The role of 5' splice sites on nuclear mechanisms of regulated gene expression in retroviruses and cellular genes

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The generation of messenger RNA from a DNA template is a complex, stepwise process. mRNAs acquire different features including a cap-structure and a poly A tail. In addition, introns are spliced out to build a continuous open reading frame for protein synthesis. Finally the RNA has to leave the nucleus via active RNA export through the nuclear pore. One of the reasons for all these steps is the implementation of “check-points” to monitor the functionality of the mRNA. These surveillance mechanisms are essential to protect the cell from defective RNAs. We intend to study two examples where cellular quality control plays a central role. One is a point mutation within the 3’UTR of the p14 gene leading to a novel primary immunodeficiency syndrome. This mutation may create a cryptic 5’ splice site (ss) causing reduced mRNA levels. Binding of the 5’ss by the spliceosomal component U1 snRNP seems to interfere with normal 3’ end processing of the p14 mRNA and leads to rapid degradation, possibly by the nuclear exosome. In the second case, we plan to elucidate how murine leukemia virus (MLV), the paradigm of simple gammaretroviruses, regulates the export of its introncontaining, genomic RNA. The presence of unused splice sites on this RNA would normally prevent export as another means of quality control. The acquired knowledge will contribute to improving gene transfer vectors. Moreover, the identification of the underlying mechanism of the p14 immunodeficiency will provide valuable insights into gene regulation in general, and may also lead to a treatment strategy for the affected patients.