Talk abstracts

Talk on Saturday 09:20-09:40am submitted by Ravi Singh

Molecular mechanism of genotoxic stress induced alternative splicing of MDM2

Ravi K Singh (Center for Childhood Cancer,Childrens Research Institute and department of pediatrics, The Ohio State University, Columbus, OH.), Dawn S Chandler (Center for Childhood Cancer,Childrens Research Institute and department of pediatrics, The Ohio State University, Columbus, OH.)

Abstract:
MDM2, murine double-minute 2, and its related family member MDM4 are important for regulating the levels and activity of tumor suppressor protein, p53. Expression of alternatively spliced forms of these p53 regulators has been associated with various cancers. We have identified alternatively spliced forms of MDM2 and MDM4, which are induced after certain kinds of genotoxic stress.
Using MDM2 as a model, we are trying to understand the mechanism of alternative splicing of MDM2 and MDM4 in response to specific cell stress like UV exposure and cisplatin treatment. We have used minimal conserved genomic sequences from human and mice to create mini-gene constructs. These constructs contain the cis elements required to direct the alternative splicing in response to UV exposure in vivo. Using in vitro UV cross-linking with normal and cisplatin treated nuclear extract, we found factors that bind differently to our mini-gene constructs. Factors which bind to pre-mRNA of our mini-gene constructs in normal nuclear extract, but are displaced in the cisplatin treated nuclear extract may facilitate recognition of all MDM2 exons by the splicing machinery, and thus acts as a positive regulators of splicing. Likewise, we have identified specific proteins that bind to the MDM2 pre-mRNA only in the damaged extracts. These proteins are putative negative regulators of MDM2 splicing. These data underscore the dynamic nature of splicing machinery, which determines the splicing of not only MDM2 and MDM4, but also other genes. The UV-induced splicing machinery results from events initiated in response to stress and these RNA binding proteins may be the effectors of the damage-induced spliceome.
As opposed to a gene-specific approach to studying alternative splicing in cancer, this study will lead to a better understanding of how global signaling affects regulation of RNA processing factors involved in the generation of a damage-induced spliceome.

Keywords: MDM2, alternative splicing, spliceome