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Nat Commun. 2015 Sep 25;6:8391. doi: 10.1038/ncomms9391.

Evolutionarily conserved intercalated disc protein Tmem65 regulates cardiac conduction and connexin 43 function.

Author information

1
Department of Physiology, University of Toronto, Toronto General Hospital Research Institute, Toronto, Ontario, Canada M5G 1L7.
2
Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada M5S 1A8.
3
McEwen Centre for Regenerative Medicine, University Health Network, Toronto, Ontario, Canada M5G 1L7.
4
Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada M5G 1L7.
5
Departments of Molecular Genetics and Biochemistry, Donnelly Centre,, University of Toronto, Toronto, Ontario, Canada M5S 3E1.
6
Department of Mechanical and Industrial Engineering, Advanced Micro and Nanosystems Laboratory, University of Toronto, Toronto, Ontario, Canada M5S 3G8.
7
Institute of Pharmacology, University Medical Center Göttingen and DZHK (German Center for Cardiovascular Research) partner site Göttingen, Göttingen 37075, Germany.
8
The Hospital for Sick Children, Toronto, Ontario, Canada M5G 1X8.
9
Ted Rogers Centre for Heart Research, Toronto, Ontario, Canada M5G 1L7.
10
Toronto General Hospital, University Health Network, Toronto, Ontario, Canada M5G 1L7.
11
Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada M5G 2M9.

Abstract

Membrane proteins are crucial to heart function and development. Here we combine cationic silica-bead coating with shotgun proteomics to enrich for and identify plasma membrane-associated proteins from primary mouse neonatal and human fetal ventricular cardiomyocytes. We identify Tmem65 as a cardiac-enriched, intercalated disc protein that increases during development in both mouse and human hearts. Functional analysis of Tmem65 both in vitro using lentiviral shRNA-mediated knockdown in mouse cardiomyocytes and in vivo using morpholino-based knockdown in zebrafish show marked alterations in gap junction function and cardiac morphology. Molecular analyses suggest that Tmem65 interaction with connexin 43 (Cx43) is required for correct localization of Cx43 to the intercalated disc, since Tmem65 deletion results in marked internalization of Cx43, a shorter half-life through increased degradation, and loss of Cx43 function. Our data demonstrate that the membrane protein Tmem65 is an intercalated disc protein that interacts with and functionally regulates ventricular Cx43.

PMID:
26403541
DOI:
10.1038/ncomms9391
[Indexed for MEDLINE]

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