Increased Expression of SAM68 in SV40 Transformed Human Diploid Fibroblast Cells and the Effect on Downstream Splicing Targets
Student Research Paper
Dr. Jane Cavender
Alternative splicing has emerged as a major player in cancer initiation and progression. The altered splicing factor profiles and the effect on their subsequent targets have been reported in several cancers. Comparing tumor samples and tumor cell lines to build protein profiles of specific isoforms has elucidated common alterations that may indicate the important drivers of tumorigenesis. However, when using primary tumors and tumor cell lines there is often not a matched control to directly compare the results. But by employing the simian virus 40 (SV40) DNA tumor virus model, studies can be designed to investigate the isoform profiles before and after cellular transformation. For this study, human diploid fibroblasts immortalized with telomerase (HDF(tert)) were stably transfected with plasmid encoding the early region SV40 (HDF(tert+T)). The two created clones were compared to the parental line through RT-PCR and Western Blotting. It was found that the level of T-antigen was directly correlated to increased levels of the splicing factor SAM68. To elucidate the possible role of SAM68 in viral transformation, downstream splicing targets were also assessed using RT-PCR and Western Blotting. The results show that the SAM68 target, SRSF1, has an increased expression of only the specific SRSF1-208 isoform in the transformed lines. To further investigate SRSF1 expression and modifications, an immunofluorescence assay was performed. Preliminary results suggest that SRSF1 is located in the cytoplasm and that there are no phosphorylation pattern differences between SRSF1 isoforms, but studies are still ongoing to solidify this finding. Overall, this data suggests that increased SAM68 expression could contribute to the aggressive growth of SV40 T-antigen transformed human diploid fibroblasts, and this could be through the preferential splicing of the SRSF1-208 isoform. These findings are significant as they could indicate that SAM68 could potentially be a chemotherapeutic target for virally-induced cancers, and they expand the literature on SAM68 as increased levels of SAM68 and the subsequent downstream altered proteins have been implicated as a driver of tumorigenesis in other cancers.
Caroland, Kailey, "Increased Expression of SAM68 in SV40 Transformed Human Diploid Fibroblast Cells and the Effect on Downstream Splicing Targets" (2022). Biology: Student Scholarship & Creative Works. 20.
Honors Senior Thesis; Honors in the Discipline