Set2 and Prp28 function in forward and reverse coupling of transcription and splicing
Kress, Tracy L.
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There is a series of steps that must take place for proper gene expression to occur. Two of those steps, transcription and splicing, have been shown to be coordinated. Although not much is known about the mechanism by which these two steps are connected, research has shown that the spliceosome, which is comprised of small nucleoprotein complexes (snRNPs,) loads onto the pre-mRNA as the pre-mRNA strand is created by RNA polymerase II. In order to determine the affect that transcription and splicing have on each other, we performed genetic and biochemical studies using Saccharomyces cerevisiae. Specifically, we looked for interactions between Set2, a histone methyltransferase responsible for methylated lysine 36 on histone 3 (H3K36) to regulate transcription, and Prp28, a helicase protein that facilitates the exchange of the U1 snRNP for the triple snRNP. Genetic interaction studies showed that Set2 and Prp28 negatively interact, leading to reduced fitness. Additionally, analysis by qRT-PCR found deletion of SET2 reduces splicing and that there was a greater accumulation of pre-mRNA in the set2D prp28-1 double mutant strains, consistent with our genetic results. Taken together, these results suggest that transcription influences the splicing of pre-mRNA. To test the converse, we monitored growth of yeast splicing mutants on a drug, 6-azauracil (6AU) , that limits the amount of rGTP, whereby slowing transcription. Wildtype yeast display impaired growth on 6AU and mutation of Prp28 further impaired growth, implicating Prp 28 in promoting transcription. Furthermore, we show a stalling of RNA polymerase II in cells where Prp 28 is mutated. Thus, our data suggest that splicing has an impact on the rate of transcription and that Set2 and Prp28 may work together to coordinate steps in gene expression.
Department of Biology
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