Internalized gap junctions are degraded by autophagy.
Direct intercellular communication mediated by gap junctions (GJs) is a hallmark of normal cell and tissue physiology. In addition, GJs significantly contribute to physical cell-cell adhesion. Clearly, these cellular functions require precise modulation. Typically, GJs represent arrays of hundreds to thousands of densely packed channels, each one assembled from two half-channels (connexons), that dock head-on in the extracellular space to form the channel arrays that link neighboring cells together. Interestingly, docked GJ channels cannot be separated into connexons under physiological conditions, posing potential challenges to GJ channel renewal and physical cell-cell separation. We described previously that cells continuously-and effectively after treatment with natural inflammatory mediators-internalize their GJs in an endo-/exocytosis process that utilizes clathrin-mediated endocytosis components, thus enabling these critical cellular functions. GJ internalization generates characteristic cytoplasmic double-membrane vesicles, described and termed earlier annular GJs (AGJs) or connexosomes. Here, using expression of the major fluorescent-tagged GJ protein, connexin 43 (Cx43-GFP/YFP/mApple) in HeLa cells, analysis of endogenously expressed Cx43, ultrastructural analyses, confocal colocalization microscopy, pharmacological and molecular biological RNAi approaches depleting cells of key-autophagic proteins, we provide compelling evidence that GJs, following internalization, are degraded by autophagy. The ubiquitin-binding protein p62/sequestosome 1 was identified in targeting internalized GJs to autophagic degradation. While previous studies identified proteasomal and endo-/lysosomal pathways in Cx43 and GJ degradation, our study provides novel molecular and mechanistic insights into an alternative GJ degradation pathway. Its recent link to health and disease lends additional importance to this GJ degradation mechanism and to autophagy in general.
Department of Biological Sciences, Lehigh University, Bethlehem, PA, USA.
John T
JT
Fong
Rachael M
RM
Kells
Anna M
AM
Gumpert
Jutta Y
JY
Marzillier
Michael W
MW
Davidson
Matthias M
MM
Falk
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D001343:Autophagy; D015415:Biomarkers / Q000378:metabolism; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D018031:Connexin 43 / Q000378:metabolism; D004705:Endocytosis; D005453:Fluorescence; D017629:Gap Junctions / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008954:Models, Biological; D010588:Phagosomes / Q000378:metabolism / Q000648:ultrastructure; D021381:Protein Transport; D034622:RNA Interference; D000071456:Sequestosome-1 Protein
D048868:Adaptor Proteins, Signal Transducing; D015415:Biomarkers; D018031:Connexin 43; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein