MOLECULAR CHARACTERIZATION AND SPATIO-TEMPORAL EXPRESSION PATTERN OF THE SHH GENE AND ITS REGULATION IN MUSCLE GROWTH IN SINIPERCA CHUATSI

Sonic hedgehog (Shh) is one of the important signaling molecules in the Hedgehog gene family that regulates cell proliferation and differentiation during the early embryo development. In this study, the DNA sequence of the Shh gene was obtained from Siniperca chuatsi genome, and the protein encoded by the gene and the homologous evolutionary features were analysed. The Siniperca chuatsi Shh gene has an open reading frame (ORF) of 1242 bp, encoding 413 amino acids, with molecular weight of 46.01 kD, isoelectric point (pI) of 6.57, lipolysis coefficient of 82.83, hydrophilic mean coefficient of −0.292, and one transmembrane structure, making it a hydrophilic membrane-bound protein. The Shh protein has two structural domains, Hh-N and Hh-C. The Siniperca chuatsi Shh protein shows a high degree of homology with that of the perciforms. The spatio-temporal expression pattern of the Shh gene in Siniperca chuatsi was analysed by using Real-time fluorescence quantitative PCR. The results showed that the expression of Shh was significantly up-regulated at the neurula stage and remained at a high level during the middle and later stages of embryonic development. Shh was differentially expressed in different tissues of Siniperca chuatsi, with higher expression in brain and intestine, and lower expression in fast muscle, slow muscle and other tissues. The expression of muscle development related genes was analysed in Siniperca chuatsi embryos which treated with Cyclopamine. The results showed that the expression of Pax3, Pax7, Myomaker, MRFs, MyLC, MyHC and MLCK, were significantly reduced after inhibiting the Shh signalling pathway, suggesting that Shh is involved in regulating the early differentiation and development of Siniperca chuatsi muscle cells. The study will help us to understand the molecular mechanism of Shh signalling in regulating fish muscle development at the molecular level and provide a reference for fish developmental biology and healthy breeding.