Intracellular Na+ Regulates Dopamine and Angiotensin II Receptors Availability at the Plasma Membrane and Their Cellular Responses in Renal Epithelia

Abstract

The balance and cross-talk between natruretic and antinatruretic hormone receptors plays a critical role in the regulation of renal Na ؉ homeostasis, which is a major determinant of blood pressure. Dopamine and angiotensin II have antagonistic effects on renal Na ؉ and water excretion, which involves regulation of the Na ؉ ,K ؉ -ATPase activity. Herein we demonstrate that angiotensin II (Ang II) stimulation of AT1 receptors in proximal tubule cells induces the recruitment of Na ؉ ,K ؉ -ATPase molecules to the plasmalemma, in a process mediated by protein kinase C␤ and interaction of the Na ؉ ,K ؉ -ATPase with adaptor protein 1. Ang II stimulation led to phosphorylation of the ␣ subunit Ser-11 and Ser-18 residues, and substitution of these amino acids with alanine residues completely abolished the Ang II-induced stimulation of Na ؉ ,K ؉ -ATPase-mediated Rb ؉ transport. Thus, for Ang II-dependent stimulation of Na ؉ ,K ؉ -ATPase activity, phosphorylation of these serine residues is essential and may constitute a triggering signal for recruitment of Na ؉ ,K ؉ -ATPase molecules to the plasma membrane. When cells were treated simultaneously with saturating concentrations of dopamine and Ang II, either activation or inhibition of the Na ؉ ,K ؉ -ATPase activity was produced dependent on the intracellular Na ؉ concentration, which was varied in a very narrow physiological range (9 -19 mM). A small increase in intracellular Na ؉ concentrations induces the recruitment of D1 receptors to the plasma membrane and a reduction in plasma membrane AT1 receptors. Thus, one or more proteins may act as an intracellular Na ؉ concentration sensor and play a major regulatory role on the effect of hormones that regulate proximal tubule Na ؉ reabsorption.

G-protein-coupled Receptor-mediated Traffic of Na,K-ATPase to the Plasma Membrane Requires the Binding of Adaptor Protein 1 to a Tyr-255-based Sequence in the α-Subunit

“…We have previously demonstrated that stimulation of AT1Rs induces the phosphorylation of Ser-11 and Ser-18 of NKA ␣1 and that mutation of these amino acids prevents the Ang II-dependent regulation of NKA activity (16,17). Consistent with this, Fig.…”

“…For this, we used the technique of plasma membrane protein biotinylation, which has been validated in several of our previous publications (16,21,24). After treatment of the cells with either PMA or Ang II, the cells were rapidly transferred to a 4°C bath, the temperature at which the plasma membrane proteins were labeled with N-hydroxysulfosuccinimide-biotin.…”

“…These results, as well as the observation that preventing the Ang II-induced phosphorylation of NKA ␣1 impairs the NKA ␣1/AP-1 interaction, support the conclusion that the NKA ␣1/AP-1 interaction is a very specific response to the activation of AT1Rs. Ang II-dependent plasma membrane recruitment of NKA molecules requires the protein kinase C␤-mediated phosphorylation of NKA ␣1 (16). The fact that the Ang II-dependent increment of FRET N was impaired when the cells were treated with the protein kinase C inhibitor chelerythrine chloride is evidence of the causal link between the specific phosphorylation of ␣1 and the NKA/AP-1 interaction.…”

“…After separation, the agarose beads were washed four times with IP buffer containing 0.2% Triton X-100 and 0.2% SDS and once with 50 mM Tris-HCl, pH 7.4, and finally resuspended in Laemmli sample buffer. Electrophoresis, Western blot analysis with ExtrAvidin peroxidase conjugate, and densitometric analysis were performed as described previously (16,21,24).…”

The 14-3-3 Protein Translates the NA+,K+-ATPase α1-Subunit Phosphorylation Signal into Binding and Activation of Phosphoinositide 3-Kinase during Endocytosis

“…Thus, a balance between the increase or decrease in the number of copies within the plasma membrane determines the cell Na ϩ ,K ϩ -ATPase activity. The responses to different agonists regulating the intracellular distribution of Na ϩ ,K ϩ -ATPase are tissue-specific (3), and in renal tubule cells, the direction of the physiological response (increase versus decrease) in response to hormones that influence urinary sodium excretion appears to depend largely on the concentration of intracellular sodium (4,5).…”