The human platelet ADP receptor activates Gi2 proteins

Area of Science:

• Biochemistry
• Molecular Biology
• Pharmacology

Background:

• Platelet activation by adenosine diphosphate (ADP) involves G-protein coupled receptors.
• Previous studies demonstrated ADP receptor coupling to G-proteins using [35S]guanosine-5'-[gamma-thio]triphosphate ([35S]GTPγS) binding assays.

Purpose of the Study:

• To identify the specific G-proteins activated by ADP in human platelets.
• To elucidate the downstream signaling pathways mediated by ADP receptors.

Main Methods:

• Photolabelling of G-proteins with 4-azidoanilido-[alpha-32P]GTP.
• Immunoprecipitation of G-protein alpha-subunits using subtype-specific antibodies.
• Measurement of [35S]GTPγS binding to human platelet membranes.

Main Results:

• ADP stimulation increased 4-azidoanilido-[alpha-32P]GTP incorporation into G alpha i and specifically G alpha 12 immunoprecipitates.
• Thromboxane analogue U46619 activated G alpha q, while thrombin activated both G alpha i and G alpha q.
• ADP-induced activation was concentration-dependent, inhibited by ATP, and did not involve G alpha q activation of phospholipase C (PLC).

Conclusions:

• ADP inhibits cyclic AMP formation through the activation of G alpha 12 proteins in human platelets.
• Human platelet ADP receptors do not appear to activate phospholipase C (PLC) via Gq activation.