Lysosomal and proteasomal degradation play distinct roles in the life cycle of Cx43 in gap junctional intercellular communication-deficient and -competent breast tumor cells.
The present study was designed to determine the specific roles played by lysosomes and proteasomes in the degradation of Cx43 in both gap junctional intercellular communication-deficient MDA-MB-231 and -competent BICR-M1Rk cells. In MDA-MB-231 cells, immunolocalization and brefeldin A protein transport blocking studies revealed that there was a propensity for newly synthesized Cx43 to be transported to lysosomes. On the other hand, light and electron microscopic analysis of BICR-M1Rk cells showed that Cx43 gap junctions were prevalent with a subpopulation of intracellular Cx43 localized to lysosomes. In both cell types, Western blots revealed a notable increase in total cellular Cx43 in response to lysosome inhibitors. Interestingly, lactacystin inhibition of proteosomal degradation in MDA-MB-231 cells resulted in a marked increase in phosphorylated Cx43 at the expense of non-phosphorylated Cx43, and this change corresponded with an increase in "oversized" gap junction plaques. In BICR-M1Rk cells, lactacystin treatment partially prevented the BFA-induced loss of gap junctions. Together, our data suggests that lysosomes play a key role in not only degrading internalized gap junction in BICR-M1Rk cells but also in degrading Cx43 delivered from early secretory compartments to lysosomes in MDA-MB-231 cells. Overall proteasomal degradation regulates the stability of phosphorylated Cx43 and appears to promote the internalization of Cx43 from the cell surface.
Qin
Hong
H
Department of Anatomy and Cell Biology, University of Western Ontario, London, Ontario N6A 5C1, Canada.
Shao
Qing
Q
Igdoura
Suleiman A
SA
Alaoui-Jamali
Moulay A
MA
Laird
Dale W
DW
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology*; D020126:Brefeldin A / Q000494:pharmacology; D002450:Cell Communication; D002462:Cell Membrane / Q000378:metabolism; D018031:Connexin 43 / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D017629:Gap Junctions / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008247:Lysosomes / Q000378:metabolism*; D015674:Mammary Neoplasms, Animal / Q000378:metabolism; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D009097:Multienzyme Complexes / Q000378:metabolism*; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D021381:Protein Transport; D051381:Rats; D015398:Signal Transduction; D013997:Time Factors; D014407:Tumor Cells, Cultured
D018031:Connexin 43; D009097:Multienzyme Complexes; D011500:Protein Synthesis Inhibitors; C067713:lactacystin; D020126:Brefeldin A; D003513:Cycloheximide; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine