A Hypertrophic Stimulus Drives Age-Dependent Transcript Isoform Switching With and Without Differential Gene Expression in Skeletal Muscle.

Aged skeletal muscle has a reduced ability to adapt to hypertrophic stimuli concomitant with dysregulation across several molecular layers. Whether altered RNA splicing associates with impaired adaptive potential is not well studied. We hypothesized that aging disrupts the splicing response to involuntary muscle mechanical loading. Using deep short-read Illumina RNA-sequencing (SRS, 60 million read pairs) and third generation Oxford Nanopore long-read RNA-sequencing (LRS) combined with a murine model of acute muscle mechanical overload (MOV), we illustrate how aged muscle has a reduced or possibly delayed potential for isoform switching in response to stress. SRS identified hundreds of genes with isoform switching events in adult MOV relative to sham but only 37 in aged MOV. LRS identified 19 genes (including Cacnb1, Naca, Psap) with true isoform switching in adult MOV and none in aged MOV. A subset of genes showed significant isoform switching without corresponding changes in overall gene expression, suggesting post-transcriptional regulation in response to MOV. These data highlight the importance of analyzing the transcriptome at the isoform level to uncover transcript diversity and "unseen genes" that may have unique functions and mechanisms influencing hypertrophic adaptations.