G-PROTEIN EXPRESSION IN HUMAN FETOPLACENTAL VASCULARIZATION - FUNCTIONAL EVIDENCE FOR G(S)ALPHA AND G(I)ALPHA SUBUNITS

GTP-binding proteins are key elements in coupling receptors to various effector systems. Using ADP-ribosylation by cholera (CTX) and pertuss is (PTX) toxins and an immunodetection technique, we investigated the G protein expression profile in smooth muscle of stem villi vessels ob tained from human term placentae. In placental vascular smooth muscle, we report the presence of two CTX-protein substrates of 42 and 45 kDa recognized by G(s) alpha antibodies, and three G(i) alpha isoforms, s ubstrates of PTX, identified as G(i1)alpha, G(i3)alpha (two proteins o f 41 kDa) and G(i2)alpha (a 40-kDa protein), We also characterized ano ther target of PTX, a 40-kDa G(0) alpha-immunoreactive protein and det ected the PTX-insensitive G(q)-G(11)alpha proteins. To assess the func tional significance of the G alpha proteins identified in this tissue, we measured the adenylyl cyclase activity in the presence of guanyl n ucleotides alone or with increasing concentrations of vasoactive intes tinal peptide (VIP), and examined whether VIP-bound sites, in the pres ence of GTP gamma S, promote the release of G alpha proteins from the membranes of vascular smooth muscle. At low concentrations (0.1 nM to 0.01 mu M), guanyl nucleotides stimulated adenylyl cyclase activity in a dose-dependent manner, while at higher concentrations (10 mu M to 1 mM) the stimulation rate of cAMP production by guanyl nucleotides dec reased. In a dose-dependent manner, VIP in the presence of GTP gamma S increased adenylyl cyclase activity and specifically promoted the rel ease of both G,(s) alpha isoforms. In contrast, the release of G(i1) a nd G(i2)alpha isoforms was not significantly increased in the presence of VIP, while GTP gamma S alone stimulated their release. Our data sh ow physical evidence of the activation of G(s) proteins by VIP-bound m embrane receptors, resulting in dissociation and release of G(s) alpha subunits in the soluble fraction. They assess the specific coupling o f the two G(s) alpha isoforms to VIP receptors in smooth muscle wall o f placental stem villi vessels. It would be of interest to investigate whether changes in G(2) alpha expression and/or function are associat ed with the placental angiogenesis process during pregnancy. (C) 1996 Academic Press Limited