Cloning and functional expression of the first Drosophila melanogaster sulfakinin receptor DSK-R1.
Described in this report is a successful cloning and characterization of a functionally active Drosophila sulfakinin receptor designated DSK-R1. When expressed in mammalian cells, DSK-R1 was activated by a sulfated, Met(7-->Leu(7)-substituted analog of drosulfakinin-1, FDDY(SO(3)H)GHLRF-NH(2) ([Leu(7)]-DSK-1S). The interaction of [Leu(7)]-DSK-1S with DSK-R1 led to a dose-dependent intracellular calcium increase with an EC(50) in the low nanomolar range. The observed Ca(2+) signal predominantly resulted from activation of pertussis toxin (PTX)-insensitive signaling pathways pointing most likely to G(q/11) involvement in coupling to the activated receptor. The unsulfated [Leu(7)]-DSK-1 was ca. 3000-fold less potent than its sulfated counterpart which stresses the importance of the sulfate moiety for the biological activity of drosulfakinin. The DSK-R1 was specific for the insect sulfakinin since two related vertebrate sulfated peptides, human CCK-8 and gastrin-II, were found inactive when tested at concentrations up to 10(-5) M. To our knowledge, the cloned DSK-R1 receptor is the first functionally active Drosophila sulfakinin receptor reported to date.
Kubiak
Teresa M
TM
Animal Health Discovery Research, Kalamazoo, Michigan 49001, USA. teresa.m.kubiak@pharmacia.com
Larsen
Martha J
MJ
Burton
Katherine J
KJ
Bannow
Carol A
CA
Martin
Roger A
RA
Zantello
Marjorie R
MR
Lowery
David E
DE
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D029721:Drosophila Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology; D010802:Phylogeny; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D011949:Receptors, Cholecystokinin; D043562:Receptors, G-Protein-Coupled
C449649:CCKLR-17D3 protein, Drosophila; D029721:Drosophila Proteins; D009479:Neuropeptides; D009842:Oligopeptides; D011956:Receptors, Cell Surface; D011949:Receptors, Cholecystokinin; D043562:Receptors, G-Protein-Coupled; C449650:drosulfakinin 1, sulfoTyr(4)-Leu(7)-; C056936:drosulfakinin 1; D002118:Calcium