Protein Phosphatase 2A Interacts with the Na+,K+-ATPase and Modulates Its Trafficking by Inhibition of Its Association with Arrestin

Background: The P-type ATPase family constitutes a collection of ion pumps that form phosphorylated intermediates during ion transport. One of the best known members of this family is the Na +,K +-ATPase. The catalytic subunit of the Na +,K +-ATPase includes several functional domains that determine its enzymatic and trafficking properties. Methodology/Principal Findings: Using the yeast two-hybrid system we found that protein phosphatase 2A (PP2A) catalytic C-subunit is a specific Na +,K +-ATPase interacting protein. PP-2A C-subunit interacted with the Na +,K +-ATPase, but not with the homologous sequences of the H +,K +-ATPase. We confirmed that the Na +,K +-ATPase interacts with a complex of A- and C-subunits in native rat kidney. Arrestins and G-protein coupled receptor kinases (GRKs) are important regulators of G-protein coupled receptor (GPCR) signaling, and they also regulate Na +,K +-ATPase trafficking through direct association. PP2A inhibits association between the Na +,K +-ATPase and arrestin, and diminishes the effect of arrestin on Na +,K +-ATPase trafficking. GRK phosphorylates the Na +,K +-ATPase and PP2A can at least partially reverse this phosphorylation. Conclusions/Significance: Taken together, these data demonstrate that the sodium pump belongs to a growing list of io

Are there several isoforms of Na,K-ATPase alpha subunit in the rabbit kidney?

International audiencePrevious pharmacologic and kinetic studies have demonstrated the axial heterogeneity of the rabbit kidney tubule with regard to Na,K-ATPase. To evaluate whether this heterogeneity might reflect the presence of distinct isoforms of the alpha subunit of Na,K-ATPase, we used two monoclonal antibodies, IIC9 and IIE2 (G8), specific for the alpha 1 and alpha 3 isoforms, respectively, as probes for changes in the specific activity of Na,K-ATPase at the level of successive segments of the rabbit nephron. Single, well defined nephron segments were obtained by microdissection of collagenase-treated kidney. Results indicate that IIC9 antibody inhibited Na,K-ATPase activity by > 90% in all the segments of the nephron except the collecting duct. Conversely, IIE2 (G8) antibody abolished Na,K-ATPase activity in the collecting duct, whereas it had no effect in other nephron segments. These findings suggest that the rabbit collecting duct preferentially expresses a distinct isoform of Na,K-ATPase catalytic subunit, which is presumably alpha 3-like, in agreement with previous pharmacologic and kinetic observations, whereas other nephron segments would express the alpha 1 isoform

Oligodendrocytes in brain and optic nerve express the β3 subunit isoform of Na,K-ATPase

The Na,K-ATPase, which catalyzes the active transport of Na+ and K+, has two principal subunits (α and β) that have several genetically distinct isoforms. Most of these isoforms are expressed in the nervous system, but certain ones are preferentially expressed in glia and others in neurons. Of the β isoforms, β1 predominates in neurons and β2 in astrocytes, although there are some exceptions. Here we demonstrate that β3 is expressed in rat and mouse white matter oligodendrocytes. Immunofluorescence microscopy identified β3 in oligodendrocytes of rat brain white matter in typical linear arrays of cell bodies between fascicles of axons. The intensity of stain peaked at 20 postnatal days. β3 was identified in cortical oligodendrocytes grown in culture, where it was expressed in processes and colocalized with antibody to galactocerebroside. In the mouse and rat optic nerve, β3 stain was seen in oligodendrocytes, where it colocalized with carbonic anhydrase II. For comparison, optic nerve was stained for the β1 and β2 subunits, showing distinct patterns of labelling of axons (β1) and astrocytes (β2). The C6 glioma cell line was also found to express the β3 isoform preferentially. Since β3 was not found at detectable levels in astrocytes, this suggests that C6 is closer to oligodendrocytes than astrocytes in the glial cell lineage. (C) 2000 Wiley-Liss, Inc.Peer Reviewe

Presence of two isoforms of Na, K-ATPase with different pharmacological and immunological properties in the rat kidney

International audiencePrevious studies have demonstrated the presence of two populations of Na,K-ATPase with distinct kinetic, pharmacological and immunological characteristics along the rabbit nephron, indicating that the proximal segments of the nephron express exclusively the alpha 1 isoform of the catalytic subunit, whereas the collecting duct expresses an alpha 3-like isoform. Because pharmacological studies have shown the existence of two populations of Na,K-ATPase with different sensitivities to ouabain in the rat cortical collecting duct, which may result from the presence in the same nephron segment of the two isoforms demonstrated in the different segments of the rabbit nephron, the present study was undertaken to characterize the properties of Na,K-ATPase along the rat nephron. Results indicate that each segment of the rat nephron contains two subpopulations of Na,K-ATPase: a component highly sensitive to ouabain (IC50 approximately 5.10(-6) M) which is recognized by an anti-alpha 3 antibody and another moiety of lower affinity for ouabain (IC50 approximately 5.10(-4) M) which is recognized by an anti-alpha 1 antibody. Whether these two subpopulations correspond to different isoforms of the alpha subunit of Na,K-ATPase (alpha 1 and alpha 3-like) remains to be determined