VIP activates G(s) and G(i3) in rat alveolar macrophages and G(s) in HEK293 cells transfected with the human VPAC(1) receptor.
We have characterized vasoactive intestinal peptide (VIP) receptor/G-protein coupling in rat alveolar macrophage (AM) membranes and find that pertussis toxin treatment and antisera against G(alphai3) and G(alphas) reduce high-affinity (125)I-VIP binding, indicating that both G(alphas) and G(alphai3) couple to the VIP-receptor. The predominant VIP-receptor subtype in AM is VPAC(1) and we examined the G-protein interactions of the human VPAC(1) that had been transfected into HEK293 cells. VPAC(1) has a molecular mass of 56 kDa; GTP analogs reduced (125)I-VIP binding to this protein demonstrating that high-affinity binding of VIP to the receptor requires coupling to G-protein. Functional VIP/VPAC(1)/G-protein complexes were captured by covalent cross-linking and analyzed by Western blotting. The transfected human VPAC(1) receptor in HEK293 was found to be coupled to G(alphas) but not G(alphai) or G(alphaq). Furthermore, pertussis toxin treatment had no effect on VPAC(1)/G-protein coupling in these cells. These observations suggest that the G-proteins activated by VPAC(1) may be dependent upon species and cell type.
Shreeve
S M
SM
Department of Pharmacology, University of Vermont, Burlington 05405, USA. Martin.Shreeve@vtmednet.org
Sreedharan
S P
SP
Hacker
M P
MP
Gannon
D E
DE
Morgan
M J
MJ
D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D020962:Heterotrimeric GTP-Binding Proteins / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006801:Humans; D007106:Immune Sera / Q000494:pharmacology; D016676:Macrophages, Alveolar / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009928:Organ Specificity; D037342:Pertussis Toxin; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D018005:Receptors, Vasoactive Intestinal Peptide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051238:Receptors, Vasoactive Intestinal Polypeptide, Type I; D015398:Signal Transduction / Q000187:drug effects; D013045:Species Specificity; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D003432:Cross-Linking Reagents; D007106:Immune Sera; D018005:Receptors, Vasoactive Intestinal Peptide; D051238:Receptors, Vasoactive Intestinal Polypeptide, Type I; D010566:Virulence Factors, Bordetella; D014660:Vasoactive Intestinal Peptide; D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D019205:GTP-Binding Protein alpha Subunits, Gs; D020962:Heterotrimeric GTP-Binding Proteins