in choanoflagellates and other organisms. Our data contribute to investigations into choanoflagellate signaling and how it informs metazoan evolution.
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Duan Y, Li G, Xu M, Qi X, Deng M, Lin X, Lei Z, Hu Y, Jia Z, Yang Q, Cao G, Liu Z, Wen Q, Li Z, Tang J, Zhang WK, Huang P, Zheng L, Flavell RA, Hao J, Yin Z. CFTR is a negative regulator of γδ T cell IFN-γ production and antitumor immunity. Cell Mol Immunol 2021; 18:1934-1944. [PMID: 32669666 PMCID: PMC8322328 DOI: 10.1038/s41423-020-0499-3] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/17/2020] [Accepted: 06/24/2020] [Indexed: 11/09/2022] Open
Abstract
CFTR, a chloride channel and ion channel regulator studied mostly in epithelial cells, has been reported to participate in immune regulation and likely affect the risk of cancer development. However, little is known about the effects of CFTR on the differentiation and function of γδ T cells. In this study, we observed that CFTR was functionally expressed on the cell surface of γδ T cells. Genetic deletion and pharmacological inhibition of CFTR both increased IFN-γ release by peripheral γδ T cells and potentiated the cytolytic activity of these cells against tumor cells both in vitro and in vivo. Interestingly, the molecular mechanisms underlying the regulation of γδ T cell IFN-γ production by CFTR were either TCR dependent or related to Ca2+ influx. CFTR was recruited to TCR immunological synapses and attenuated Lck-P38 MAPK-c-Jun signaling. In addition, CFTR was found to modulate TCR-induced Ca2+ influx and membrane potential (Vm)-induced Ca2+ influx and subsequently regulate the calcineurin-NFATc1 signaling pathway in γδ T cells. Thus, CFTR serves as a negative regulator of IFN-γ production in γδ T cells and the function of these cells in antitumor immunity. Our investigation suggests that modification of the CFTR activity of γδ T cells may be a potential immunotherapeutic strategy for cancer.
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Affiliation(s)
- Yuanyuan Duan
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Guangqiang Li
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Miaomiao Xu
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Xiaofei Qi
- Department of Physiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, 510515, Guangdong, China
| | - Mingxia Deng
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Xuejia Lin
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Zhiwei Lei
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Yi Hu
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Zhenghu Jia
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
- The First Affiliated Hospital, Jinan University, Guangzhou, 510632, China
| | - Quanli Yang
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
- The First Affiliated Hospital, Jinan University, Guangzhou, 510632, China
| | - Guangchao Cao
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Zonghua Liu
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Qiong Wen
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Zhenhua Li
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China
| | - Jie Tang
- Department of Physiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, 510515, Guangdong, China
| | - Wei Kevin Zhang
- School of Pharmaceutical Sciences, South-Central University for Nationalities, Wuhan, 430074, Hubei, China
| | - Pingbo Huang
- Division of Life Science, Hong Kong University of Science and Technology (HKUST), Hong Kong, China
| | - Limin Zheng
- State Key Laboratory of Oncology in Southern China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, 510060, China
| | - Richard A Flavell
- Department of Immunobiology, School of Medicine, Yale University, New Haven, CT, 06520, USA
| | - Jianlei Hao
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China.
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China.
| | - Zhinan Yin
- Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China.
- The Biomedical Translational Research Institute, Faculty of Medical Science, Jinan University, Guangzhou, 510632, Guangdong, China.
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Abstract
PURPOSE OF REVIEW Organ transplantation research has led to the discovery of several interesting individual mechanistic pathways, molecules and potential drug targets but there are still no comprehensive studies that have addressed how these varied mechanisms work in unison to regulate the posttransplant immune response that drives kidney rejection and dysfunction. RECENT FINDINGS Systems biology is a rapidly expanding field that aims to integrate existing knowledge of molecular concepts and large-scale genomic and clinical datasets into networks that can be used in cutting edge computational models to define disease mechanisms in a holistic manner. Systems biology approaches have brought a paradigm shift from a reductionist view of biology to a wider agnostic assessment of disease from several lines of evidence. Although the complex nature of the posttransplant immune response makes it difficult to pinpoint mechanisms, systems biology is enabling discovery of unknown biological interactions using the cumulative power of genomic data sets, clinical data and endpoints, and improved computational methods for the systematic deconvolution of this response. SUMMARY An integrative systems biology approach that leverages genomic data from varied technologies, such as DNA sequencing, copy number variation, RNA sequencing, and methylation profiles along with long-term clinical follow-up data has the potential to define a framework that can be mined to provide novel insights for developing therapeutic interventions in organ transplantation.
