Alternative splicing is one of the major cellular processes that determine the tissue-specific expression of protein variants.However,it remains challenging to identify physiologically relevant and tissue-selective pr...Alternative splicing is one of the major cellular processes that determine the tissue-specific expression of protein variants.However,it remains challenging to identify physiologically relevant and tissue-selective proteins that are generated by alternative splicing.Hence,we investigated the target spectrum of the splicing factor Rbfox1 in the cardiac muscle context in more detail.By using a combination of in silico target prediction and in-cell validation,we identified several focal adhesion proteins as alternative splicing targets of Rbfox1.We focused on the alternative splicing patterns of vinculin(metavinculin isoform)and paxillin(extended paxillin isoform)and identified both as potential Rbfox1 targets.Minigene analyses suggested that both isoforms are promoted by Rbfox1 due to binding in the introns.Focal adhesions play an important role in the cardiac muscle context,since they mainly influence cell shape,cytoskeletal organization,and cell–matrix association.Our data confirmed that depletion of Rbfox1 changed cardiomyoblast morphology,cytoskeletal organization,and multinuclearity after differentiation,which might be due to changes in alternative splicing of focal adhesion proteins.Hence,our results indicate that Rbfox1 promotes alternative splicing of focal adhesion genes in cardiac muscle cells,which might contribute to heart disease progression,where downregulation of Rbfox1 is frequently observed.展开更多
基金supported by grants from the Deutsche Forschungsgemeinschaft[DFG,German Research FoundationRTG 2467,project number 391498659‘Intrinsically disordered proteins—molecular principles,cellular functions,and diseases’+1 种基金FOR 5433,project number 468534282‘RNA in focus(RIF):from mechanisms to novel therapeutic strategies in cancer treatment’]the Wilhelm-Roux-Program(Medical Faculty,Martin-Luther-University Halle–Wittenberg).
文摘Alternative splicing is one of the major cellular processes that determine the tissue-specific expression of protein variants.However,it remains challenging to identify physiologically relevant and tissue-selective proteins that are generated by alternative splicing.Hence,we investigated the target spectrum of the splicing factor Rbfox1 in the cardiac muscle context in more detail.By using a combination of in silico target prediction and in-cell validation,we identified several focal adhesion proteins as alternative splicing targets of Rbfox1.We focused on the alternative splicing patterns of vinculin(metavinculin isoform)and paxillin(extended paxillin isoform)and identified both as potential Rbfox1 targets.Minigene analyses suggested that both isoforms are promoted by Rbfox1 due to binding in the introns.Focal adhesions play an important role in the cardiac muscle context,since they mainly influence cell shape,cytoskeletal organization,and cell–matrix association.Our data confirmed that depletion of Rbfox1 changed cardiomyoblast morphology,cytoskeletal organization,and multinuclearity after differentiation,which might be due to changes in alternative splicing of focal adhesion proteins.Hence,our results indicate that Rbfox1 promotes alternative splicing of focal adhesion genes in cardiac muscle cells,which might contribute to heart disease progression,where downregulation of Rbfox1 is frequently observed.
