c-IAP1 is cleaved by caspases to produce a proapoptotic C-terminal fragment

Although:human c-IAP1 and c-IAP2 have been reported to-possess antiapoptoti c activity against a variety of stimuli in several mammalian cell types, we observed that full-length c-IAP1 and c-IAP2 failed to protect cells from a poptosis induced by Bax overexpression, tumor necrosis factor alpha treatme nt or Sindbis virus infection. However, deletion of the C-terminal RING dom ains:of c-IAP1 and c-IAP2 restored antiapoptotic activity, indicating that this region negatively regulates the antiapoptotic function of the N-termin al BIR domain. This finding is consistent with the observation by others th at the spacer region and RING domain of c-IAP1 functions as an E3 ligase, p romoting autoubiquitination and degradation of c-IAP1. In addition, we foun d that c-IAP1 is cleaved during apoptosis to 52- and 35-kDa fragments. Both fragments contain the C-terminal end of c-IAP1 including the-RING finger. In vitro cleavage of c-IAP1 with apoptotic cell extracts or with purified r ecombinant caspase-3 produced similar fragments. Furthermore, transfection of cells with the spacer-RING domain alone Suppressed the antiapoptotic fun ction of the N-terminal BIR domain of c-IAP1 and induced apoptosis, Optimal death-inducing activity of the spacer-RING required both the spacer region and the zinc-binding RING domain of c-IAP1 but did not require the caspase recruitment domain located within the spacer region, To the contrary, dele tion of the caspase recruitment domain increased proapoptotic activity, app arently by stabilizing the C-terminal fragment.