Reixachs, MarinaRuiz-Orera, JorgeAlba, M. MarEyras, Eduardo2026-01-282026-01-28Reixachs-Solé, M., Ruiz-Orera, J., Albà, M.M. et al. Ribosome profiling at isoform level reveals evolutionary conserved impacts of differential splicing on the proteome. Nat Commun 11, 1768 (2020). https://doi.org/10.1038/s41467-020-15634-w2041-1723https://hdl.handle.net/1885/733804991The differential production of transcript isoforms from gene loci is a key cellular mechanism. Yet, its impact in protein production remains an open question. Here, we describe ORQAS (ORF quantification pipeline for alternative splicing), a pipeline for the translation quantification of individual transcript isoforms using ribosome-protected mRNA fragments (ribosome profiling). We find evidence of translation for 40-50% of the expressed isoforms in human and mouse, with 53% of the expressed genes having more than one translated isoform in human, and 33% in mouse. Differential splicing analysis revealed that about 40% of the splicing changes at RNA level are concordant with changes in translation. Furthermore, orthologous cassette exons between human and mouse preserve the directionality of the change, and are enriched in microexons in a comparison between glia and glioma. ORQAS leverages ribosome profiling to uncover a widespread and evolutionarily conserved impact of differential splicing on translation, particularly of microexon-containing isoforms.We acknowledge funding from the Spanish Government and FEDER with grants BFU2015-65235-P, BIO2017-85364-R, and MDM-2014-0370, and by Catalan Government (AGAUR) with grant SGR2017-1020. MR-S had funding from an FI grant from the Catalan Government with reference 2018FI_B1_00126 for part of this work.application/pdfen-AU© 2020 The Author(s)http://creativecommons.org/licenses/by/4.0/202010.1038/s41467-020-15634-w2023-10-22Creative Commons Attribution 4.0 International License