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Manipulating Cx43 expression triggers gene reprogramming events in dermal fibroblasts from oculodentodigital dysplasia patients.
Biochem J. 2015 Nov 15; 472(1):55-69.BJ

Abstract

Oculodentodigital dysplasia (ODDD) is primarily an autosomal dominant disorder linked to over 70 GJA1 gene [connexin43 (Cx43)] mutations. For nearly a decade, our laboratory has been investigating the relationship between Cx43 and ODDD by expressing disease-linked mutants in reference cells, tissue-relevant cell lines, 3D organ cultures and by using genetically modified mouse models of human disease. Although salient features of Cx43 mutants have been revealed, these models do not necessarily reflect the complexity of the human context. To further overcome these limitations, we have acquired dermal fibroblasts from two ODDD-affected individuals harbouring D3N and V216L mutations in Cx43, along with familial controls. Using these ODDD patient dermal fibroblasts, which naturally produce less GJA1 gene product, along with RNAi and RNA activation (RNAa) approaches, we show that manipulating Cx43 expression triggers cellular gene reprogramming. Quantitative RT-PCR, Western blot and immunofluorescent analysis of ODDD patient fibroblasts show unusually high levels of extracellular matrix (ECM)-interacting proteins, including integrin α5β1, matrix metalloproteinases as well as secreted ECM proteins collagen-I and laminin. Cx43 knockdown in familial control cells produces similar effects on ECM expression, whereas Cx43 transcriptional up-regulation using RNAa decreases production of collagen-I. Interestingly, the enhanced levels of ECM-associated proteins in ODDD V216L fibroblasts is not only a consequence of increased ECM gene expression, but also due to an apparent deficit in collagen-I secretion which may further contribute to impaired collagen gel contraction in ODDD fibroblasts. These findings further illuminate the altered function of Cx43 in ODDD-affected individuals and highlight the impact of manipulating Cx43 expression in human cells.

Authors+Show Affiliations

Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON, Canada, N6A-5C1.Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON, Canada, N6A-5C1.Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON, Canada, N6A-5C1.Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON, Canada, N6A-5C1.Department of Neurology, Stanford University Medical Center, Palo Alto, CA 94305, U.S.A.Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON, Canada, N6A-5C1 dale.laird@schulich.uwo.ca.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

26349540

Citation

Esseltine, Jessica L., et al. "Manipulating Cx43 Expression Triggers Gene Reprogramming Events in Dermal Fibroblasts From Oculodentodigital Dysplasia Patients." The Biochemical Journal, vol. 472, no. 1, 2015, pp. 55-69.
Esseltine JL, Shao Q, Huang T, et al. Manipulating Cx43 expression triggers gene reprogramming events in dermal fibroblasts from oculodentodigital dysplasia patients. Biochem J. 2015;472(1):55-69.
Esseltine, J. L., Shao, Q., Huang, T., Kelly, J. J., Sampson, J., & Laird, D. W. (2015). Manipulating Cx43 expression triggers gene reprogramming events in dermal fibroblasts from oculodentodigital dysplasia patients. The Biochemical Journal, 472(1), 55-69. https://doi.org/10.1042/BJ20150652
Esseltine JL, et al. Manipulating Cx43 Expression Triggers Gene Reprogramming Events in Dermal Fibroblasts From Oculodentodigital Dysplasia Patients. Biochem J. 2015 Nov 15;472(1):55-69. PubMed PMID: 26349540.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Manipulating Cx43 expression triggers gene reprogramming events in dermal fibroblasts from oculodentodigital dysplasia patients. AU - Esseltine,Jessica L, AU - Shao,Qing, AU - Huang,Tao, AU - Kelly,John J, AU - Sampson,Jacinda, AU - Laird,Dale W, Y1 - 2015/09/08/ PY - 2015/06/05/received PY - 2015/09/08/accepted PY - 2015/9/10/entrez PY - 2015/9/10/pubmed PY - 2016/4/2/medline KW - collagen KW - connexin43 KW - extracellular matrix KW - fibroblast KW - gap junctional intercellular communication KW - oculodentodigital dysplasia SP - 55 EP - 69 JF - The Biochemical journal JO - Biochem. J. VL - 472 IS - 1 N2 - Oculodentodigital dysplasia (ODDD) is primarily an autosomal dominant disorder linked to over 70 GJA1 gene [connexin43 (Cx43)] mutations. For nearly a decade, our laboratory has been investigating the relationship between Cx43 and ODDD by expressing disease-linked mutants in reference cells, tissue-relevant cell lines, 3D organ cultures and by using genetically modified mouse models of human disease. Although salient features of Cx43 mutants have been revealed, these models do not necessarily reflect the complexity of the human context. To further overcome these limitations, we have acquired dermal fibroblasts from two ODDD-affected individuals harbouring D3N and V216L mutations in Cx43, along with familial controls. Using these ODDD patient dermal fibroblasts, which naturally produce less GJA1 gene product, along with RNAi and RNA activation (RNAa) approaches, we show that manipulating Cx43 expression triggers cellular gene reprogramming. Quantitative RT-PCR, Western blot and immunofluorescent analysis of ODDD patient fibroblasts show unusually high levels of extracellular matrix (ECM)-interacting proteins, including integrin α5β1, matrix metalloproteinases as well as secreted ECM proteins collagen-I and laminin. Cx43 knockdown in familial control cells produces similar effects on ECM expression, whereas Cx43 transcriptional up-regulation using RNAa decreases production of collagen-I. Interestingly, the enhanced levels of ECM-associated proteins in ODDD V216L fibroblasts is not only a consequence of increased ECM gene expression, but also due to an apparent deficit in collagen-I secretion which may further contribute to impaired collagen gel contraction in ODDD fibroblasts. These findings further illuminate the altered function of Cx43 in ODDD-affected individuals and highlight the impact of manipulating Cx43 expression in human cells. SN - 1470-8728 UR - https://www.unboundmedicine.com/medline/citation/26349540/Manipulating_Cx43_expression_triggers_gene_reprogramming_events_in_dermal_fibroblasts_from_oculodentodigital_dysplasia_patients_ L2 - https://portlandpress.com/biochemj/article-lookup/doi/10.1042/BJ20150652 DB - PRIME DP - Unbound Medicine ER -