Caspase mediated cleavage of p130Cas is essential during apoptosis of intestinal epithelial cells

Multidomain adaptor p130Cas (Crk‐associated substrate) has been implicated as a key signaling protein in apoptosis. Inhibition of ornithine decarboxylase by alpha‐difluromethylornithine (DFMO) confers resistance to apoptosis in intestinal epithelial cells. DFMO induced p130Cas (Tyr‐410) phosphorylation, which was blocked by the addition of exogenous putrescine. Sustained time dependent p130Cas phosphorylation in polyamine depleted cells correlated with a decrease in TNF‐α‐induced apoptosis. Progression of apoptosis in IEC‐6 cells led to caspase‐3 dependent cleavage of p130Cas protein and generated a 31‐Kda fragment. Conversely, decreased caspase‐3 activation after polyamine depletion significantly attenuated p130Cas cleavage. The cleaved 31‐Kda protein fragment was generated in the cytoplasm and translocated to the nucleus. The E47 family of transciption factor regulates p21 expression and is crucially involved in cell survival. E47 associates with the 31‐Kda fragment after apoptosis. Polyamine depletion prevented p130Cas degradation and association with E47 during apoptosis. Increased p21 waf/Kip protein was mis‐localized in the cytoplasm in polyamine depleted cells. Inhibition of Src using a dominant negative construct or a pharmacologic inhibitor PP2, decreased p130Cas phosphorylation in control and polyamine depleted cells, increased p130Cas cleavage, and sensitized cells to TNF‐α‐induced apoptosis. These data suggest that polyamine depletion increases p130Cas phosphorylation and inhibits its degradation during apoptosis, and thereby promotes survival. Supported by NIDDK grant DK‐16505.