Uncleaved BAP31 in association with A4 protein at the endoplasmic reticulum is an inhibitor of Fas-initiated release of cytochrome c from mitochondria.
BAP31 is a polytopic integral protein of the endoplasmic reticulum membrane and, like BID, is a preferred substrate of caspase-8. Upon Fas/CD95 stimulation, BAP31 is cleaved within its cytosolic domain, generating proapoptotic p20 BAP31. In human KB epithelial cells expressing the caspase-resistant mutant crBAP31, Fas stimulation resulted in cleavage of BID and insertion of BAX into mitochondrial membrane, but subsequent oligomerization of BAX and BAK, egress of cytochrome c to the cytosol, and apoptosis were impaired. Bap31-null mouse cells expressing crBAP31 cannot generate the endogenous p20 BAP31 cleavage product, yet crBAP31 conferred resistance to cellular condensation and cytochrome c release in response to activation of ectopic FKBPcasp8 by FK1012z. Full-length BAP31, therefore, is a direct inhibitor of these caspase-8-initiated events, acting independently of its ability to sequester p20, with which it interacts. Employing a novel split ubiquitin yeast two-hybrid screen for BAP31-interacting membrane proteins, the putative ion channel protein of the endoplasmic reticulum, A4, was detected and identified as a constitutive binding partner of BAP31 in human cells. Ectopic A4 that was introduced into A4-deficient cells cooperated with crBAP31 to resist Fas-induced egress of cytochrome c from mitochondria and cytoplasmic apoptosis.
Wang
Bing
B
Department of Biochemistry, McGill University, Montreal, Quebec H3G 1Y, Canada.
Nguyen
Mai
M
Breckenridge
David G
DG
Stojanovic
Marina
M
Clemons
Paul A
PA
Kuppig
Stephan
S
Shore
Gordon C
GC
D000251:Adenosine Triphosphatases / Q000378:metabolism; D017209:Apoptosis; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism*; D003593:Cytoplasm / Q000473:pathology; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D062792:MARVEL Domain-Containing Proteins; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism* / Q000473:pathology; D008954:Models, Biological; D011233:Precipitin Tests; D011485:Protein Binding; D011510:Proteolipids; D013997:Time Factors; D014162:Transfection; D020798:Two-Hybrid System Techniques; D025801:Ubiquitin / Q000378:metabolism; D019014:fas Receptor / Q000378:metabolism
C091247:BCAP31 protein, human; C489642:Bcap31 protein, mouse; D003574:Cytochrome c Group; D062792:MARVEL Domain-Containing Proteins; D008565:Membrane Proteins; C094765:PLP2 protein, human; D011510:Proteolipids; D025801:Ubiquitin; D019014:fas Receptor; D003513:Cycloheximide; C505530:CASP8 protein, human; C506433:CASP9 protein, human; C505526:Casp8 protein, mouse; C506434:Casp9 protein, mouse; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D000251:Adenosine Triphosphatases