TY - JOUR
T1 - Na,K-ATPase α1-subunit dephosphorylation by protein phosphatase 2A is necessary for its recruitment to the plasma membrane
AU - Lecuona, Emilia
AU - Dada, Laura
AU - Sun, Haiying
AU - Butti, Maria L.
AU - Zhou, Guofei
AU - Chew, Teng Leong
AU - Sznajder, Jacob I
PY - 2006/12
Y1 - 2006/12
N2 - In alveolar epithelial cells, G-protein coupled-receptors agonists (GPCR) induce the recruitment of the Na,K-ATPase to the plasma membrane. Here we report that for the recruitment of the Na,K-ATPase to occur, dephosphorylation of its α1-subunit at serine 18 is necessary, as demonstrated by in vitro phosphorylation, mutation of the serine 18 to alanine, and use of a specific phospho-antibody. Several approaches strongly suggest dephosphorylation to be mediated by protein phosphatase 2A (PP2A): 1) Na,K-ATPase dephosphorylation and recruitment were prevented by okadaic acid (OA); 2) the Na,K-ATPase α1-subunit is an in vitro substrate for PP2A; and 3) glutathione S-transferase (GST)-fusion proteins binding assays demonstrate a direct interaction between the catalytic subunit of PP2A and the first 90 amino acids of the Na,K-ATPase α1-subunit. Finally, GPCR agonists induced a rapid translocation of PP2A from the cytosol to the membrane fraction, which corresponded with increased coimmunoprecipitation and colocalization of PP2A and the Na,K-ATPase. Accordingly, we provide evidence that GPCR agonists promote PP2A translocation to the membrane fraction, leading to the dephosphorylation of the Na,K-ATPase α1-subunit at the serine 18 residue and its recruitment to the cell plasma membrane, which is of biological and physiological importance.
AB - In alveolar epithelial cells, G-protein coupled-receptors agonists (GPCR) induce the recruitment of the Na,K-ATPase to the plasma membrane. Here we report that for the recruitment of the Na,K-ATPase to occur, dephosphorylation of its α1-subunit at serine 18 is necessary, as demonstrated by in vitro phosphorylation, mutation of the serine 18 to alanine, and use of a specific phospho-antibody. Several approaches strongly suggest dephosphorylation to be mediated by protein phosphatase 2A (PP2A): 1) Na,K-ATPase dephosphorylation and recruitment were prevented by okadaic acid (OA); 2) the Na,K-ATPase α1-subunit is an in vitro substrate for PP2A; and 3) glutathione S-transferase (GST)-fusion proteins binding assays demonstrate a direct interaction between the catalytic subunit of PP2A and the first 90 amino acids of the Na,K-ATPase α1-subunit. Finally, GPCR agonists induced a rapid translocation of PP2A from the cytosol to the membrane fraction, which corresponded with increased coimmunoprecipitation and colocalization of PP2A and the Na,K-ATPase. Accordingly, we provide evidence that GPCR agonists promote PP2A translocation to the membrane fraction, leading to the dephosphorylation of the Na,K-ATPase α1-subunit at the serine 18 residue and its recruitment to the cell plasma membrane, which is of biological and physiological importance.
KW - Alveolar epithelial cells
KW - G-protein-coupled receptor agonists
KW - Intracellular trafficking
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U2 - 10.1096/fj.06-6503fje
DO - 10.1096/fj.06-6503fje
M3 - Article
C2 - 17065225
AN - SCOPUS:33845625148
SN - 0892-6638
VL - 20
SP - E2146-E2155
JO - FASEB Journal
JF - FASEB Journal
IS - 14
ER -