Asynchronous activation of 10 muscle-specific protein (MSP) genes during zebrafish somitogenesis

In the present study, 10 zebrafish cDNA clones coding for muscle-specific proteins (MSPs) were characterized and most of them encode fast skeletal muscle isoforms. They are skeletal muscle alpha-actin (acta1), fast skeletal muscle a-tropomyosin (tpma), fast skeletal muscle troponin C (tnnc), fast skeletal muscle troponin T (tnnt), fast skeletal muscle myosin heavy chain (myhz1), fast skeletal muscle myosin light chain 2 (mylz2), fast skeletal muscle myosin light chain 3 (mylz3), muscle creatine kinase (ckm), parvalbumin (pvalb), and desmin (desm). Using these cDNA probes, their expression patterns in developing embryos and adults were compared by Northern blot hybridization and whole-mount in situ hybridization. All of the 10 genes are expressed in both embryos and adult fish, and the expression is highly abundant in skeletal muscle. Among them, acta1, tpma, tnnc, tnnt, myhz1, mylz2, mylz3 and pvalb, are expressed specifically in fast skeletal muscle while ckm and desm are expressed in both fast and slow skeletal muscles. In addition, tpma, ckm, and desm are also expressed in the heart. Ontogenetically, the onset of expression of these MSP genes in zebrafish skeletal muscle varies and the expression occurs rostral-caudally in developing somites. Shortly after the expression of myoD, desm is the first to be activated at approximately 9 hpf, followed by tpma (approximately 10 hpf), tnnc (approximately 12 hpf), acta1 (approximately 12 hpf), ckm (approximately 14 hpf), myhz1 (approximately 14 hpf), mylz2 (approximately 16 hpf), mylz3 (approximately 16.5 hpf), tnnt (approximately 16.5 hpf), and pvalb (approximately 16.5 hpf). At later stages (after 48 hpf), these MSP genes are also expressed in fin buds and head muscles including eye, jaw, and gill muscles. Thus, our experiment demonstrated the order of expression of the 10 MSP genes, which may reflect the sequence of muscle filament assembly. In spite of the asynchrony in activation of these MSP genes, the timing of expression for each individual MSP gene appears to be synchronous to somite development as each somite has an identical timetable to express the set of MSP genes.