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Hofmann F. The cGMP system: components and function. Biol Chem 2021; 401:447-469. [PMID: 31747372 DOI: 10.1515/hsz-2019-0386] [Citation(s) in RCA: 51] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2019] [Accepted: 10/30/2019] [Indexed: 12/29/2022]
Abstract
The cyclic guanosine monophosphate (cGMP) signaling system is one of the most prominent regulators of a variety of physiological and pathophysiological processes in many mammalian and non-mammalian tissues. Targeting this pathway by increasing cGMP levels has been a very successful approach in pharmacology as shown for nitrates, phosphodiesterase (PDE) inhibitors and stimulators of nitric oxide-guanylyl cyclase (NO-GC) and particulate GC (pGC). This is an introductory review to the cGMP signaling system intended to introduce those readers to this system, who do not work in this area. This article does not intend an in-depth review of this system. Signal transduction by cGMP is controlled by the generating enzymes GCs, the degrading enzymes PDEs and the cGMP-regulated enzymes cyclic nucleotide-gated ion channels, cGMP-dependent protein kinases and cGMP-regulated PDEs. Part A gives a very concise introduction to the components. Part B gives a very concise introduction to the functions modulated by cGMP. The article cites many recent reviews for those who want a deeper insight.
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Affiliation(s)
- Franz Hofmann
- Pharmakologisches Institut, Technische Universität München, Biedersteiner Str. 29, D-80802 München, Germany
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Abstract
Cystic Fibrosis (CF) is the commonest inherited genetic disorder in Caucasians due to a mutation in the gene CFTR (Cystic Fibrosis Transmembrane Conductance Regulator), and it should be considered as an Inherited Colorectal Cancer (CRC) Syndrome. In the United States, physicians of CF Foundation established the “Developing Innovative Gastroenterology Speciality Training Program” to increase the research on CF in gastrointestinal and hepatobiliary diseases. The risk to develop a CRC is 5–10 times higher in CF patients than in the general population and even greater in CF patients receiving immunosuppressive therapy due to organ transplantation (30-fold increased risk relative to the general population). Colonoscopy should be considered the best screening for CRC in CF patients. The screening colonoscopy should be started at the age of 40 in CF patients and, if negative, a new colonoscopy should be performed every 5 years and every 3 years if adenomas are detected. For transplanted CF patients, the screening colonoscopy could be started at the age of 35, in transplanted patients at the age of 30 and, if before, at the age of 30. CF transplanted patients, between the age of 35 and 55, must repeat colonoscopy every 3 years. Our review draws attention towards the clinically relevant development of CRC in CF patients, and it may pave the way for further screenings and studies.
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Stanke F, Janciauskiene S, Tamm S, Wrenger S, Raddatz EL, Jonigk D, Braubach P. Effect of Alpha-1 Antitrypsin on CFTR Levels in Primary Human Airway Epithelial Cells Grown at the Air-Liquid-Interface. Molecules 2021; 26:molecules26092639. [PMID: 33946490 PMCID: PMC8125203 DOI: 10.3390/molecules26092639] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2021] [Accepted: 04/29/2021] [Indexed: 11/16/2022] Open
Abstract
The cystic fibrosis transmembrane conductance regulator (CFTR) gene is influenced by the fundamental cellular processes like epithelial differentiation/polarization, regeneration and epithelial–mesenchymal transition. Defects in CFTR protein levels and/or function lead to decreased airway surface liquid layer facilitating microbial colonization and inflammation. The SERPINA1 gene, encoding alpha1-antitrypsin (AAT) protein, is one of the genes implicated in CF, however it remains unknown whether AAT has any influence on CFTR levels. In this study we assessed CFTR protein levels in primary human lung epithelial cells grown at the air-liquid-interface (ALI) alone or pre-incubated with AAT by Western blots and immunohistochemistry. Histological analysis of ALI inserts revealed CFTR- and AAT-positive cells but no AAT-CFTR co-localization. When 0.5 mg/mL of AAT was added to apical or basolateral compartments of pro-inflammatory activated ALI cultures, CFTR levels increased relative to activated ALIs. This finding suggests that AAT is CFTR-modulating protein, albeit its effects may depend on the concentration and the route of administration. Human lung epithelial ALI cultures provide a useful tool for studies in detail how AAT or other pharmaceuticals affect the levels and activity of CFTR.
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Affiliation(s)
- Frauke Stanke
- Department of Pediatric Pneumology, Neonatology and Allergology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.T.); (E.L.R.)
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Centre for Lung Research, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.J.); (S.W.); (D.J.); (P.B.)
- Correspondence: ; Tel.: +49-511-5326722
| | - Sabina Janciauskiene
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Centre for Lung Research, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.J.); (S.W.); (D.J.); (P.B.)
- Department of Respiratory Medicine, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany
| | - Stephanie Tamm
- Department of Pediatric Pneumology, Neonatology and Allergology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.T.); (E.L.R.)
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Centre for Lung Research, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.J.); (S.W.); (D.J.); (P.B.)
| | - Sabine Wrenger
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Centre for Lung Research, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.J.); (S.W.); (D.J.); (P.B.)
