Roles of atypical protein kinase C in lysophosphatidic acid-induced type II adenylyl cyclase activation in RAW 264.7 macrophages

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Abstract
Lysophosphatidic acid (LPA) has been widely studied as a naturally occurring and multifunctional phospholipid messenger in diverse tissue and cell types and shown to inhibit adenylyl cyclase (AC) by a G protein-mediated mechanism. In type II AC-expressing mouse RAW 264.7 macrophages, we showed that LPA at 3–50 μM increased cyclic AMP formation in a concentration-dependent manner, the effect being additive with that of forskolin or cholera toxin, and synergistic with that of prostaglandin E1 (PGE1) or isoproterenol. The potentiation effect of LPA was unaffected by the removal of serum or pertussis toxin treatment. Both colchicine and cytochalasin B potentiated the cyclic AMP response to PGE1, the effect being additive to that of LPA. On studying the regulation of type II AC by protein kinase C (PKC), phorbol 12-myristate-13 acetate (PMA) potentiated the PGE1-elicited cyclic AMP response, this effect being non-additive to that of LPA, suggesting that PKC activation was the common mechanism involved in AC potentiation by LPA and PMA. PKC inhibitor Ro 31-8220, but not Go 6976, significantly inhibited the LPA-induced cyclic AMP potentiation. The potentiation effect of LPA was unaffected by long-term treatment with PMA, which resulted in the down-regulation of PKCα, βI, βII and PKCδ, but not PKCε, μ, λ and ζ. By in situ kinase assay, we found a marked increase in atypical PKC activity after LPA treatment. Taken together, we conclude that LPA can elicit a unique signalling cascade in RAW 264.7 macrophages and increase type II AC activity via the activation of atypical PKC.