Bwchtrmca et Bwph~swa Acta, 1096 (1991) 127-133 © 1991 Elsewer Soence Pubhshers B V 0925-4439/91/$03 50 ADONIS 0925443991000573 127 BBADIS 61028 Altered G-protein expression and adenylate cyclase activity in platelets of non-insulin-dependent diabetic (NIDDM) male subjects Callum Dvlngstone 1, Alastmr R McLellan 2, Mary-Anne McGregor 1, Andrew Wilson 1t, John M C Connell 2, Michael Small 4, Graeme Mdhgan 1, Kenneth R. Paterson 3 and Miles D Houslay 1 t Molecular Pharmacologr Group, Department of Btochemtstr~, Unwerstt~ of Glasgow,, Glasgow 2 MRC Blood Pressure Umt, Western lnftrmar~, Glasgow ~ Dtabetes Umt, Royal Infirmary, Glasgow and 4 Gartnavel General Hospital, Glasgo~ (U K ) (Recewed 8 October 1990) Key words G protein, Adenylate cyclase Diabetes Protein expression, (Rat membrane) Adenylate cyclase activity and levels of guanine nucleotide regulatory proteins (G-proteins) were compared in platelets from normal and non-insulin-dependent diabetic (NIDDM) male subjects. Whilst no differences were noted in basal and NaF-stimulated adenylate cyclase activities the degree of stimulation achieved by both forskolin and prostaglandin, E t was lower by some 34 and 52% respectively, in platelet membranes from diabetic subjects compared with those from normal control subjects Altered a~-adrenoceptor-mediated inhibition of prostaglandin E i-stimulated adenylate cyclase activity was evident; it being some 34% lower in platelet membranes from diabetic subjects compared to controls. Analysis of G-protein a-subunits, using specific anti-peptide antisera, showed that platelets from all subjects exhibited the G,-2 and G,-3, but not the G,-1 forms of the inhibitory G-protein 'G,' and all expressed the 42 kDa species of a-subunit of the stimulatory G-protein G s. Whilst platelets of diabetic subjects had levels of G s which were comparable to those found in control subjects their levels of G,-2 and G,-3 were some 49 and 75%, respectively, of those found in platelets from control subjects. It is suggested that changes in adenylate cyclase functioning and G-protein expression may contribute to altered platelet functioning in non-insulin-dependent diabetic subjects. Introduction It is generally considered that diabetes is associated with an increased mortahty due to cardiovascular dis- ease [see e g, Refs 1 and 2] Platelets have been im- plicated in the atherogemc process [1] and there is now a body of evidence suggesting that platelets from dia- betic subjects are more sensitive to agomst-induced aggregation [see eg, Refs 1, 3-8] It has even been suggested [9-13] that ttus may precede the appearance of clinically evident vascular disease The molecular basis of these changes in platelet functioning has yet to be ascertained However, alterations in the functioning of plasma membrane signalling systems coupled to both t Deceased Thursday June 28th 1990 Correspondence M D Houslay, Molecular Pharmacology Group, Department of Blochermstry, University of Glasgow, Glasgow G12 8QQ, U K adenylate cyclase and phospholnosltldase (phospho- llpase C) rmght be expected to have profound effects on platelet activation [14,15] Interestingly, recent observa- tions have demonstrated that platelets from non-in- sulin-dependent diabetic patients exhibit reduced mem- brane fluidity [16] Such alterations in membrane fluid° ~ty can be expected to have effects on the functiomng of integral membrane proteins [17,18] including those, such as adenylate cyclase [18-22], whose vanous hgand- stimulation activities have been shown to be particularly affected by such changes In platelets, adenylate cyclase IS regulated by stlmu- latory receptors, such as that for prostaglandm E I (PGE1) and by lntubitory receptors, such as the a 2- adrenoceptor, which exert functions through stlmula- tory (G~) and Inhibitory (G,) guanine nucleoude regu- latory proteins, respectively [14,15,23,24] These are het- erotrimerlc proteins consisting of a-, fl- and y-subumts The a-subunlts can bind GTP when activated through interaction with an occupied receptor, serving to couple