SIK1/SOS2 networks: decoding sodium signals via calcium-responsive protein kinase pathways

Changes in cellular ion levels can modulate distinct signaling networks aimed at correcting major disruptions in ion balances that might otherwise threaten cell growth and development. Salt-inducible kinase 1 (SIK1) and salt overly sensitive 2 (SOS2) are key protein kinases within such networks in mammalian and plant cells, respectively. In animals, SIK1 expression and activity are regulated in response to the salt content of the diet, and in plants SOS2 activity is controlled by the salinity of the soil. The specific ionic stress (elevated intracellular sodium) is followed by changes in intracellular calcium; the calcium signals are sensed by calcium-binding proteins and lead to activation of SIK1 or SOS2. These kinases target major plasma membrane transporters such as the Na+,K+-ATPase in mammalian cells, and Na+/H+ exchangers in the plasma membrane and membranes of intracellular vacuoles of plant cells. Activation of these networks prevents abnormal increases in intracellular sodium concentration

Involvement of SIK3 in Glucose and Lipid Homeostasis in Mice

Salt-inducible kinase 3 (SIK3), an AMP-activated protein kinase-related kinase, is induced in the murine liver after the consumption of a diet rich in fat, sucrose, and cholesterol. To examine whether SIK3 can modulate glucose and lipid metabolism in the liver, we analyzed phenotypes of SIK3-deficent mice. Sik3−/− mice have a malnourished the phenotype (i.e., lipodystrophy, hypolipidemia, hypoglycemia, and hyper-insulin sensitivity) accompanied by cholestasis and cholelithiasis. The hypoglycemic and hyper-insulin-sensitive phenotypes may be due to reduced energy storage, which is represented by the low expression levels of mRNA for components of the fatty acid synthesis pathways in the liver. The biliary disorders in Sik3−/− mice are associated with the dysregulation of gene expression programs that respond to nutritional stresses and are probably regulated by nuclear receptors. Retinoic acid plays a role in cholesterol and bile acid homeostasis, wheras ALDH1a which produces retinoic acid, is expressed at low levels in Sik3−/− mice. Lipid metabolism disorders in Sik3−/− mice are ameliorated by the treatment with 9-cis-retinoic acid. In conclusion, SIK3 is a novel energy regulator that modulates cholesterol and bile acid metabolism by coupling with retinoid metabolism, and may alter the size of energy storage in mice

Multiple Polymorphisms Affect Expression and Function of the Neuropeptide S Receptor (NPSR1)

Peer reviewe

The Dopamine Paradox in Lung and Kidney Epithelia: Sharing the Same Target but Operating Different Signaling Networks

Stimulation of dopamine receptors in the lung or kidney epithelia has distinct and opposite effects on the function of Na,K-ATPase, which results in increased Na+ absorption across the alveolar epithelium and increased sodium excretion via the kidney epithelium. In the lung, dopamine increases Na,K-ATPase by increasing cell basolateral surface expression of Na+,K+-ATPase molecules, whereas in the kidney epithelia it decreases Na+,K+-ATPase activity by removing active units from the plasma membrane by endocytosis. The opposite effects of dopamine over the same target (the Na+,K+-ATPase) involve the activation of a distinct signaling network that it is target specific, and has a different spatial resolution. Understanding the specific signaling pathways involved in these actions of dopamine and their hierarchical organization may facilitate the drug discovery process that could lead to the design of new therapeutic approaches to clear lung edema in patients with acute lung injury and to decrease fluid overload during congestive heart failure and hypertension

Receptor-mediated inhibition of renal Na +

The mechanisms involved in receptor-mediated inhibition of Na + -K + -ATPase remain poorly understood. In this study, we evaluate whether inhibition of proximal tubule Na + -K + -ATPase activity by dopamine is linked to its removal from the plasma membrane and internalization into defined intracellular compartments. Clathrin-coated vesicles were isolated by sucrose gradient centrifugation and negative lectin selection, and early and late endosomes were separated on a flotation gradient. Inhibition of Na + -K + -ATPase activity by dopamine, in contrast to its inhibition by ouabain, was accompanied by a sequential increase in the abundance of the α-subunit in clathrin-coated vesicles (1 min), early endosomes (2.5 min), and late endosomes (5 min), suggesting its stepwise translocation between these organelles. A similar pattern was found for the β-subunit. The increased incorporation of both subunits in all compartments was blocked by calphostin C. The results demonstrate that the dopamine-induced decrease in Na + -K + -ATPase activity in proximal tubules is associated with internalization of its α- and β-subunits into early and late endosomes via a clathrin-dependent pathway and that this process is protein kinase C dependent. The presence of Na + -K + -ATPase subunits in endosomes suggests that these compartments may constitute normal traffic reservoirs during pump degradation and/or synthesis

Phosphatidylinositol 3-Kinase-mediated Endocytosis of Renal Na(+),K(+)-ATPase α Subunit in Response to Dopamine

