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

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


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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Chromatography, High Pressure Liquid
  • Connexin 43 / metabolism*
  • Fluorescent Antibody Technique
  • Gap Junctions / metabolism*
  • Gap Junctions / ultrastructure
  • Gene Knockdown Techniques
  • Heart Conduction System / metabolism*
  • Heart Conduction System / physiology
  • Heart Ventricles / metabolism*
  • In Vitro Techniques
  • Membrane Proteins / genetics*
  • Membrane Proteins / metabolism
  • Mice
  • Microscopy, Electron
  • Myocytes, Cardiac / metabolism*
  • Myocytes, Cardiac / physiology
  • Myocytes, Cardiac / ultrastructure
  • Proteomics
  • Silicon Dioxide
  • Zebrafish
  • Zebrafish Proteins / genetics*
  • Zebrafish Proteins / metabolism


  • Connexin 43
  • Membrane Proteins
  • Tmem65 protein, mouse
  • Zebrafish Proteins
  • tmem65 protein, zebrafish
  • Silicon Dioxide