Molecular evolution of neuropeptides isolated from the brain of the Pacific salmon (Oncorhynchus SPP.)




Coe, Imogen Ruth

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The molecular evolution of three neuropeptides genes isolated from Pacific salmon (Oncorhynchus spp.) brain was investigated using both immunocytochemistry and molecular biology. The distribution of gonadotropin-releasing hormone (GnRH), a key reproductive neurohormone, was studied in protochordates, fishes and rat. The presence of GnRH-like immunoreactivity in the protochordates suggests that this hormone is phylogenetically ancient. The similar distribution of the peptide in the brains of fishes and mammals suggests that its location, and possibly function, has been conserved during evolution. The molecular structure of GnRH in salmon was investigated by isolation and characterization of one form, salmon (s) GnRH, from a sockeye salmon genomic library using a complementary (c) DNA for sGnRH originally isolated from cichlid. The entire protein coding region was sequenced and found to be distributed on three exons interrupted by two introns. The gene shows a high degree of sequence identity to an Atlantic salmon sGnRH gene and structural similarity to rat and human mammalian (m) GnRH genes. This suggests that both genes may be derived from a common ancestor. Two cDNAs encoding vasotocin, an important osmoregulatory neurohormone, were isolated from a chum salmon brain cDNA library using short degenerate probes against fish vasotocin. These clones have 65% sequence identity to each other at the protein level. The presence of two vasotocin precursors is probably due to the ancestral salmonid line becoming tetraploid at some point. Both GnRH and vasotocin exhibit typical features of members of evolutionarily ancient and diverse neuropeptide families. The hormone coding portion of the precursor is highly conserved in terms of protein sequence but the associated proteins, the signal and cryptic peptides, are not. This implies that the hormone portion of the precursor is under stricter constraint than the other regions. Three different cDNAs encoding α-tubulin were also isolated from the chum salmon brain cDNA library. These clones showed very high sequence identity with each other and with previously isolated α-tubulins from a variety of organisms. It appears that structural proteins, such as tubulin, are under strict constraint and limited in the extent to which their sequences can evolve before they become nonfunctional. Tubulin genes appear to have been duplicated repeatedly producing a multigene family. The tubulin clones show variations in their carboxy terminal regions and possess typical microtubule associated binding protein sites. These studies demonstrate that proteins, and portions of protein precursors, can evolve at different rates, which may be influenced by events such as gene duplication or tetraploidy.



Pacific salmon, Tubulin, Immunocytochemistry, Molecular biology