- Department of Respiratory Medicine, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany
| | - Ellen Luise Raddatz
- Department of Pediatric Pneumology, Neonatology and Allergology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.T.); (E.L.R.)
| | - Danny Jonigk
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Centre for Lung Research, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.J.); (S.W.); (D.J.); (P.B.)
- Institute of Pathology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany
| | - Peter Braubach
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Centre for Lung Research, Carl-Neuberg-Str. 1, 30625 Hannover, Germany; (S.J.); (S.W.); (D.J.); (P.B.)
- Institute of Pathology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany
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37
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Lopes-Pacheco M, Pedemonte N, Veit G. Discovery of CFTR modulators for the treatment of cystic fibrosis. Expert Opin Drug Discov 2021; 16:897-913. [PMID: 33823716 DOI: 10.1080/17460441.2021.1912732] [Citation(s) in RCA: 37] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
INTRODUCTION Cystic fibrosis (CF) is a life-threatening inherited disease caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR) protein, an anion channel expressed at the apical membrane of secretory epithelia. CF leads to multiorgan dysfunction with progressive deterioration of lung function being the major cause of untimely death. Conventional CF therapies target only symptoms and consequences downstream of the primary genetic defect and the current life expectancy and quality of life of these individuals are still very limited. AREA COVERED CFTR modulator drugs are novel-specialized therapies that enhance or even restore functional expression of CFTR mutants and have been approved for clinical use for individuals with specific CF genotypes. This review summarizes classical approaches used for the pre-clinical development of CFTR correctors and potentiators as well as emerging strategies aiming to accelerate modulator development and expand theratyping efforts. EXPERT OPINION Highly effective CFTR modulator drugs are expected to deeply modify the disease course for the majority of individuals with CF. A multitude of experimental approaches have been established to accelerate the development of novel modulators. CF patient-derived specimens are valuable cell models to predict therapeutic effectiveness of existing (and novel) modulators in a precision medicine approach.
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Affiliation(s)
| | | | - Guido Veit
- Department of Physiology, McGill University, Montréal, Canada
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Pathophysiology of Lung Disease and Wound Repair in Cystic Fibrosis. PATHOPHYSIOLOGY 2021; 28:155-188. [PMID: 35366275 PMCID: PMC8830450 DOI: 10.3390/pathophysiology28010011] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2020] [Revised: 03/08/2021] [Accepted: 03/08/2021] [Indexed: 12/11/2022] Open
Abstract
Cystic fibrosis (CF) is an autosomal recessive, life-threatening condition affecting many organs and tissues, the lung disease being the chief cause of morbidity and mortality. Mutations affecting the CF Transmembrane Conductance Regulator (CFTR) gene determine the expression of a dysfunctional protein that, in turn, triggers a pathophysiological cascade, leading to airway epithelium injury and remodeling. In vitro and in vivo studies point to a dysregulated regeneration and wound repair in CF airways, to be traced back to epithelial CFTR lack/dysfunction. Subsequent altered ion/fluid fluxes and/or signaling result in reduced cell migration and proliferation. Furthermore, the epithelial-mesenchymal transition appears to be partially triggered in CF, contributing to wound closure alteration. Finally, we pose our attention to diverse approaches to tackle this defect, discussing the therapeutic role of protease inhibitors, CFTR modulators and mesenchymal stem cells. Although the pathophysiology of wound repair in CF has been disclosed in some mechanisms, further studies are warranted to understand the cellular and molecular events in more details and to better address therapeutic interventions.
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39
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Structural dynamics of ABC transporters: molecular simulation studies. Biochem Soc Trans 2021; 49:405-414. [PMID: 33634827 DOI: 10.1042/bst20200710] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2020] [Revised: 02/02/2021] [Accepted: 02/03/2021] [Indexed: 12/31/2022]
Abstract
The biological activities of living organisms involve various inputs and outputs. The ATP-driven substances (biomolecules) responsible for these kinds of activities through membrane (i.e. uptake and efflux of substrates) include ATP-binding cassette (ABC) transporters, some of which play important roles in multidrug resistance. The basic architecture of ABC transporters comprises transmembrane domains (TMDs) and nucleotide-binding domains (NBDs). The functional dynamics (substrate transport) of ABC transporters are realized by concerted motions, such as NBD dimerization, mechanical transmission via coupling helices (CHs), and the translocation of substrates through TMDs, which are induced by the binding and/or hydrolysis of ATP molecules and substrates. In this mini-review, we briefly discuss recent progresses in the structural dynamics as revealed by molecular simulation studies at all-atom (AA), coarse-grained (CG), and quantum mechanics/molecular mechanics (QM/MM) levels.
