Poster abstracts

Poster number 124 submitted by Whitney Smith-Kinnaman

RNA polymerase II and 3’ end processing machinery in S. cerevisiae

Whitney R. Smith-Kinnaman (Department of Biochemistry and Molecular Biology, IU School of Medicine), Melanie J. Fox (Department of Biochemistry and Molecular Biology, IU School of Medicine), Megan A. Zimmerly (Department of Biochemistry and Molecular Biology, IU School of Medicine), Amber L. Mosley (Department of Biochemistry and Molecular Biology & Center for Computational Biology and Bioinformatics, IU School of Medicine)

Abstract:
RNA Polymerase II (RNAPII) transcribes mRNA and small non coding RNA. The C-Terminal domain (CTD) of Rbp1 plays a large role in the events and timing of transcription. The repetitive sequence YSPTSPS (26 repeats in S.cerevisiae) and its phosphorylation pattern as RNAPII moves across the gene is the main orchestrator of when proteins are recruited to the CTD and in turn controls the timing of transcription events such as termination and 3’ end processing. Messenger RNA cleavage and polyadenylation are two important processes that are coupled with RNAPII transcription termination in eukaryotes. In yeast the complexes implicated in these tasks include: cleavage factor IA (CF IA), Hrp1 (cleavage factor IB), the Nrd1-Nab3-Sen1 (NNS) complex, the cleavage and polyadenylation factor (CPF), and the nuclear exosome. Although the major subunits of each complex have been defined by previous studies, the degree of crosstalk between these complexes and RNAPII has not been determined. By utilizing affinity purifications coupled with mass spectrometry (MudPIT) and quantitative proteomics analysis, we will determine the degree of involvement RNAPII plays in 3’ end processing complex formation, the recruitment dynamics of subunits involved, , and any post translation modifications that may occur. We will present evidence showing that many of the 3’ end processing complexes interact with RNAPII. However, we observe limited co-purification of subunits from different 3’end processing complexes suggesting that RNAPII and/or the nascent RNA likely provide the framework for 3’ end processing complex crosstalk. In addition, we have identified a number of novel candidates associated with specific mRNA processing complexes that we plan to characterize further.

Keywords: RNA Polymerase II, 3 end processing, mRNA