G-PROTEINS IN GUINEA-PIG AIRWAY SMOOTH-MUSCLE - IDENTIFICATION AND FUNCTIONAL INVOLVEMENT

In airway smooth muscle, G-proteins have not been identified directly as yet, This study was an attempt to detect various types of conventio nal G(i)- and G(s)-proteins in purified membranes of guinea pig airway smooth muscle and to assess the involvement of the G-proteins in agon ist-induced contractile response of the smooth muscle. Immunoblotting using AS/7 antibody which recognizes G(i-1/2) demonstrated the presenc e of polypeptides of M(r) 34, 41 and 75 kDa. Polypeptides of M(r) 43, 46 and 48 kDa were identified with RM/1 antibody that detects G(s)-typ e G-proteins. The AG/1 antibody that recognizes alpha-subunits common to all G(i)- and G(s)- proteins detected two polypeptides of M(r) 41 a nd 75 kDa. Heterotrimeric structure of the G-proteins was confirmed by the identification of a single dense beta-subunit band at M(r) 39 kDa with SW/1 antibody, Pertussis toxin (PT) ADP-ribosylated three G(i) a lpha polypeptides of M(r) 41, 43, and 62 kDa. On the other hand, chole ra toxin (CT) catalysed the ADP-ribosylation of two G(s) alpha polypep tides of M(r) 46 and 62 kDa. Both PT and CT attenuated the maximum con tractile responses of the airway smooth muscle to the muscarinic agoni st, methacholine. Pretreatment of the tissues with the sulphydryl alky lating G-protein inhibitor, N-ethylmaleimide, also inhibited the maxim um contractions to methacholine. These data suggest that plasma membra nes of guinea pig airway smooth muscle contain a variety of convention al, including G(s) and G(i), and other types of G-proteins, and at lea st a portion of the proteins present may be involved in mediating the contractile responses of the smooth muscle to an agonist such as metha choline. (C) 1996 The Italian Pharmacological Society