Meeting Abstract

P3.133  Jan. 6  Cloning and expression of recombinant crustacean hyperglycemic hormone isoforms from land crab, Gecarcinus lateralis ZARUBIN, T.*; MELZER, S.A.; DENNIS, D.; DORAN, K.J. Lee; MYKLES, D.L.; Colorado State University; Colorado State University; Colorado State University; Colorado State University; Colorado State University tzarubin@colostate.edu

Crustacean hyperglycemic hormone (CHH) regulates carbohydrate metabolism, molting, and ion and water transport. We have previously shown that two groups of CHH isoforms exist in the blackback land crab, Gecarcinus lateralis: EG-CHH group, which consists of four isoforms, were expressed in eyestalk ganglia (EG) and HT-CHH, which was similar to the pericardial organ CHH isoform in other decapods, was expressed in hindgut and testis (HT). The HT-CHH deduced amino acid (aa) sequence was identical to the EG-CHH isoforms through aa #40 of the mature peptide. The divergent aa sequence between positions #41 and #73 was encoded by an insertion of a novel 111-bp sequence absent in EG-CHH cDNAs. Based on the differences in sequence and tissue expression, we hypothesize that the two isoforms differ in hyperglycemic and molt inhibitory activities. To test this hypothesis, we used a yeast expression system to synthesize recombinant EG-CHH and HT-CHH mature peptides. To date, recombinant Myc-tagged EG-CHH has been expressed and its levels of expression have been determined; recombinant EG-CHH has been expressed but levels of expression have not been determined; and HT-CHH is currently being cloned into our yeast expression vector. Once expressed, we will determine the biological activities of EG-CHH and HT-CHH in vitro and in vivo. Future research will use these recombinant peptides for the characterization of membrane receptors and signaling mechanisms regulating molting in crustaceans. Supported by NSF (IBN-0342982).