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40
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Abdel Hameid R, Cormet-Boyaka E, Kuebler WM, Uddin M, Berdiev BK. SARS-CoV-2 may hijack GPCR signaling pathways to dysregulate lung ion and fluid transport. Am J Physiol Lung Cell Mol Physiol 2021; 320:L430-L435. [PMID: 33434105 PMCID: PMC7938641 DOI: 10.1152/ajplung.00499.2020] [Citation(s) in RCA: 28] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
The tropism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a virus responsible for the ongoing coronavirus disease 2019 (COVID-19) pandemic, toward the host cells is determined, at least in part, by the expression and distribution of its cell surface receptor, angiotensin-converting enzyme 2 (ACE2). The virus further exploits the host cellular machinery to gain access into the cells; its spike protein is cleaved by a host cell surface transmembrane serine protease 2 (TMPRSS2) shortly after binding ACE2, followed by its proteolytic activation at a furin cleavage site. The virus primarily targets the epithelium of the respiratory tract, which is covered by a tightly regulated airway surface liquid (ASL) layer that serves as a primary defense mechanism against respiratory pathogens. The volume and viscosity of this fluid layer is regulated and maintained by a coordinated function of different transport pathways in the respiratory epithelium. We argue that SARS-CoV-2 may potentially alter evolutionary conserved second-messenger signaling cascades via activation of G protein-coupled receptors (GPCRs) or by directly modulating G protein signaling. Such signaling may in turn adversely modulate transepithelial transport processes, especially those involving cystic fibrosis transmembrane conductance regulator (CFTR) and epithelial Na+ channel (ENaC), thereby shifting the delicate balance between anion secretion and sodium absorption, which controls homeostasis of this fluid layer. As a result, activation of the secretory pathways including CFTR-mediated Cl− transport may overwhelm the absorptive pathways, such as ENaC-dependent Na+ uptake, and initiate a pathophysiological cascade leading to lung edema, one of the most serious and potentially deadly clinical manifestations of COVID-19.
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Affiliation(s)
- Reem Abdel Hameid
- Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, United Arab Emirates
| | | | - Wolfgang M Kuebler
- Institute of Physiology, Charité-Universitätsmedizin Berlin, Berlin, Germany
| | - Mohammed Uddin
- Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, United Arab Emirates.,The Centre for Applied Genomics, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Bakhrom K Berdiev
- Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, United Arab Emirates
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Aukema HM. Prostaglandins as potential targets for the treatment of polycystic kidney disease. Prostaglandins Leukot Essent Fatty Acids 2021; 164:102220. [PMID: 33285393 DOI: 10.1016/j.plefa.2020.102220] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/13/2020] [Revised: 11/24/2020] [Accepted: 11/24/2020] [Indexed: 12/15/2022]
Abstract
Polycystic kidney disease (PKD) is characterized by the proliferation of fluid-filled kidney cysts that enlarge over time, causing damage to the surrounding kidney and ultimately resulting in kidney failure. Both increased cell proliferation and fluid secretion are stimulated by increased cyclic adenosine monophosphate (cAMP) in PKD kidneys, so many treatments for the disease target cAMP lowering. Prostaglandins (PG) levels are elevated in multiple animal models of PKD and mediate many of their effects by elevating cAMP levels. Inhibiting the production of PG with cyclooxygenase 2 (COX2) inhibitors reduces PG levels and reduces disease progression. However, COX inhibitors also block beneficial PG and can cause nephrotoxicity. In an orthologous model of the main form of PKD, PGD2 and PGI2 were the two PG highest in kidneys and most affected by a COX2 inhibitor. Future studies are needed to determine whether specific blockage of PGD2 and/or PGI2 activity would lead to more targeted and effective treatments with fewer undesirable side-effects.
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Affiliation(s)
- Harold M Aukema
- Department of Food and Human Nutritional Sciences, University of Manitoba, MB R3T 2N2, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St Boniface Albrechtsen Research Centre, Winnipeg, MB R2H 2A6, Canada.
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42
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Loureiro CA, Pinto FR, Barros P, Matos P, Jordan P. A SYK/SHC1 pathway regulates the amount of CFTR in the plasma membrane. Cell Mol Life Sci 2020; 77:4997-5015. [PMID: 31974654 PMCID: PMC11105000 DOI: 10.1007/s00018-020-03448-4] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2019] [Revised: 12/06/2019] [Accepted: 01/02/2020] [Indexed: 11/24/2022]
Abstract
Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene cause the recessive genetic disease cystic fibrosis, where the chloride transport across the apical membrane of epithelial cells mediated by the CFTR protein is impaired. CFTR protein trafficking to the plasma membrane (PM) is the result of a complex interplay between the secretory and membrane recycling pathways that control the number of channels present at the membrane. In addition, the ion transport activity of CFTR at the PM is modulated through post-translational protein modifications. Previously we described that spleen tyrosine kinase (SYK) phosphorylates a specific tyrosine residue in the nucleotide-binding domain 1 domain and this modification can regulate the PM abundance of CFTR. Here we identified the underlying biochemical mechanism using peptide pull-down assays followed by mass spectrometry. We identified in bronchial epithelial cells that the adaptor protein SHC1 recognizes tyrosine-phosphorylated CFTR through its phosphotyrosine-binding domain and that the formation of a complex between SHC1 and CFTR is induced at the PM in the presence of activated SYK. The depletion of endogenous SHC1 expression was sufficient to promote an increase in CFTR at the PM of these cells. The results identify a SYK/SHC1 pathway that regulates the PM levels of CFTR channels, contributing to a better understanding of how CFTR-mediated chloride secretion is regulated.
