Alternative splicing (AS) is a mechanism that generates protein diversity from a limited number of genes. Through the differential assortment of exons, AS allows the production of different mRNAs from most human genes, thereby increasing genome complexity. Splicing abnormalities have profound impact on several human diseases, including cancer. A growing body of evidence suggests that the splicing machinery is an important target for misregulation in cancer, and that altered expression of several splicing factors correlates with cancer development and progression. Sam68 is an RNA-binding protein involved in alternative splicing regulation. Sam68 has a pro-oncogenic function and it is frequently up-regulated in human cancer, including prostate cancer (PCa), wherein Sam68 supports cell proliferation, migration and survival. In the present work, we have analysed by mass spectrometry proteins that co-immunoprecipitate with endogenous Sam68 from PCa cells and we identified p100/SND1 as a novel Sam68-interacting protein. P100/SND1 is a transcriptional co-activator, which also interacts with components of the spliceosome, suggesting a role in coupling transcription and splicing. We found that p100/SND1 is over-expressed in PCa cells like Sam68. Sedimentation assays and co-immunoprecipitation experiments showed that p100/SND1 and Sam68 form a complex. Moreover, the functional relevance of this interaction was demonstrated by splicing assays using a CD44 minigene, a well established target of Sam68 splicing activity. Up-regulation of p100/SND1 enhances Sam68-dependent exon v5 inclusion in CD44 mRNA, which is known to correlate with increased proliferation and invasiveness of cancer cells. In line with these observations, depletion of p100/SND1 by RNAi reduced migration of PCa cells, similarly to what observed with Sam68 depletion. These results strongly suggest that p100/SND1 is a novel positive regulator of Sam68 splicing activity that promotes PCa cell growth and survival.
The transcriptional regulator p100SND1 interacts Sam68 and regulates CD44 alternative splicing in prostate cancer cells.
Paronetto MP;
2010-01-01
Abstract
Alternative splicing (AS) is a mechanism that generates protein diversity from a limited number of genes. Through the differential assortment of exons, AS allows the production of different mRNAs from most human genes, thereby increasing genome complexity. Splicing abnormalities have profound impact on several human diseases, including cancer. A growing body of evidence suggests that the splicing machinery is an important target for misregulation in cancer, and that altered expression of several splicing factors correlates with cancer development and progression. Sam68 is an RNA-binding protein involved in alternative splicing regulation. Sam68 has a pro-oncogenic function and it is frequently up-regulated in human cancer, including prostate cancer (PCa), wherein Sam68 supports cell proliferation, migration and survival. In the present work, we have analysed by mass spectrometry proteins that co-immunoprecipitate with endogenous Sam68 from PCa cells and we identified p100/SND1 as a novel Sam68-interacting protein. P100/SND1 is a transcriptional co-activator, which also interacts with components of the spliceosome, suggesting a role in coupling transcription and splicing. We found that p100/SND1 is over-expressed in PCa cells like Sam68. Sedimentation assays and co-immunoprecipitation experiments showed that p100/SND1 and Sam68 form a complex. Moreover, the functional relevance of this interaction was demonstrated by splicing assays using a CD44 minigene, a well established target of Sam68 splicing activity. Up-regulation of p100/SND1 enhances Sam68-dependent exon v5 inclusion in CD44 mRNA, which is known to correlate with increased proliferation and invasiveness of cancer cells. In line with these observations, depletion of p100/SND1 by RNAi reduced migration of PCa cells, similarly to what observed with Sam68 depletion. These results strongly suggest that p100/SND1 is a novel positive regulator of Sam68 splicing activity that promotes PCa cell growth and survival.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
