Poster abstracts
Poster number 9 submitted by Grace Eramo
Alternative splicing in the 5` untranslated region of mRNA as a mechanism of translational control
Grace A. Eramo (Department of Pharmacology, University of Michigan), Ben T. Pockrass (Department of Biological Chemistry, University of Michigan), Michelle L. Hastings (Department of Pharmacology, University of Michigan), Rachel O. Niederer (Department of Biological Chemistry, University of Michigan)
Abstract:
Neurons have a high demand to regulate their proteome. They accomplish this in part by employing highly specialized splicing and translation regulation programs. We hypothesize these programs are interconnected in the regulation of splicing of the 5’ untranslated region (5’UTR) of mRNA. Regulatory features in the 5’UTR of mRNA play a critical role in translational control, including RNA-binding protein motifs, secondary structures, and upstream start codons and open reading frames. Therefore, though alternative splicing in the 5’UTR may not affect the coding sequence, it can alter the cis-regulatory content of the mRNA, leading to alterations in translation. To investigate the contribution of alternative splicing to translation, we employed a high-throughput ribosome recruitment assay, DART (Direct Analysis of Ribosome Targeting). We use ribosome recruitment scores to identify gene transcripts with splice isoforms and regulatory regions associated with changes in translation initiation. Our preliminary data show that splice isoforms result in up to 10-fold differences in ribosome recruitment. To explore the regulatory role of this phenomenon, we have refined the assay to enrich for 5’ UTR spliced isoforms in neuronal and non-neuronal lysates to determine the effect of tissue-specific alternative splicing on ribosome recruitment. Luciferase reporters are being used in in vitro and in vivo translation assays to validate these results. Understanding the role of introns and alternative splicing in the 5’UTR of mRNA will lead to a better understanding of the basic mechanisms of translation and will reveal potential regulatory sequences that can be targeted for therapeutic modulation of gene expression.
Keywords: alternative splicing, 5UTR, translation