Dopamine (DA) inhibition of Na(+),K(+)-ATPase in proximal tubule cells is associated with increased endocytosis of its α and β subunits into early and late endosomes via a clathrin vesicle-dependent pathway. In this report we evaluated intracellular signals that could trigger this mechanism, specifically the role of phosphatidylinositol 3-kinase (PI 3-K), the activation of which initiates vesicular trafficking and targeting of proteins to specific cell compartments. DA stimulated PI 3-K activity in a time- and dose-dependent manner, and this effect was markedly blunted by wortmannin and LY 294002. Endocytosis of the Na(+),K(+)-ATPase α subunit in response to DA was also inhibited in dose-dependent manner by wortmannin and LY 294002. Activation of PI 3-K generally occurs by association with tyrosine kinase receptors. However, in this study immunoprecipitation with a phosphotyrosine antibody did not reveal PI 3-K activity. DA-stimulated endocytosis of Na(+),K(+)-ATPase α subunits required protein kinase C, and the ability of DA to stimulate PI 3-K was blocked by specific protein kinase C inhibitors. Activation of PI 3-K is mediated via the D(1) receptor subtype and the sequential activation of phospholipase A(2), arachidonic acid, and protein kinase C. The results indicate a key role for activation of PI 3-K in the endocytic sequence that leads to internalization of Na(+),K(+)-ATPase α subunits in response to DA, and suggest a mechanism for the participation of protein kinase C in this process

Hypoxia-induced endocytosis of Na,K-ATPase in alveolar epithelial cells is mediated by mitochondrial reactive oxygen species and PKC-ζ

During ascent to high altitude and pulmonary edema, the alveolar epithelial cells (AEC) are exposed to hypoxic conditions. Hypoxia inhibits alveolar fluid reabsorption and decreases Na,K-ATPase activity in AEC. We report here that exposure of AEC to hypoxia induced a time-dependent decrease of Na,K-ATPase activity and a parallel decrease in the number of Na,K-ATPase α(1) subunits at the basolateral membrane (BLM), without changing its total cell protein abundance. These effects were reversible upon reoxygenation and specific, because the plasma membrane protein GLUT1 did not decrease in response to hypoxia. Hypoxia caused an increase in mitochondrial reactive oxygen species (ROS) levels that was inhibited by antioxidants. Antioxidants prevented the hypoxia-mediated decrease in Na,K-ATPase activity and protein abundance at the BLM. Hypoxia-treated AEC deficient in mitochondrial DNA (ρ(0) cells) did not have increased levels of ROS, nor was the Na,K-ATPase activity inhibited. Na,K-ATPase α(1) subunit was phosphorylated by PKC in hypoxia-treated AEC. In AEC treated with a PKC-ζ antagonist peptide or with the Na,K-ATPase α(1) subunit lacking the PKC phosphorylation site (Ser-18), hypoxia failed to decrease Na,K-ATPase abundance and function. Accordingly, we provide evidence that hypoxia decreases Na,K-ATPase activity in AEC by triggering its endocytosis through mitochondrial ROS and PKC-ζ–mediated phosphorylation of the Na,K-ATPase α(1) subunit

Phosphoinositide-3 kinase binds to a proline-rich motif in the Na(+),K(+)-ATPase α subunit and regulates its trafficking

Endocytosis of Na(+),K(+)-ATPase molecules in response to G protein-coupled receptor stimulation requires activation of class I(A) phosphoinositide-3 kinase (PI3K-I(A)) in a protein kinase C-dependent manner. In this paper, we report that PI3K-I(A), through its p85α subunit-SH3 domain, binds to a proline-rich region in the Na(+),K(+)-ATPase catalytic α subunit. This interaction is enhanced by protein kinase C-dependent phosphorylation of a serine residue that flanks the proline-rich motif in the Na(+),K(+)-ATPase α subunit and results in increased PI3K-I(A) activity, an effect necessary for adaptor protein 2 binding and clathrin recruitment. Thus, Ser-phosphorylation of the Na(+),K(+)-ATPase catalytic subunit serves as an anchor signal for regulating the location of PI3K-I(A) and its activation during Na(+),K(+)-ATPase endocytosis in response to G protein-coupled receptor signals

Phosphorylation of the Catalyic α-Subunit Constitutes a Triggering Signal for Na+,K+-ATPase Endocytosis

Inhibition of Na+,K+-ATPase activity by dopamine is an important mechanism by which renal tubules modulate urine sodium excretion during a high salt diet. However, the molecular mechanisms of this regulation are not clearly understood. Inhibition of Na+,K+-ATPase activity in response to dopamine is associated with endocytosis of its α- and β-subunits, an effect that is protein kinase C-dependent. In this study we used isolated proximal tubule cells and a cell line derived from opossum kidney and demonstrate that dopamine-induced endocytosis of Na+,K+-ATPase and inhibition of its activity were accompanied by phosphorylation of the α-subunit. Inhibition of both the enzyme activity and its phosphorylation were blocked by the protein kinase C inhibitor bisindolylmaleimide. The early time dependence of these processes suggests a causal link between phosphorylation and inhibition of enzyme activity. However, after 10 min of dopamine incubation, the α-subunit was no longer phosphorylated, whereas enzyme activity remained inhibited due to its removal from the plasma membrane. Dephosphorylation occurred in the late endosomal compartment. To further examine whether phosphorylation was a prerequisite for subunit endocytosis, we used the opossum kidney cell line transfected with the rodent α-subunit cDNA. Treatment of this cell line with dopamine resulted in phosphorylation and endocytosis of the α-subunit with a concomitant decrease in Na+,K+-ATPase activity. In contrast, none of these effects were observed in cells transfected with the rodent α-subunit that lacks the putative protein kinase C-phosphorylation sites (Ser11 and Ser18). Our results support the hypothesis that protein kinase C-dependent phosphorylation of the α-subunit is essential for Na+,K+-ATPase endocytosis and that both events are responsible for the decreased enzyme activity in response to dopamine