Conclusion
The procaspase-8 Q482H mutation in AML patients abolishes caspase-8-mediated apoptosis by impairing procaspase-8 dimerization.
Methods
Polymerase chain reaction (PCR) amplification and Sanger sequencing of the procaspase-8 gene (CASP8) were performed. Apoptotic rates were analyzed with Annexin V-FITC staining in cells expressing wild-type (WT) procaspase-8, the Q482H or C360S mutant, or control vector after treatment with or without tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Western blot analysis was performed to detect activation of procaspase-8 and downstream apoptotic signaling pathway components in those cells. The Co-immunoprecipitation (Co-IP) assays were performed to detect interaction between WT and mutant procaspase-8 proteins.
Results
AML patients carrying the Q482H mutation were likely to develop chemotherapy resistance. Similar to C360S, The Q482H mutation abolished caspase-8-mediated apoptotic signaling and inhibited TRAIL-induced apoptosis. The Q482H mutation impaired procaspase-8 dimerization, thus preventing the self-activation of procaspase-8.