NGFFFamide and echinotocin: structurally unrelated myoactive neuropeptides derived from neurophysin-containing precursors in sea urchins

Abstract

SUMMARYThe myoactive neuropeptide NGIWYamide was originally isolated from the holothurian (sea cucumber) Apostichopus japonicus but there is evidence that NGIWYamide-like peptides also occur in other echinoderms. Here we report the discovery of a gene in the sea urchin Strongylocentrotus purpuratus that encodes two copies of an NGIWYamide-like peptide: Asn-Gly-Phe-Phe-Phe-(NH 2 ) or NGFFFamide. Interestingly, the C-terminal region of the NGFFFamide precursor shares sequence similarity with neurophysins, carrier proteins hitherto uniquely associated with precursors of vasopressin/oxytocin-like neuropeptides. Thus, the NGFFFamide precursor is the first neurophysin-containing neuropeptide precursor to be discovered that does not contain a vasopressin/oxytocin-like peptide. However, it remains to be determined whether neurophysin acts as a carrier protein for NGFFFamide. The S. purpuratus genome also contains a gene encoding a precursor comprising a neurophysin polypeptide and 'echinotocin' (CFISNCPKGamide) -the first vasopressin/oxytocin-like peptide to be identified in an echinoderm. Therefore, in S. purpuratus there are two genes encoding precursors that have a neurophysin domain but which encode neuropeptides that are structurally unrelated. Furthermore, both NGFFFamide and echinotocin cause contraction of tube foot and oesophagus preparations from the sea urchin Echinus esculentus, consistent with the myoactivity of NGIWYamide in sea cucumbers and the myoactivity of vasopressin/oxytocin-like peptides in other animal phyla. Presumably the NGFFFamide precursor acquired its neurophysin domain following partial or complete duplication of a gene encoding a vasopressin/oxytocin-like peptide, but it remains to be determined when in evolutionary history this occurred.

NG peptides: A novel family of neurophysin-associated neuropeptides

“…Strongylocentrotus purpuratus there are two neuropeptide precursors that have a neurophysin domain: a precursor comprising neurophysin in association with a vasopressin/oxytocin-type neuropeptide ("echinotocin") and the NGFFFamide precursor (Elphick and Rowe, 2009). …”

“…Genes encoding precursors of vasopressin/oxytocin neuropeptides with a C-terminal neurophysin domain have been identified throughout the bilateria in both protostomian and deuterostomian invertebrate species (Van Kesteren et al, 1992;Stafflinger et al, 2008;Ukena et al, 2008;Elphick and Rowe, 2009) If NG peptide precursor genes evolved from a complete vasopressin/oxytocin precursor gene then one would expect to observe sequence similarity in N-terminal neuropeptide-containing regions of NG peptide precursors and vasopressin/oxytocin precursors. However, there is little if any such sequence similarity.…”

“…However, this is not the case; the genes are similar in having an intron that precedes the codon for the first residue of the neurophysin domain but the positions of other introns are different. In NG peptide precursor genes there is an intron separating the exon encoding the signal peptide and NG peptides, whereas in vasopressin/oxytocin-type neuropeptide precursor genes the signal peptide and vasopressin/oxytocin peptide are encoded by a single exon (Ivell and Richter, 1984;Elphick and Rowe, 2009). Conversely, in vertebrate and invertebrate vasopressin/oxytocin-type neuropeptide precursor genes the neurophysinencoding domain is typically interrupted by an intron, (Ivell and Richter, 1984;Stafflinger et al, 2008;Elphick and Rowe, 2009), whereas the neurophysin-encoding domain of NG peptide precursor genes is intronless.…”

“…However, this textbook perspective on neurophysins changed with the discovery of a gene in the sea urchin Strongylocentrotus purpuratus (phylum Echinodermata) encoding a precursor protein comprising a neurophysin domain in association with a myoactive neuropeptide that is structurally unrelated to vasopressin/oxytocin-type neuropeptides -Asn-Gly-PhePhe-Phe-NH 2 or NGFFFamide (Elphick and Rowe, 2009)…”

Discovery of a second SALMFamide gene in the sea urchin Strongylocentrotus purpuratus reveals that L-type and F-type SALMFamide neuropeptides coexist in an echinoderm species

“…To assess the utility of this EST collection as a resource for identification of cDNAs encoding neuropeptide precursors, known sea urchin neuropeptide precursor sequences were submitted as queries in tBLASTn searches of dbEST. Using this approach a cDNA (RNSP-5L15) encoding the precursor of the myoactive neuropeptide NGFFFamide was identified based upon both 5' (GI: 109403168) and 3' (GI: 109402129) EST data, as reported previously [24]. In addition, a cDNA (RNSP-5H7) encoding the precursor of the F-type SALMFamides SpurS1-SpurS7 [1] was identified based upon both 5' (GI:109402985) and 3' (GI: 109402020) EST data.…”