title: The 14-3-3 protein translates the NA+,K+-ATPase {alpha}1-subunit phosphorylation signal into binding and activation of phosphoinositide 3-kinase during endocytosis.
abstract: Clathrin-dependent endocytosis of Na(+),K(+)-ATPase molecules in response to G protein-coupled receptor signals is triggered by phosphorylation of the alpha-subunit and the binding of phosphoinositide 3-kinase. In this study, we describe a molecular mechanism linking phosphorylation of Na(+),K(+)-ATPase alpha-subunit to binding and activation of phosphoinositide 3-kinase. Co-immunoprecipitation studies, as well as experiments using confocal microscopy, revealed that dopamine favored the association of 14-3-3 protein with the basolateral plasma membrane and its co-localization with the Na(+),K(+)-ATPase alpha-subunit. The functional relevance of this interaction was established in opossum kidney cells expressing a 14-3-3 dominant negative mutant, where dopamine failed to decrease Na(+),K(+)-ATPase activity and to promote its endocytosis. The phosphorylated Ser-18 residue within the alpha-subunit N terminus is critical for 14-3-3 binding. Activation of phosphoinositide 3-kinase by dopamine during Na(+),K(+)-ATPase endocytosis requires the binding of the kinase to a proline-rich domain within the alpha-subunit, and this effect was blocked by the presence of a 14-3-3 dominant negative mutant. Thus, the 14-3-3 protein represents a critical linking mechanism for recruiting phosphoinositide 3-kinase to the site of Na(+),K(+)-ATPase endocytosis.
authors.affiliation: Department of Medicine, Atherosclerosis Research Unit, Membrane Signaling Networks, Karolinska Institutet, Karolinska University Hospital-Solna, S-171 76 Stockholm, Sweden.
authors.forename: Riad
authors.initials: R
authors.lastname: Efendiev
authors.forename: Zongpei
authors.initials: Z
authors.lastname: Chen
authors.forename: Rafael T
authors.initials: RT
authors.lastname: Krmar
authors.forename: Sabine
authors.initials: S
authors.lastname: Uhles
authors.forename: Adrian I
authors.initials: AI
authors.lastname: Katz
authors.forename: Carlos H
authors.initials: CH
authors.lastname: Pedemonte
authors.forename: Alejandro M
authors.initials: AM
authors.lastname: Bertorello
mesh_terms: D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs; D000818:Animals; D001665:Binding Sites; D004705:Endocytosis / Q000502:physiology*; D018014:Gene Transfer Techniques; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
chemical_list: D048948:14-3-3 Proteins; D019869:Phosphatidylinositol 3-Kinases; D043562:Receptors, G-Protein-Coupled; D000254:Sodium-Potassium-Exchanging ATPase