RIP3 (receptor-interacting protein 3) is a serine/threonine kinase that promotes apoptosis in various cell types. The C-terminal domain of RIP3 is critical for its apoptosis induction. In this study, we showed that a truncated form of RIP3 (tRIP3) containing only the unique C-terminal region (aa 224-518) induced significant apoptosis in human hepatocellular carcinoma cells QGY-7703. A FADD-dominant negative (FADD-DN) was shown to significantly block apoptosis induced by tRIP3. In contrast, the RIP3 dominant negative (RIP3-DN) was found unable to block FADD-induced apoptosis. Thus, we conclude that tRIP3 may function upstream of FADD to induce apoptosis in TNFR-1 signaling pathway. Additionally, sequence alignments of RIP3 with other death domain (DD)-containing proteins revealed that amino acids Leu 477 and Leu 488 in RIP3 are highly conserved, nonetheless, neither mutational change at Leu 477 nor at Leu 488 confers obvious effect on cell death, indicating that these two amino acids might not be critical for its pro-apoptotic activity as expected. (c) 2006 Elsevier Inc. All rights reserved.