Dr. Lucas Horan
I think the “RNA World Hypothesis” is the most reasonable explanation for the origin of life on this planet. If “life” was once primarily composed of RNA molecules, then the functionality of RNA in biology is undoubtedly under-appreciated. Alternative splicing of mRNA is a great example of how the most subtle of changes in RNA sequence can amplify expression diversity from a single gene.
My current research uses the P-element splicing silencer complex as a model system to investigate mechanisms of alternative-splicing. Earlier studies on the regulation of the Drosophila P-element transposase revealed one of the first examples of tissue-specific alternative splicing. In somatic cells, splicing of the P-element third intron (IVS3) is repressed by formation of a splicing silencer complex on the 5’ exon RNA. This creates an mRNA isoform that retains the intron and translates into a P-element ORF with an early termination codon. Biochemical and genetic experiments have previously demonstrated that the hnRNP proteins PSI and hrp48 silence IVS3 splicing by interacting with U1snRNP near the 5’ splice site. A recent RNAi screen identified 3 additional proteins that are functionally required for efficient silencing of the IVS3 splice site.
Our first aim is to elucidate the molecular mechanism of these proteins in IVS3 splicing silencing. To address this aim I am reconstituting the silencing complex for biochemical and structural analysis. Our second aim is to better understand the role of these proteins in alternative splicing within endogenous genes. To address this aim we are coupling next-generation RNA sequencing with immuno-precipitation to identify the binding sites of these silencing proteins within the transcriptome.