Transcript isoform switching during embryonic and post-hatch development of broiler breast muscle

Gene-level analysis of RNA-Seq data provides only a partial view of muscle transcriptional regulation because gene isoforms may be differentially used without changes in total gene expression. This study evaluated the extent and potential role of differential transcript usage (DTU) during breast muscle development from the late embryonic stage to market age. Publicly available RNA-Seq data from breast muscle of Cornish, White Plymouth Rock, and their crossbred progeny sampled at embryonic day 17 (ED17) and post-hatch days 1, 21, and 42 were analyzed using The R package IsoformSwitchAnalyzeR (n = 6 samples per genetic background and stage, total = 72 samples). The highest number of isoform switches (|Δ isoform fraction| > 0.1 and FDR < 0.05) occurred during ED17-to-D1 transition (933 switches), followed by D1-to-D21 (631 switches), whereas only 46 occurred between D21 and D42. Exon skipping, alternative transcription start sites, and alternative transcription termination sites were the most frequent splicing events associated with these switches (n = 1,042, 964, and 817, respectively). Changes in coding sequence completeness, intrinsically disordered regions, and protein domains were the most frequently predicted functional consequences (n = 497, 472, and 461 events, respectively). Gene ontology enrichment (P < 0.05) indicated that ED17-to-D1 switching genes were involved in sarcomeric and cytoskeletal organization, redox and metabolic adaptation, signaling, chromatin regulation, and RNA processing, with top genes including OBSCN, DTNA, ABLIM3, and LIMCH1. D1-to-D21 switching genes were associated with cytoskeletal restructuring, adhesion and mechanosensing, RNA processing, protein turnover, autophagy, metabolic adaptation, and stress signaling, with TNNT3, FHL1, and FHL3 among the top-ranked genes. Although fewer genes exhibited isoform switching during the D21-to-D42 transition, the top-ranked genes included MYH1B, SMYD1, and MRTFA, which are involved in contractile or muscle regulatory processes. In conclusion, isoform switching represents an additional layer of transcriptional regulation of muscle development and it may contribute to hatch-related maturation, rapid post-hatch growth, and later refinement of contractile function. Future studies should experimentally validate candidate switches and their predicted functional consequences.