Quantitative Profiling of Peptides from RNAs

Only a small fraction of the mammalian genome codes for messenger RNAs destined to be translated into proteins, and it is generally assumed that a large portion of transcribed sequences including introns and several classes of non-coding RNAs (ncRNAs) do not give rise to peptide products. A systematic examination of translation and physiological regulation of ncRNAs has not been conducted. Here, we use computational methods to identify the products of non-canonical translation in mouse neurons by analyzing unannotated transcripts in combination with proteomic data. This study supports the existence of non-canonical translation products from both intragenic and extragenic genomic regions, including peptides derived from anti-sense transcripts and introns. Moreover, the studied novel translation products exhibit temporal regulation similar to that of proteins known to be involved in neuronal activity processes. These observations highlight Correspondence: J.S. (judith.steen@childrens.harvard.edu) or G.K. (gabriel.kreiman@childrens.harvard.edu). 5Current address: Institute for Biochemistry and Molecular Biology Nussallee 11, 53115 Bonn, Germany 6Current address: School of Biological Sciences, University of Auckland, Auckland 1010, New Zealand. 7Current address: Merck Millipore, Im Laternenacker 5, 8200 Schaffhausen, Switzerland 8Current address: Department of Neurobiology, Harvard Medical School †These authors contributed equally Authors contributions: SP: designed and performed the final mass spectrometry and bioinformatics experiments, analyzed the data and wrote the manuscript; MH: developed concept, designed and carried out bioinformatics data analysis, and wrote the manuscript; RC: designed and carried out mass spectrometry validation experiments, molecular biology and biochemistry follow-up experiments, and wrote the manuscript; DW: designed and carried out the quantitative proteomics experiments; RYTC: carried out initial pilot experiments and tested strategies; CD: designed and performed initial neurobiologyexperiments; EH: designed and performed initial neurobiological experiments; HS: contributed to the mass spectrometric experiments and critically evaluated the data; JG: contributed to the conceptual design and critically evaluated the results; GK: developed concept, and supervised the RNA-seq part of the project and wrote the manuscript. JS: developed concept, designed initial experiment, supervised the proteomics part of the project and wrote the manuscript. Accession codes: RNA-seq data is already published (References: 5 and 16). The data can be found on GEO with the code GSE21161 Competing financial interests: The authors declare no competing fiancial interest. HHS Public Access Author manuscript Nat Commun. Author manuscript; available in PMC 2015 May 18. Published in final edited form as: Nat Commun. ; 5: 5429. doi:10.1038/ncomms6429. A uhor M anscript