Protein kinase C-induced phosphorylation modulates the Na+-ATPase activityfrom proximal tubules

This study describes the modulation of the ouabain-insensitive Na--ATPase a ctivity from renal proximal tubule basolateral membranes (BLM) by protein k inase C (PKC). Two PKC isoforms were identified in BLM, one of 75 kDa and t he other of 135 kDa. The former correlates with the PKC isoforms described in the literature but the latter seems to be a novel isoform. not yet ident ified. Both PE;C isoforms of BLM are functional since a protein kinase C ac tivator, TPA, increased the total hydroxylamine-resistant P-32(i) incorpora tion from [gamma-P-32]ATP into the BLM. In parallel, TPA stimulated the Na-ATPase activity from BLM in a dose-dependent manner. the effect being reve rsed by the PKC inhibitor sphingosine. The stimulatory; effect of TPA on Na +-ATPase involved an increase in the IVmax (t`rom 13.4 +/- 0.6 nmol P-i mg( -1) min(-1) to 25.2 +/- 1.4 nmol P-i mg(-1) min(-1), in the presence of TPA . P < 0.05) but did not change the apparent affinity for Na+ (K-0.5 = 14.5 +/- 2.1 nM in control and 10.0 +/- 2.1 mM in the presence of TPA. P > 0.07) . PKC involvement was further confirmed by stimulation of the Na+-ATPase ac tivity by the catalytic subunit of PKC (PKC-M). Finally, the phosphorylatio n of an approx. 100 kDa protein in the BLM (the suggested molecular mass of Na+-ATPase [1]) was induced by TPA.. Taken together, these findings indica te that PKCs resident in BLM stimulate Na+-ATPase activity which could repr esent an important mechanism of regulation of proximal tubule Na- reabsorpt ion. (C) 2001 Elsevier Science B.V. All rights reserved.