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Affiliation(s)
- Cláudia Almeida Loureiro
- Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Avenida Padre Cruz, 1649-016, Lisbon, Portugal
- BioISI-Biosystems and Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Lisbon, Portugal
| | - Francisco R Pinto
- BioISI-Biosystems and Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Lisbon, Portugal
- Department of Chemistry and Biochemistry, Faculty of Sciences, University of Lisbon, Lisbon, Portugal
| | - Patrícia Barros
- Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Avenida Padre Cruz, 1649-016, Lisbon, Portugal
- BioISI-Biosystems and Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Lisbon, Portugal
| | - Paulo Matos
- Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Avenida Padre Cruz, 1649-016, Lisbon, Portugal
- BioISI-Biosystems and Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Lisbon, Portugal
- Department of Chemistry and Biochemistry, Faculty of Sciences, University of Lisbon, Lisbon, Portugal
| | - Peter Jordan
- Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Avenida Padre Cruz, 1649-016, Lisbon, Portugal.
- BioISI-Biosystems and Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Lisbon, Portugal.
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43
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Material properties of interfacial films of mucoid and nonmucoid Pseudomonas aeruginosa isolates. Acta Biomater 2020; 118:129-140. [PMID: 33053427 DOI: 10.1016/j.actbio.2020.10.010] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2020] [Revised: 09/20/2020] [Accepted: 10/07/2020] [Indexed: 12/19/2022]
Abstract
Chronic lung infection with bacterial biofilms is a leading cause of death in cystic fibrosis (CF) patients. Pseudomonas aeruginosa, one of the many species colonizing the lung airways, can undergo pathoadaptation, leading to a mucoid phenotype with interesting material properties. We hypothesize that the surface properties and extracellular materials of mucoid P. aeruginosa cells greatly influence the mechanical behavior of their films at fluid interfaces. In this study, we investigate the interfacial properties of films formed by nonmucoid (PANT) and mucoid (PASL) strains of P. aeruginosa isolated from CF patients. We use pendant drop elastometry to analyze the interfacial response of the films formed by PANT and PASL at the hexadecane-water interface. The dynamic rheological analyses of the films highlight the distinctive signature of the mucoid strains at fluid interfaces. The mucoid films exhibit greater relaxation following a compressive strain than a tensile one, while a full hysteresis response is achieved by the nonmucoid films; this indicates that the material properties of the PANT films are conserved under both compression and tension. The wrinkling and shape analyses of the interfacial bacterial films elucidate that the mucoid strain exhibits remarkable viscoelastic properties, enabling the remodeling of the living films and dissipation of the compressive stress. The comparative analysis of the material properties of mucoid and nonmucoid P. aeruginosa cells indicates that mucoid switch can play an important role in protecting the bacteria from interfacial stresses. Further characterization of interfacial bacterial films will provide new insights into the development of methods for controlling interfacial films of bacteria.
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Amaral MD. How to determine the mechanism of action of CFTR modulator compounds: A gateway to theranostics. Eur J Med Chem 2020; 210:112989. [PMID: 33190956 DOI: 10.1016/j.ejmech.2020.112989] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2020] [Revised: 11/02/2020] [Accepted: 11/02/2020] [Indexed: 12/12/2022]
Abstract
The greatest challenge of 21st century biology is to fully understand mechanisms of disease to drive new approaches and medical innovation. Parallel to this is the huge biomedical endeavour of treating people through personalized medicine. Until now all CFTR modulator drugs that have entered clinical trials have been genotype-dependent. An emerging alternative is personalized/precision medicine in CF, i.e., to determine whether rare CFTR mutations respond to existing (or novel) CFTR modulator drugs by pre-assessing them directly on patient's tissues ex vivo, an approach also now termed theranostics. To administer the right drug to the right person it is essential to understand how drugs work, i.e., to know their mechanism of action (MoA), so as to predict their applicability, not just in certain mutations but also possibly in other diseases that share the same defect/defective pathway. Moreover, an understanding the MoA of a drug before it is tested in clinical trials is the logical path to drug discovery and can increase its chance for success and hence also approval. In conclusion, the most powerful approach to determine the MoA of a compound is to understand the underlying biology. Novel large datasets of intervenients in most biological processes, namely those emerging from the post-genomic era tools, are available and should be used to help in this task.
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Affiliation(s)
- Margarida D Amaral
- BioISI - Biosystems & Integrative Sciences Institute, Lisboa, Faculty of Sciences, University of Lisboa, Portugal.
