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We have isolated MLC1(F) (tMLC1(F)), MLC3(F) (tMLC3(F)) and skeletal actin cDNAs from the teleost Oncorhynchus mykiss. Sequence analysis indicates that tMLC1(F) and tMLC3(F) are not produced from differentially spliced mRNAs as reported in avians and rodents but are encoded by different genes. Results from RNase protection analysis showed that the corresponding transcripts are expressed in fast skeletal muscles. Whole-mount in situ hybridisation revealed distinct expression patterns of the myosin alkali light chains and skeletal actin genes during skeletal muscle development in the embryo.

FGF6, a member of the fibroblast growth factor (FGF) family, is specifically expressed in developing skeletal muscle and may participate in muscle maintenance and regeneration. Until now, no convincing evidence for the existence of an FGF6 gene in non-mammalian vertebrates has been put forward. Only a hybrid growth factor containing features characteristic of both FGF4 and FGF6 has been identified in frogs and chickens, suggesting that the step of duplication which created FGF4 and FGF6 took place with the emergence of mammals. In this study, we report the isolation and characterization of a genomic clone encoding the trout (Oncorhynchus mykiss) fibroblast growth factor 6 (TFGF6). An initial cDNA clone was generated by PCR amplification using degenerate oligo primers corresponding to a conserved region of protein found in the mouse and human homologs. The screening of a genomic library with the cloned PCR product led to the isolation of a clone composed of three exons encoding a putative protein of 206 amino acids which exhibits a potential signal peptide and shows 64.6 and 63.6% similarity with mouse and human FGF6, respectively (77% over the carboxy two-thirds of the protein) and only 46.5% similarity with mouse and human FGF4 (62% over the carboxy two-thirds of the protein). The splice position of the three exons was found to be analogous to the human and mouse FGF6 and the start translation site of TFGF6 was preceded by a long stretch of nucleotides that is highly and specifically conserved in mammalian FGF6. Furthermore, a comparative reverse transcriptase-linked PCR assay revealed that the expression pattern of TFGF6 is close to that of mammals, TFGF6 transcripts being present in muscle (fast-twitch and to a lesser extent slow-twitch fibers), heart, testis and brain. Interestingly, the prolonged phase of muscle fiber hyperplasia which occurs in trout is accompanied by the lasting expression of TFGF6 up to the adult stage suggesting that TFGF6 may participate in the continuous generation of muscle fibers within the myotomal musculature of post larval animals.

We have isolated the cDNA encoding a myogenic factor expressed in embryonic trout muscle by hybridization with a Xenopus MyoD cDNA. Nucleotide sequence analysis and amino acid comparison showed that this cDNA called TMyoD encodes a polypeptide of 276 amino acids with 70% identity to the entire Xenopus MyoD protein and 92% identity within the basic and myc-like region. Results from Northern blotting showed that the corresponding transcript is expressed both in adult and embryonic skeletal musculature and in an in vitro myogenesis system, but is undetectable in cardiac and smooth muscles and in non muscle tissues.