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Background/Aims: Gq protein-coupled receptors (GqPCRs) regulate various cellular processes including mainly proliferation and differentiation. In a previous study, we found that in prostate cancer cells, the GqPCR of GnRH induces apoptosis by reducing the PKC-dependent AKT activity and elevating JNK phosphorylation. Since it was thought that GqPCR induces mainly activation of AKT, we undertook to examine how general is this phenomenon and understand its signaling. Methods: We used various cells to follow the phosphorylation of signaling components using western blotting. Results: In a screen of 21 cell lines, we found that PKC activation results in the reduction of AKT activity, which correlates nicely to JNK activation and in some cases to apoptosis. To further understand the signaling pathways involved in this stimulation, we studied in detail the SVOG-4O and αT3-1 cells. We found that PGF2α and GnRH agonist (GnRH-a) indeed induce significant Gq- and PKC- dependent apoptosis in these cells. This is mediated by two signaling branches downstream of PKC, which converge at the level of MLK3 upstream of JNK. One branch consists on c-Src activation of the JNK cascade and the second involves reduction of AKT activity that alleviates its inhibitory effect on MLK3, to allow the flow of the c-Src signal to JNK. At the MAPKK level, we found that the signal is transmitted by MKK7 and not MKK4. Conclusion: Our results present a general mechanism that mediates a GqPCR-induced, death receptors-independent, apoptosis in physiological, as well as cancer-related systems.

Background G protein-coupled receptors (GPCRs) usually regulate cellular processes via activation of intracellular signaling pathways. However, we have previously shown that in several cell lines, GqPCRs induce immediate inactivation of the AKT pathway, which leads to JNK-dependent apoptosis. This apoptosis-inducing AKT inactivation is essential for physiological functions of several GqPCRs, including those for PGF2α and GnRH. Methods Here we used kinase activity assays of PI3K and followed phosphorylation state of proteins using specific antibodies. In addition, we used coimmunoprecipitation and proximity ligation assays to follow protein–protein interactions. Apoptosis was detected by TUNEL assay and PARP1 cleavage. Results We identified the mechanism that allows the unique stimulated inactivation of AKT and show that the main regulator of this process is the phosphatase PP2A, operating with the non-canonical regulatory subunit IGBP1. In resting cells, an IGBP1-PP2Ac dimer binds to PI3K, dephosphorylates the inhibitory pSer608-p85 of PI3K and thus maintains its high basal activity. Upon GqPCR activation, the PP2Ac-IGBP1 dimer detaches from PI3K and thus allows the inhibitory dephosphorylation. At this stage, the free PP2Ac together with IGBP1 and PP2Aa binds to AKT, causing its dephosphorylation and inactivation. Conclusion Our results show a stimulated shift of PP2Ac from PI3K to AKT termed “PP2A switch” that represses the PI3K/AKT pathway, providing a unique mechanism of GPCR-stimulated dephosphorylation.

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