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45
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Rodrat M, Jantarajit W, Ng DRS, Harvey BSJ, Liu J, Wilkinson WJ, Charoenphandhu N, Sheppard DN. Carbon monoxide-releasing molecules inhibit the cystic fibrosis transmembrane conductance regulator Cl - channel. Am J Physiol Lung Cell Mol Physiol 2020; 319:L997-L1009. [PMID: 32936026 DOI: 10.1152/ajplung.00440.2019] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
The gasotransmitter carbon monoxide (CO) regulates fluid and electrolyte movements across epithelial tissues. However, its action on anion channels is incompletely understood. Here, we investigate the direct action of CO on the cystic fibrosis transmembrane conductance regulator (CFTR) by applying CO-releasing molecules (CO-RMs) to the intracellular side of excised inside-out membrane patches from cells heterologously expressing wild-type human CFTR. Addition of increasing concentrations of tricarbonyldichlororuthenium(II) dimer (CORM-2) (1-300 μM) inhibited CFTR channel activity, whereas the control RuCl3 (100 μM) was without effect. CORM-2 predominantly inhibited CFTR by decreasing the frequency of channel openings and, hence, open probability (Po). But, it also reduced current flow through open channels with very fast kinetics, particularly at elevated concentrations. By contrast, the chemically distinct CO-releasing molecule CORM-3 inhibited CFTR by decreasing Po without altering current flow through open channels. Neither depolarizing the membrane voltage nor raising the ATP concentration on the intracellular side of the membrane affected CFTR inhibition by CORM-2. Interestingly, CFTR inhibition by CORM-2, but not by CFTRinh-172, was prevented by prior enhancement of channel activity by the clinically approved CFTR potentiator ivacaftor. Similarly, when added after CORM-2, ivacaftor completely relieved CFTR inhibition. In conclusion, CORM-2 has complex effects on wild-type human CFTR consistent with allosteric inhibition and open-channel blockade. Inhibition of CFTR by CO-releasing molecules suggests that CO regulates CFTR activity and that the gasotransmitter has tissue-specific effects on epithelial ion transport. The action of ivacaftor on CFTR Cl- channels inhibited by CO potentially expands the drug's clinical utility.
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Affiliation(s)
- Mayuree Rodrat
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom.,Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand.,Department of Physiology, Faculty of Science, Mahidol University, Bangkok, Thailand
| | - Walailak Jantarajit
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom.,Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand.,Department of Physiology, Faculty of Science, Mahidol University, Bangkok, Thailand
| | - Demi R S Ng
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom
| | - Bartholomew S J Harvey
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom
| | - Jia Liu
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom
| | | | - Narattaphol Charoenphandhu
- Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand.,Department of Physiology, Faculty of Science, Mahidol University, Bangkok, Thailand.,Institute of Molecular Biosciences, Mahidol University, Nakhon Pathom, Thailand.,The Academy of Science, The Royal Society of Thailand, Dusit, Bangkok, Thailand
| | - David N Sheppard
- School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom
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Cytoskeleton regulators CAPZA2 and INF2 associate with CFTR to control its plasma membrane levels under EPAC1 activation. Biochem J 2020; 477:2561-2580. [PMID: 32573649 DOI: 10.1042/bcj20200287] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2020] [Revised: 06/19/2020] [Accepted: 06/22/2020] [Indexed: 02/06/2023]
Abstract
Cystic Fibrosis (CF), the most common lethal autosomic recessive disorder among Caucasians, is caused by mutations in the gene encoding the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) protein, a cAMP-regulated chloride channel expressed at the apical surface of epithelial cells. Cyclic AMP regulates both CFTR channel gating through a protein kinase A (PKA)-dependent process and plasma membane (PM) stability through activation of the exchange protein directly activated by cAMP1 (EPAC1). This cAMP effector, when activated promotes the NHERF1:CFTR interaction leading to an increase in CFTR at the PM by decreasing its endocytosis. Here, we used protein interaction profiling and bioinformatic analysis to identify proteins that interact with CFTR under EPAC1 activation as possible regulators of this CFTR PM anchoring. We identified an enrichment in cytoskeleton related proteins among which we characterized CAPZA2 and INF2 as regulators of CFTR trafficking to the PM. We found that CAPZA2 promotes wt-CFTR trafficking under EPAC1 activation at the PM whereas reduction of INF2 levels leads to a similar trafficking promotion effect. These results suggest that CAPZA2 is a positive regulator and INF2 a negative one for the increase of CFTR at the PM after an increase of cAMP and concomitant EPAC1 activation. Identifying the specific interactions involving CFTR and elicited by EPAC1 activation provides novel insights into late CFTR trafficking, insertion and/or stabilization at the PM and highlighs new potential therapeutic targets to tackle CF disease.
