Biological activity of the predicted red pigment-concentrating hormone of Daphnia pulex in a crustacean and an insect

The elucidation of the genome of the waterflea Daphnia pulex made it possible to search for orthologue genes for the crustacean red pigment-concentrating hormone (named Panbo-RPCH after the species Pandalus borealis in which the red pigment-concentrating hormone was first identified); Panbo-RPCH is a member of the adipokinetic hormone (AKH)/red pigment-concentrating hormone (RPCH) peptide family. The information pointed to a putative mature RPCH octapeptide in D. pulex with the primary sequence of pGlu-Val-Asn-Phe-Ser-Thr-Ser-Trp amide (=Dappu-RPCH). Since Panbo-RPCH is endogenous in decapod crustaceans and in the green stink bug Nezara viridula, we assayed Dappu-RPCH in the shrimp Palaemon pacificus and in N. viridula. Here we show that this variant member of the AKH/RPCH family has no activity to concentrate the red, brown, yellow and blue pigments in the epithelium of the shrimp at physiological doses but is effective in mobilising lipids in the green stink bug N. viridula. Moreover, since Panbo-RPCH and Dappu-RPCH differ structurally at three positions, viz. Leu2 to Val2; Pro6 to Thr6; Gly7 to Ser7, we tested other members of the peptide family which have single or dual amino acid substitutions at the appropriate positions, for their chromatophorotropic action at physiological doses. These studies show unequivocally that a single change from Gly7 to Ser7 (as in the peptide Corpu-AKH) does not inflict any loss of biological activity, and the same is true for a single change from Pro6 to Thr6 (represented by the peptide Schgr-AKH-II). The change from Leu2 to Val2 (embodied in Manto-CC), however, is accompanied with a substantial loss of chromatophorotropic activity; combinations of Val2 and Ser7 (as in Anaim-AKH) or Val2 and Thr6 (as in Grybi-AKH) result in almost complete loss of biological activity. Dappu-RPCH with its three substitutions is not active at all in the shrimp at the tested concentration range of up to 30 pmol.