Abstract
The eukaryotic spliceosome is composed of five small nuclear ribonucleoproteins (snRNPs) and many other associated protein splicing factors. The human U2 snRNP‐associated protein, SF3B1, allows the U2 snRNP to associate with and define the branchpoint region of precursor‐messenger RNA (pre‐mRNA). Recent evidence has shown that mutations to SF3B1 are linked to a variety of different myelodysplastic syndromes (MDS). These mutations are found in regions of high conservation between SF3B1 and the yeast homolog, Hsh155p. We have constructed a genetic system in S. cerevisiae to interrogate the effects of these mutations on splicing and cell proliferation. We used plasmid shuffling to introduce Hsh155p mutant proteins into a HSH155Δ yeast strain and growth assays to assess the impact of the mutations on cell proliferation. Several of the mutations surveyed thus far (H331D, K335N, P369E, D450G) have no obvious effect on yeast growth despite being correlated with MDS in humans. This may imply that the effects of these mutations are subtle and do not allow splicing to become limiting for yeast growth. To further understand the effects of these mutations, a Reverse Transcription‐Polymerase Chain Reaction (RT‐PCR) analysis will be used to compare pre‐mRNA and mRNA levels of several transcripts from Hsh155p mutant strains relative to the WT. The results from these experiments will be critical for establishing yeast as a model system for studying MDS. This research has been supported by startup funds from the U. Wisconsin‐Madison, WARF, and the Department of Biochemistry.
Published Version
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