The ASPP2 tumor suppressor is a pro-apoptotic member of a family of p53 binding proteins. It functions in part by selectively enhancing p53 transcriptional activation of pro-apoptotic target genes. Although the ASPP2 C-terminal domain binds the p53 core domain, little is known about the physical and functional interactions of other proteins to the highly structured ASPP2 N-terminus. Moreover, mounting evidence from our lab and others demonstrate that ASPP2 also mediates non-apoptotic p53-independent functions. However, the molecular mechanisms remain poorly understood. Previous reports have found that ASPP2 can localize to the cytoplasm and have identified a potential Ras-association domain in the ASPP2 N-terminus. Therefore, we wondered if novel ASPP2 functions could be mediated by ASPP2 N-terminal interaction with cytoplasmic Ras. Using co-immunoprecipitation and immunofluorescence microscopy, we identified ASPP2 in a complex with active Ras on the plasma membrane. Moreover, we also found that this interaction required the ASPP2 N-terminus since BBP, a naturally occurring alternatively spliced ASPP2 isoform lacking the N-terminus, showed markedly reduced ability to bind Ras. To determine if ASPP2 could modulate Ras function, we ectopically expressed ASPP2 in cells and found enhancement of Ras activation and subsequent stimulation of ERK phosphorylation. Conversely, ASPP2 knockdown by siRNA drastically reduced ERK phosphorylation and activation by EGF. Although it is oncogenic in tumor cells, constitutive Ras activation in normal cells leads to premature senescence in a Raf-MEK-ERK activation dependent manner. To explore if ASPP2 enhanced senescence, we knocked down ASPP2 expression with siRNA and found that senescence induced by RasV12 lentiviral infection in normal human fibroblasts was significantly attenuated. This was similar to the attenuation of senescence achieved by the MEK inhibitor PD98059. Together, our results demonstrate a novel function of the ASPP2 N-terminus in promoting cellular senescence via Ras-interaction and stimulation of Raf-MEK-ERK activity. These findings suggest a novel mechanism by which ASPP2-induced senescence may mediate its tumor suppression ability, and opens new avenues for investigation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-128. doi:10.1158/1538-7445.AM2011-LB-128

©2011 American Association for Cancer Research
2011