Ceramide-activated protein kinases A and C zeta inhibit kidney proximal tubule cell Na+-ATPase

Abstract

The basolateral membranes of kidney proximal tubule cells have (Na(+)+K(+))-ATPase and Na(+)-ATPase activities, involved in Na(+) reabsorption. We showed that ceramide (Cer) modulates protein kinase A (PKA) and protein kinase C (PKC), which are involved in regulating ion transporters. Here we show that ceramide, promotes 60% inhibition of Na(+)-ATPase activity (I(50) approximately 100nM). This effect was completely reversed by inhibiting PKA but did not involve the classic PKC signaling pathway. In these membranes we found the Cer-activated atypical PKC zeta (PKCzeta) isoform. When PKCzeta is inhibited, Cer ceases to inhibit the Na(+)-ATPase, allowing the cAMP/PKA signaling pathway to recover its stimulatory effect on the pump. There were no effects on the (Na(+)+K(+))-ATPase. These results reveal Cer as a potent physiological modulator of the Na(+)-ATPase, participating in a regulatory network in kidney cells and counteracting the stimulatory effect of PKA via PKCzeta.

Altered signaling pathways linked to angiotensin II underpin the upregulation of renal Na+-ATPase in chronically undernourished rats

“…7) would also contribute to activation of the pump because this kinase has been implicated in 2 different inhibitory pathways of Na + -ATPase, i.e. those linked to AT 2 R [39,42] and ceramide receptors [49]. As discussed above, these alterations may be a consequence of increased local activity of Ang II in the tubulointerstitium, which can stimulate translocation of the upregulated PKCs to the membranes after binding to AT 1 R [50].…”

“…Second, Ang III is functionally associated with an AT 2 R-mediated natriuretic responses [47,48] rather than with an increased AT 1 R-stimulated Na + reabsorption, as that mediated by the ouabain-resistant Na + -ATPase. The important simultaneous upregulation of PKC ζ, which is associated with a ceramide-induced inhibition of Na + -ATPase [49], can be viewed as a compensatory counter-regulatory -but not sufficientmechanism that opposes to PKC α, ε and λ. Downregulation of PKA (Fig. 7) would also contribute to activation of the pump because this kinase has been implicated in 2 different inhibitory pathways of Na + -ATPase, i.e.…”

Mechanisms Involving Ang II and MAPK/ERK1/2 Signaling Pathways Underlie Cardiac and Renal Alterations during Chronic Undernutrition

“…SDS-PAGE and imunoblotting for AT 1 R, AT 2 R, ERK1 and phospho-ERK1/2 were carried out as in [8], [36] using the specific antibodies. Briefly, the proteins of renal proximal tubule cells and cardiomyocytes were separated on 10% SDS-PAGE [37] and transferred to nitrocellulose membranes.…”

“…The activities of PKC and PKA were measured as in [8], [36].The activities of PKC and PKA associated with the isolated membranes of renal and cardiac origin were measured by incorporation of the γ-phosphoryl group of (γ- 32 P)ATP into histone in the absence and presence of their respective inhibitors: 10 nM calphostin C (Calbiochem) for PKC, 10 nM PKAi (5–24) peptide (Sigma-Aldrich) for PKA [8], [30], [36]. The reaction was started by adding (γ- 32 P)ATP (10 µM; specific activity ∼4.5 µCi/nmol) to the reaction medium (0.1 ml) containing 20 mM Hepes-Tris (pH 7.0), 4 mM MgCl 2 , 1.5 mg/ml histone 2S (Sigma-Aldrich) and 0.7 mg/ml membrane protein.…”

p53 and Ceramide as Collaborators in the Stress Response

“…Additionally, since SM gets synthesized in the cis and medial Golgi apparatus, the signaling pool of SM resides also in the inner leaflet of the plasma membrane or on the cytoplasmic face of a subcellular fraction (such as Golgi or endosomes) [36]. Indeed, many signaling lipid-regulated enzymes and lipases are associated with the inner leaflet of the plasma membrane; these include the ceramide-activated protein kinase C zeta (PKCz) [37] and ceramide-activated protein phosphatase (CAPP), as well as phospholipases C and A2 [35]. The exogenous use of bacterial sphingomyelinase allowed the examination of the role of outer leaflet sphingomyelin in inducing apoptotic signaling.…”