Limiting transport steps and novel interactions of Connexin-43 along the secretory pathway.
Connexins are four-transmembrane-domain proteins expressed in all vertebrates which form permeable gap junction channels that connect cells. Here, we analysed Connexin-43 (Cx43) transport to the plasma membrane and studied the effects of small GTPases acting along the secretory pathway. We show that both GTP- and GDP-restricted Sar1 prevents exit of Cx43 from the endoplasmic reticulum (ER), but only GTP-restricted Sar1 arrests Cx43 in COP II-coated ER exit sites and accumulates 14-3-3 proteins in the ER fraction. FRET-FLIM data confirm that already in ER exit sites Cx43 exists in oligomeric form, suggesting an in vivo role for 14-3-3 in Cx43 oligomerization. Exit of Cx43 from the ER can be blocked by other factors--such as expression of the beta subunit of the COP I coat or p50/dynamitin that acts on the microtubule-based dynein motor complex. GTP-restricted Arf1 blocks Cx43 in the Golgi. Lastly, we show that GTP-restricted Arf6 removes Cx43 gap junction plaques from the cell-cell interface and targets them to degradation. These data provide a molecular explanation of how small GTPases act to regulate Cx43 transport through the secretory pathway, facilitating or abolishing cell-cell communication through gap junctions.
Majoul
Irina V
IV
Institute of Biology, Center for Structural and Cell Biology in Medicine, University of Lübeck, Ratzeburger Allee 160, 23538, Lübeck, Germany. irina.majoul@bio.uni-luebeck.de
Onichtchouk
Daria
D
Butkevich
Eugenia
E
Wenzel
Dirk
D
Chailakhyan
Levon M
LM
Duden
Rainer
R
D048948:14-3-3 Proteins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002522:Chlorocebus aethiops; D018031:Connexin 43 / Q000502:physiology*; D020285:Cryoelectron Microscopy; D004721:Endoplasmic Reticulum / Q000502:physiology / Q000648:ultrastructure; D036681:Fluorescence Recovery After Photobleaching; D017629:Gap Junctions / Q000502:physiology* / Q000648:ultrastructure; D006056:Golgi Apparatus / Q000502:physiology; D051379:Mice; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D021381:Protein Transport / Q000502:physiology; D055571:Secretory Pathway; D014709:Vero Cells
D048948:14-3-3 Proteins; D018031:Connexin 43