The Vesicle‐Mediated Transport Genes, Snap23 , Tmed2 , and Trip10 , are Alternatively Spliced During Striated Muscle Development

Abstract

The reprogramming of alternative splicing patterns during development is a hallmark of tissue maturation and identity. Heart and skeletal (striated) muscle tissues exhibit the most highly conserved splicing programs, which substantially change during postnatal development. Notably, alterations of these splicing events are associated with severe cardiac and muscular diseases. In striated muscle, a collection of genes encoding membrane trafficking proteins are spliced in a developmental stage‐specific manner. However, few splice isoforms have been characterized, and little is known about their functional roles during heart and skeletal muscle development. We hypothesize that coordinated splicing of membrane trafficking genes represents a splicing network, which functions to regulate the transport of cargo that is critical for muscle development and homeostasis. Here, we characterize three proteins involved in vesicle‐mediated transport that are alternatively spliced in striated muscle: SNAP23, TMED2, and TRIP10. The synaptosome‐associated protein 23 (SNAP23) is a SNARE protein that mediates vesicle fusion with the plasma membrane during exocytosis, the transmembrane emp23 domain‐containing protein 2 (TMED2) regulates vesicle budding during secretion, and the thyroid hormone receptor interactor 10 (TRIP10) is involved in endocytosis and membrane tubulation. First, we found that during striated muscle development, the Snap23, Tmed2, and Trip10 pre‐mRNAs undergo splicing changes of a single exon. Second, these three splicing transitions are characterized by a shift from short to long isoform expression that are regulated by the RBPs quaking (QKI) and polypyrimidine tract binding protein 1 (PTBP1). Third, depletion of SNAP23 in muscle cells results in reduced cell viability and differentiation, which can be rescued with conditioned media from differentiated cells or by seeding cells on collagen‐coated plates. Finally, exclusive expression of the short SNAP23 isoform leads to increased fusion of myoblasts into myotubes that exhibit greater myotube area and a higher nuclei‐per‐myotube ratio. Taken together, our data suggest that splicing of Snap23 is an important regulator of myogenesis, where SNAP23 isoform expression controls secretion of cargo that is necessary for muscle cell fusion and myotube size. Preliminary studies show that disruption of TMED2 and TRIP10 isoform levels in muscle cells likewise affects differentiation. Elucidating the mechanisms of regulation and the functional consequences of splicing networks will be fundamental to our understanding of muscle development and disease.