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47
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Di Pietro C, Öz HH, Murray TS, Bruscia EM. Targeting the Heme Oxygenase 1/Carbon Monoxide Pathway to Resolve Lung Hyper-Inflammation and Restore a Regulated Immune Response in Cystic Fibrosis. Front Pharmacol 2020; 11:1059. [PMID: 32760278 PMCID: PMC7372134 DOI: 10.3389/fphar.2020.01059] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2020] [Accepted: 06/30/2020] [Indexed: 12/11/2022] Open
Abstract
In individuals with cystic fibrosis (CF), lung hyper-inflammation starts early in life and is perpetuated by mucus obstruction and persistent bacterial infections. The continuous tissue damage and scarring caused by non-resolving inflammation leads to bronchiectasis and, ultimately, respiratory failure. Macrophages (MΦs) are key regulators of immune response and host defense. We and others have shown that, in CF, MΦs are hyper-inflammatory and exhibit reduced bactericidal activity. Thus, MΦs contribute to the inability of CF lung tissues to control the inflammatory response or restore tissue homeostasis. The non-resolving hyper-inflammation in CF lungs is attributed to an impairment of several signaling pathways associated with resolution of the inflammatory response, including the heme oxygenase-1/carbon monoxide (HO-1/CO) pathway. HO-1 is an enzyme that degrades heme groups, leading to the production of potent antioxidant, anti-inflammatory, and bactericidal mediators, such as biliverdin, bilirubin, and CO. This pathway is fundamental to re-establishing cellular homeostasis in response to various insults, such as oxidative stress and infection. Monocytes/MΦs rely on abundant induction of the HO-1/CO pathway for a controlled immune response and for potent bactericidal activity. Here, we discuss studies showing that blunted HO-1 activation in CF-affected cells contributes to hyper-inflammation and defective host defense against bacteria. We dissect potential cellular mechanisms that may lead to decreased HO-1 induction in CF cells. We review literature suggesting that induction of HO-1 may be beneficial for the treatment of CF lung disease. Finally, we discuss recent studies highlighting how endogenous HO-1 can be induced by administration of controlled doses of CO to reduce lung hyper-inflammation, oxidative stress, bacterial infection, and dysfunctional ion transport, which are all hallmarks of CF lung disease.
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Affiliation(s)
| | | | | | - Emanuela M. Bruscia
- Department of Pediatrics, Yale University School of Medicine, New Haven, CT, United States
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48
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Jouret F, Devuyst O. Targeting chloride transport in autosomal dominant polycystic kidney disease. Cell Signal 2020; 73:109703. [PMID: 32619563 DOI: 10.1016/j.cellsig.2020.109703] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2020] [Revised: 06/25/2020] [Accepted: 06/26/2020] [Indexed: 02/07/2023]
Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is the most frequent inherited kidney disease. Transepithelial fluid secretion is one of the key factors of cystogenesis in ADPKD. Multiple studies have suggested that fluid secretion across ADPKD cyst-lining cells is driven by the secretion of chloride, essentially mediated by the CFTR channel and stimulated by increased intracellular levels of 3',5'-cyclic adenosine monophosphate. This review focuses on the pathophysiology of fluid secretion in ADPKD based on the pioneering studies of Jared Grantham and colleagues, and on the follow-up investigations from the molecular level to the potential applications in ADPKD patients. Altogether, the studies of fluid and chloride transport in ADPKD paved the way for innovative therapeutic targets to prevent cyst volume expansion and thus, kidney disease progression.
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Affiliation(s)
- François Jouret
- Division of Nephrology, Department of Internal Medicine, ULiège Academic Hospital, Liège, Belgium,; Groupe Interdisciplinaire de Géno-protéomique Appliquée, Cardiovascular Sciences, ULiège Medical School, Liège, Belgium
| | - Olivier Devuyst
- Division of Nephrology, UCLouvain Medical School, B-1200, Brussels, Belgium,; Mechanisms of Inherited Kidney Disorders, University of Zurich, CH-8057 Zurich, Switzerland.
