von Willebrand factor type D domain mutant of SVS-1/SUSD2, vWD^m, induces apoptosis in HeLa cells

SVS-1/SUSD2 is a novel gene, which inhibits growth and reverses tumorigenic phenotypes of cancer cells in vitro. Here we report identification of a mutant of SVS-1, designated SVS-1-vWD^m, in which conserved amino acids GLLG at positions 591–594 in von Willebrand factor type D (vWD) domain are replaced by AAAA. As observed by laser confocal microscope, intracellular localization of the mutant protein has changed such that both the N-terminus and the C-terminus of SVS-1-vWD^m were localized in the inner surface of the plasma membrane, whereas the N-terminus of SVS-1 was localized in the outer surface of the plasma membrane. Additionally, SVS-1-vWD^m was processed much less efficiently and in a slightly different manner. In in vitro studies, adenovirus-mediated transduction of the SVS-1-vWD^mgene induced growth suppression of HeLa cells in a dose-dependent manner, as the wild-type gene and inhibition of anchorage-independent growth. Of great interest is the finding that the mutant protein, vWD^m, but not the wild-type one induced apoptosis, as observed by nuclear as well as DNA fragmentation. Activation of caspase-3 and -9, but not caspase-8 or -12, was also demonstrated in vWD^m-expressing cells. An inhibition of Akt phosphorylation, a major survival signaling component, also occurred in vWD^m-expressing HeLa cells. Together these data suggest that vWD^m induces apoptosis by inactivation of survival signaling component Akt and activation of caspase cascade (mitochondrial pathway) in HeLa cells. We propose SVS-1-vWD^mas an alternative gene for use in developing new therapeutic strategies for the treatment of cancer. (Cancer Sci 2007; 98: 909–915)