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Untangling protein synthesis with ribosome profiling Pavel V. Baranov
School of Biochemistry and Cell Biology, University College Cork, Cork Ireland, p.baranov@ucc.ie

We used ribosome profiling (ribo-seq) to assess the gene expression response of mammalian cells to various stresses, such as increased eIF2 phosphorylation (the key step in Integrated Stress Response)1, and Oxygen and Glucose Deprivation (OGD)2. It enabled us to delineate the rapid translational response affecting thousands of genes. The response frequently involves translation of short regulatory ORFs usually located in the 5' leaders of mRNAs. We also observed translation of unannotated long ORFs that likely leads to the synthesis of novel protein products specific to stress conditions. To assist the research community in using ribo-seq data we are developing RiboSeq.Org suite of tools (http://riboseq.org) that currently consists of the GWIPS-viz browser for the visualization of genomic alignments of ribosome footprints3 and RiboGalaxy which is a Galaxy instance specifically tailored for the analysis of ribo-seq data4. In addition we developed a simple computational approach for the characterization of ribo-seq datasets5. This technique is resistant to irregular technical noise and aberrant footprint densities caused by ribosome pauses. Application of this approach to several ribo-seq datasets revealed the strong impact of sequencing biases and translation inhibitors on the distribution of aligned ribosome footprints as well as substantial non-biological variability between datasets obtained from different laboratories. 1. D.E.Andreev, P.B.O'Connor, C.Fahey, E.M.Kenny, I.M.Terenin, S.E.Dmitriev, P.Cormican, D.W.Morris, I.N.Shatsky, P.V.Baranov (2015) Translation of 5' leaders is pervasive in genes resistant to eIF2 repression. Elife 4:e03971. 2. D.E.Andreev, P.B.O'Connor, A.V.Zhdanov, I.D.Dmitriev, I.N.Shatsky, D.B.Papkovsky, P.V.Baranov (2015) Oxygen and glucose deprivation induces widespread alterations in mRNA translation within 20 minutes Genome Biol 16:90. 3. A.M.Michel, G.Fox, A.M.Kiran, C.De Bo, P.B.O'Connor, S.M.Heaphy, J.P.Mullan, C.A.Donohue, D.G.Higgins, P.V.Baranov (2014) GWIPS-viz: development of a ribo-seq genome browser. Nucleic Acids Res 42:D859-864. 4. A.M.Michel, J.P.A. Mullan, C.A.Donohue, V.Velayudhan, P.B. O'Connor, P.V. Baranov (2015) RiboGalaxy: a platform for the alignment, analysis and visualization of ribo-seq data. Under review. 5. P.B.O'Connor, D.E.Andreev, P.V.Baranov (2015) Surveying the relative impact of mRNA features on local ribosome profiling read density in 28 datasets. BioRxiv doi: 10.1101/018762