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Rayner RE, Wellmerling J, Osman W, Honesty S, Alfaro M, Peeples ME, Cormet-Boyaka E. In vitro 3D culture lung model from expanded primary cystic fibrosis human airway cells. J Cyst Fibros 2020; 19:752-761. [PMID: 32565193 DOI: 10.1016/j.jcf.2020.05.007] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2019] [Revised: 04/13/2020] [Accepted: 05/23/2020] [Indexed: 12/12/2022]
Abstract
BACKGROUND In vitro cystic fibrosis (CF) models are crucial for understanding the mechanisms and consequences of the disease. They are also the gold standard for pre-clinical efficacy studies of current and novel CF drugs. However, few studies have investigated expansion and differentiation of primary CF human bronchial epithelial (CF-HBE) cells. Here we describe culture conditions to expand primary CF airway cells while preserving their ability to differentiate into 3D epithelial cultures expressing functional cystic fibrosis transmembrane conductance regulator (CFTR) ion channels that responds to CFTR modulators. METHODS Primary CF airway cells were expanded using PneumaCultTM-Ex Plus (StemCell Technologies) medium with no feeder cells or added Rho kinase (ROCK) inhibitor. Differentially passaged CF-HBE cells at the air-liquid interface (ALI) were characterized phenotypically and functionally in response to the CFTR corrector drug VX-661 (Tezacaftor). RESULTS CF-HBE primary cells, expanded up to six passages (~25 population doublings), differentiated into 3D epithelial cultures as evidenced by trans-epithelial electrical resistance (TEER) of >400 Ohms∙cm2 and presence of pseudostratified columnar ciliated epithelium with goblet cells. However, up to passage five cells from most donors showed increased CFTR-mediated short-circuit currents when treated with the corrector drug, VX-661. Ciliary beat frequency (CBF) also increased with the corrector VX-661. CONCLUSIONS CF donor-derived airway cells can be expanded without the use of feeder cells or additional ROCK inhibitor, and still achieve optimal 3D epithelial cultures that respond to CFTR modulators. The study of rare CF mutations could benefit from cell expansion and could lead to the design of personalized medicine/treatments.
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Affiliation(s)
- Rachael E Rayner
- Department of Veterinary Biosciences, The Ohio State University, 1925 Coffey Road, Columbus, OH 43210, USA
| | - Jack Wellmerling
- Department of Veterinary Biosciences, The Ohio State University, 1925 Coffey Road, Columbus, OH 43210, USA
| | - Wissam Osman
- Department of Veterinary Biosciences, The Ohio State University, 1925 Coffey Road, Columbus, OH 43210, USA
| | - Sean Honesty
- Department of Veterinary Biosciences, The Ohio State University, 1925 Coffey Road, Columbus, OH 43210, USA
| | - Maria Alfaro
- Institute for Genomic Medicine, Abigail Wexner Research Institute, Nationwide Children's Hospital, 700 Children's Drive, Columbus, OH 43205, USA
| | - Mark E Peeples
- Center for Vaccines and Immunity, Abigail Wexner Research Institute, Nationwide Children's Hospital, 700 Children's Drive, Columbus, OH 43205, USA; Department of Pediatrics, The Ohio State University College of Medicine, 700 Children's Drive, Columbus, OH 43205, USA
| | - Estelle Cormet-Boyaka
- Department of Veterinary Biosciences, The Ohio State University, 1925 Coffey Road, Columbus, OH 43210, USA.
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Erwood S, Laselva O, Bily TM, Brewer RA, Rutherford AH, Bear CE, Ivakine EA. Allele-Specific Prevention of Nonsense-Mediated Decay in Cystic Fibrosis Using Homology-Independent Genome Editing. Mol Ther Methods Clin Dev 2020; 17:1118-1128. [PMID: 32490033 PMCID: PMC7256445 DOI: 10.1016/j.omtm.2020.05.002] [Citation(s) in RCA: 32] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2020] [Accepted: 05/07/2020] [Indexed: 02/07/2023]
Abstract
Nonsense-mediated decay (NMD) is a major pathogenic mechanism underlying a diversity of genetic disorders. Nonsense variants tend to lead to more severe disease phenotypes and are often difficult targets for small molecule therapeutic development as a result of insufficient protein production. The treatment of cystic fibrosis (CF), an autosomal recessive disease caused by mutations in the CFTR gene, exemplifies the challenge of therapeutically addressing nonsense mutations in human disease. Therapeutic development in CF has led to multiple, highly successful protein modulatory interventions, yet no targeted therapies have been approved for nonsense mutations. Here, we have designed a CRISPR-Cas9-based strategy for the targeted prevention of NMD of CFTR transcripts containing the second most common nonsense variant listed in CFTR2, W1282X. By introducing a deletion of the downstream genic region following the premature stop codon, we demonstrate significantly increased protein expression of this mutant variant. Notably, in combination with protein modulators, genome editing significantly increases the potentiated channel activity of W1282X-CFTR in human bronchial epithelial cells. Furthermore, we show how the outlined approach can be modified to permit allele-specific editing. The described approach can be extended to other late-occurring nonsense mutations in the CFTR gene or applied as a generalized approach for gene-specific prevention of NMD in disorders where a truncated protein product retains full or partial functionality.
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Affiliation(s)
- Steven Erwood
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada
| | - Onofrio Laselva
- Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Physiology, University of Toronto, Toronto, ON, Canada
| | - Teija M.I. Bily
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
| | - Reid A. Brewer
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
| | - Alexandra H. Rutherford
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
| | - Christine E. Bear
- Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Physiology, University of Toronto, Toronto, ON, Canada
- Department of Biochemistry, University of Toronto, Toronto, ON, Canada
| | - Evgueni A. Ivakine
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Physiology, University of Toronto, Toronto, ON, Canada
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