Type A allatostatins from Drosophila melanogaster and Diplotera puncata activate two Drosophila allatostatin receptors, DAR-1 and DAR-2, expressed inCHO cells

The type-A allatostatins A (AST-A) are a group of insect peptides with a co mmon C-terminal motif Y/FXFGL-NH2. The existence of at least four putative type A Drosophila melanogaster ASTs (called type A drostatins or DST-As) ha s been predicted from the sequence of a recently cloned DST-A preprohormone [C. Lenz et al. (2000) Biochem. Biophys. Res. Commun. 273, 126-1131]. SRPY SFGL-NH2, (DST-3A), the only DST isolated from Drosophila so far, activated the first cloned DST-A GPCR (DAR-1) [N. Birgul et al. (1999) EMBO J. 18, 5 892-5900]. A newly cloned orphan Dm GPCR, which shares 47% overall and 60% transmembrane region sequence identity with DAR-1, was classified as a seco nd putative Dm DST-A receptor (DAR-2) [C. Lenz et al. (2000) Biochem. Bioph ys. Res. Commun. 273, 571-577]. Although activation of DAR-2 by DSTs has be en postulated, no experimental evidence for that has been presented to date . In this study, we expressed both DAR-1 and DAR-2 in CHO cells and used a GTP gammaS and a Ca2+ mobilization assay for pharmacological evaluation of the receptors. Synthetically prepared DST-As, as well as selected Diplotera punctata (cockroach) ASTs, activated DAR-I and DAR-2 in both functional as says indicating ligand redundancy and cross species activity. Cell pretreat ment with pertussis toxin led to some differences in the nature and magnitu de of signaling pathways at the DAR-I and DAR-2 receptors, suggesting possi ble differential coupling to cellular effector system(s) and distinct biolo gical functions of each receptor in vivo. (C) 2001 Academic Press.