The Role of Protein Arginine Methylation in the Co-transcriptional Recruitment of pre-mRNA Splicing Factors

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Intellectual Merit. In eukaryotes, proper gene expression is vital for the control of many cellular processes and responses. The initial step of gene expression requires the copying of the DNA into an intermediate molecule called the RNA via a process called transcription. During transcription, the unfinished RNA must simultaneously be processed by a multitude of specific proteins, including the splicing factors (which are members of a macromolecular protein complex called the spliceosome), in order to generate a final, finished product. As the RNA molecules are made and processed, they are packaged into a complex called messenger ribonucleoprotein (mRNP) particle and once finished, the entire mRNP is then exported out of the cell nucleus. To achieve and coordinate the coupling of these biological processes, the RNA-binding proteins must be recruited to the RNA in a very precise order during transcription. As a way to expand or to regulate the function of a protein, many RNA-binding proteins are modified chemically by specific enzymes. Arginine methylation is one such type of chemical modification that is found on many RNA binding proteins, and this modification is catalyzed by a family of enzymes known as protein arginine methyltransferases (PRMT). This enzyme family has been identified in species ranging from the budding yeast to humans, with the most conserved member of this family being PRMT1. In the budding yeast Saccharomyces cerevisiae, it was established that yeast PRMT1 (termed Hmt1) plays a functional role in promoting the recruitment of splicing factors during transcription, which is vital to mRNP formation. The goals of this project are to examine the mechanisms by which protein arginine methylation modulates the co-transcriptional recruitment of splicing factors in the simple eukaryotic model organism Saccharomyces cerevisiae. A combination of molecular biology, biochemistry, proteomic, and genomic methodologies will be used. Because of the high conservation between yeast Hmt1 and its homologs in higher eukaryotes, the information obtained from this project will provide insights that will likely be applicable to those organisms.Broader Impacts. This project will provide opportunities to train graduate students and post-doctoral fellows at University at Buffalo, which is a very diverse campus. The graduate students will receive training in the areas of molecular biology, biochemistry, and genomics. Undergraduate students will also be recruited to take part in various aspects of this project and these recruits will include members of the New York Collegiate Science & Technology Entry (CSTEP) program, an enrichment program for students who are economically disadvantaged or historically underrepresented. Since the project involves the use of molecular biology with genomics and computational analysis, the students will benefit from crossdisciplinary training. Travel funds will be used to allow students to present talks and posters at national scientific meetings. Findings from this work will be incorporated into an Advanced Molecular Biology laboratory course offered to the more senior undergraduates at University at Buffalo. The microarray data generated from this project will be made available to the public.