Stimulation by extracellular ATP and UTP of the mitogen‐activated protein kinase cascade and proliferation of rat renal mesangial cells

Abstract

Extracellular ATP and UTP have been reported to activate a nucleotide receptor that mediates phosphoinositide and phosphatidylcholine hydrolysis by phospholipases C and D, respectively. Here we report that ATP and UTP potently stimulate mesangial cell proliferation. Both nucleotides stimulate phosphorylation and activation of mitogen‐activated protein kinase and a biphasic phosphorylation of the up‐stream mitogen‐activated protein kinase kinase. When added at 100 μm, ATPγS, UTP and ATP were the most potent activators of mitogen‐activated protein kinase. βγ‐imido‐ATP was somewhat less active and ADP and 2‐methylthio‐ATP caused a weak induction of enzyme activity. Activation of mitogen‐activated protein kinase by both ATP and UTP is dose‐dependently attenuated by the P2‐receptor antagonist, suramin. The protein kinase C activator 12‐0‐tetradecanoylphorbol 13‐acetate, but not the biologically inactive 4α‐phorbol 12,13‐didecanoate, increased mitogen‐activated protein kinase activity in mesangial cells, suggesting that protein kinase C may mediate nucleotide‐induced stimulation of mitogen‐activated protein kinase. Down‐regulation of protein kinase C ‐α and‐θ isoenzymes by 4 h or 8 h treatment with phorbol ester partially inhibited ATP‐ and UTP‐triggered mitogen‐activated protein kinase activation. Moreover, a 24 h treatment of mesangial cells with phorbol ester, a regimen that also causes depletion of protein kinase C‐∍ did not further reduce the level of mitogen‐activated protein kinase stimulation. The specific protein kinase C inhibitor, CGP 41251, which displayed a selectivity for the Ca2+‐dependent isoenzymes, as compared to the Ca2+‐independent isoenzymes did not inhibit nucleotide‐stimulated mitogen‐activated protein kinase phosphorylation, thus implicating the involvement of a Ca2+‐independent protein kinase C isoform. In summary, these results suggest that ATP and UTP trigger the activation of the mitogen‐activated protein kinase signalling cascade in mesangial cells and this may be responsible for the potent mitogenic activity of both nucleotides. 1994 British Pharmacological Society