YM-254890, which was isolated from the culture broth of Chromobacterium sp., inhibits ADP-induced platelet aggregation and has antithrombotic and thrombolytic effects. YM-254890 blocks G␣ q/11 -coupled ADP receptor P2Y1-mediated Ca 2؉ mobilization. Here we report that YM-254890 is a selective G␣ q/11 inhibitor. YM-254890 blocked Ca 2؉ mobilization mediated by several G␣ q/11 -coupled receptors but not by G␣ i -or G␣ 15 -coupled receptor, indicating that phospholipase C activation and subsequent signaling molecules are not the target of YM-254890. YM-254890 completely prevented the serum response factor (SRF)-mediated gene transcription induced by G␣ q R183C, which is constitutively active in a receptor-dependent manner because of its reduced k cat of GTP hydrolysis. Conversely, YM-254890 had only a modest effect on the SRF-mediated gene transcription by G␣ q Q209L, which is GTPase-deficient (activated) G␣ q . These suggested that the acting point of YM-254890 is receptor-G␣ q interaction or the subsequent guanine nucleotide exchange step. The fact that YM-254890 (i) inhibited the SRF-mediated gene transcription by G␣ qi5 , which interacts with G␣ i -coupled receptor and possesses the effector function of G␣ q , and (ii) had no effect on the K d value of high affinity [ These data indicate that YM-254890 blocks the exchange of GDP for GTP in G␣ q/11 activation. This novel G␣ q/11 -selective inhibitor is a promising and powerful tool for studying G␣ q/11 protein activation, G␣ q/11 -coupled receptor signaling, and G␣ q/11 -mediated biological events.
G protein-coupled receptors (GPCR)1 and heterotrimeric G proteins, consisting of G␣, , and ␥, transduce extracellular stimuli, such as hormones, neurotransmitters, chemokines, and other local mediators, into appropriate intracellular responses (1, 2). The activation of G␣ proteins is related to conformational change by guanine nucleotide interaction. The GPCRs, activated by the agonist, induced exchanges of GDP for GTP on the coupled G␣ subunit. The resultant G␣-GTP complex dissociates from the G␥ subunit and activates its downstream effectors, which in turn regulate various functions such as gene transcription, mitogenesis, metabolism, muscle contractile state, and ion channel regulation. The GTPase activity of ␣-subunit turns off effector signals by hydrolyzing G␣-GTP to G␣-GDP, which re-associates with G␥.There are over 20 G␣ subunits classified into subfamilies by its sequence homology and the downstream signal. These include the major four families, G␣ q/11 , G␣ s , G␣ i/o , and G␣ 12/13 . Main effector molecules of G␣ q/11 , G␣ s , G␣ i/o , and G␣ 12/13 are thought to be phospholipase C (PLC), adenylyl cyclase (activation), adenylyl cyclase (inhibition), and small GTPase families, respectively (3). Transient intracellular Ca 2ϩ mobilization is led by PLC activation via the G␣ q/11 , G␣ 15/16 , or G␥ subunit with G␣ i . PLC hydrolyzes phosphatidylinositol bisphosphate in the plasma membrane, and the generated inositol 1,4,5-trisphosphate (IP 3 ) activates ...
