The BIR motifs mediate dominant interference and oligomerization of inhibitor of apoptosis Op-IAP.
The defining structural motif of the inhibitor of apoptosis (iap) protein family is the BIR (baculovirus iap repeat), a highly conserved zinc coordination domain of approximately 70 residues. Although the BIR is required for inhibitor-of-apoptosis (IAP) function, including caspase inhibition, its molecular role in antiapoptotic activity in vivo is unknown. To define the function of the BIRs, we investigated the activity of these structural motifs within Op-IAP, an efficient, virus-derived IAP. We report here that Op-IAP(1-216), a loss-of-function truncation which contains two BIRs but lacks the C-terminal RING motif, potently interfered with Op-IAP's capacity to block apoptosis induced by diverse stimuli. In contrast, Op-IAP(1-216) had no effect on apoptotic suppression by caspase inhibitor P35. Consistent with a mechanism of dominant inhibition that involves direct interaction between Op-IAP(1-216) and full-length Op-IAP, both proteins formed an immunoprecipitable complex in vivo. Op-IAP also self-associated. In contrast, the RING motif-containing truncation Op-IAP(183-268) failed to interact with or interfere with Op-IAP function. Substitution of conserved residues within BIR 2 caused loss of dominant inhibition by Op-IAP(1-216) and coincided with loss of interaction with Op-IAP. Thus, residues encompassing the BIRs mediate dominant inhibition and oligomerization of Op-IAP. Consistent with dominant interference by interaction with an endogenous cellular IAP, Op-IAP(1-216) also lowered the survival threshold of cultured insect cells. Taken together, these data suggest a new model wherein the antiapoptotic function of IAP requires homo-oligomerization, which in turn mediates specific interactions with cellular apoptotic effectors.
Hozak
R R
RR
Department of Biochemistry and Institute for Molecular Virology, Graduate School and College of Agricultural and Life Sciences, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
Manji
G A
GA
Friesen
P D
PD
D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D019281:Dimerization; D005786:Gene Expression Regulation; D051034:Inhibitor of Apoptosis Proteins; D012091:Repetitive Sequences, Nucleic Acid; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism*
D051034:Inhibitor of Apoptosis Proteins; D014764:Viral Proteins; C086430:inhibitor of apoptosis, Nucleopolyhedrovirus