Adenosine and its precursors, ATP and ADP, exert various physiological effects
via binding to purinergic receptors. We previously used co-immunoprecipitation, bioluminescence resonance energy transfer (BRET) and immunoelectron microscopy to demonstrate the hetero-oligomerization of purinergic receptor subtypes. Furthermore, pharmacological studies found significant changes in receptor-mediated signaling in human embryonic kidney (HEK) 293T cells co-transfected with these receptors. These findings suggest that heterodimers of purinergic receptors may have distinct pharmacological profiles, possibly due to dimerization-induced conformational changes, further suggesting that hetero-dimerization may be employed to “fine-tune” purinergic receptor signaling. Adenosine A
2A receptor (A
2AR), P2Y
1 receptor (P2Y
1R) and P2Y
12 receptor (P2Y
12R) are predominantly expressed on human platelets. ADP activates human platelets by stimulating both P2Y
1R and P2Y
12R, which act sequentially and in concert to achieve complete platelet aggregation. In contrast, adenosine stimulates Gs-coupled A
2AR, followed by activativation of adenylate cyclase, leading to an increase in intracellular cAMP levels, which potently inhibits platelet activation. We examined the hetero-oligomerization and functional interactions of A
2AR, P2Y
1R, and P2Y
12R. In HEK293T cells triply expressing all three receptors, hetero-oligomerization was observed among the three receptors. Additionally, P2Y
1R agonist-evoked Ca
2+ signaling was significantly inhibited by co-treatment with an A
2AR antagonist in HEK293T cells. In human platelets, we identified endogenous A
2AR/P2Y
1R and A
2AR/P2Y
12R heterodimers. We also observed functional Ca
2+-signaling-related cross-talk similar to those found in HEK293T cells, and found that they appeared to affect platelet shape. These results collectively suggest that intermolecular signal transduction and specific conformational changes occur among components of the hetero-oligomers formed by these three receptors.
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