We previously identified and characterized a murine BTB-containing protein, CIBZ (ZBTB38 in human), that interacts with CtBP and binds to methylated CpGs.However, its physiological function remained unknown.As CtBP is reportedly involved in p53-independent programmed cell death, we examine here whether CIBZ is associated with apoptosis.We found that CIBZ was highly expressed in proliferating C2C12 cells, but that its expression levels decreased upon induction of apoptosis by serum starvation.Knockdown of CIBZ by siRNA in C2C 12 cells induced apoptosis, as determined by an increase of annexin V/PI labeling, activation of caspase-3, and cleavage of PARP.CIBZ inhibition also activated caspase-7 and caspase-9, suggesting that CIBZ-associated apoptosis occurs through the mitochondrial pathway.Notably, knockdown of CIBZ in p53-/-MEF cells also activated caspase-3 and cleavage of PARP, indicating that CIBZ-associated apoptosis is mediated by a p53-independent pathway; however, since both common and distinct targets are regulated by CIBZ-and CtBP-associated apoptosis, we conclude that more than one pathway is involved.Finally, using mutagenesis and an in vitro caspase cleavage assay, we show that CIBZ is a novel substrate of caspase-3, and identify two caspase-3 recognition sites.These findings indicate, collectively, that CIBZ plays an important role by participating in the negative regulation of apoptosis in murine cells.