Function of FXYD proteins, regulators of Na, K-ATPase

In this short review, we summarize our work on the role of members of the FXYD protein family as tissue-specific modulators of Na, K-ATPase. FXYD1 or phospholemman, mainly expressed in heart and skeletal muscle increases the apparent affinity for intracellular Na(+) of Na, K-ATPase and may thus be important for appropriate muscle contractility. FXYD2 or gamma subunit and FXYD4 or CHIF modulate the apparent affinity for Na(+) of Na, K-ATPase in an opposite way, adapted to the physiological needs of Na(+) reabsorption in different segments of the renal tubule. FXYD3 expressed in stomach, colon, and numerous tumors also modulates the transport properties of Na, K-ATPase but it has a lower specificity of association than other FXYD proteins and an unusual membrane topology. Finally, FXYD7 is exclusively expressed in the brain and decreases the apparent affinity for extracellular K(+), which may be essential for proper neuronal excitability

The third sodium binding site of Na,K-ATPase is functionally linked to acidic pH-activated inward current

Sodium- and potassium-activated adenosine triphosphatases (Na,K-ATPase) is the ubiquitous active transport system that maintains the Na(+) and K(+) gradients across the plasma membrane by exchanging three intracellular Na(+) ions against two extracellular K(+) ions. In addition to the two cation binding sites homologous to the calcium site of sarcoplasmic and endoplasmic reticulum calcium ATPase and which are alternatively occupied by Na(+) and K(+) ions, a third Na(+)-specific site is located close to transmembrane domains 5, 6 and 9, and mutations close to this site induce marked alterations of the voltage-dependent release of Na(+) to the extracellular side. In the absence of extracellular Na(+) and K(+), Na,K-ATPase carries an acidic pH-activated, ouabain-sensitive "leak" current. We investigated the relationship between the third Na(+) binding site and the pH-activated current. The decrease (in E961A, T814A and Y778F mutants) or the increase (in G813A mutant) of the voltage-dependent extracellular Na(+) affinity was paralleled by a decrease or an increase in the pH-activated current, respectively. Moreover, replacing E961 with oxygen-containing side chain residues such as glutamine or aspartate had little effect on the voltage-dependent affinity for extracellular Na(+) and produced only small effects on the pH-activated current. Our results suggest that extracellular protons and Na(+) ions share a high field access channel between the extracellular solution and the third Na(+) binding site

Role of homologous ASP334 and GLU319 in human non-gastric H,K- and Na,K-ATPases in cardiac glycoside binding

Cardiac steroids inhibit Na,K-ATPase and the related non-gastric H,K-ATPase, while they do not interact with gastric H,K-ATPase. Introducing an arginine, the residue present in the gastric H,K-ATPase, in the second extracellular loop at the corresponding position 334 in the human non-gastric H,K-ATPase (D334R mutation) rendered it completely resistant to 2mM ouabain. The corresponding mutation (E319R) in alpha1 Na,K-ATPase produced a approximately 2-fold increase of the ouabain IC(50) in the ouabain-resistant rat alpha1 Na,K-ATPase and a large decrease of the ouabain affinity of human alpha1 Na,K-ATPase, on the other hand this mutation had no effect on the affinity for the aglycone ouabagenin. These results provide a strong support for the orientation of ouabain in its biding site with its sugar moiety interacting directly with the second extracellular loop

A novel family of transmembrane proteins interacting with beta subunits of the Na,K-ATPase

We characterized a family consisting of four mammalian proteins of unknown function (NKAIN1, 2, 3 and 4) and a single Drosophila ortholog dNKAIN. Aside from highly conserved transmembrane domains, NKAIN proteins contain no characterized functional domains. Striking amino acid conservation in the first two transmembrane domains suggests that these proteins are likely to function within the membrane bilayer. NKAIN family members are neuronally expressed in multiple regions of the mouse brain, although their expression is not ubiquitous. We demonstrate that mouse NKAIN1 interacts with the beta1 subunit of the Na,K-ATPase, whereas Drosophila ortholog dNKAIN interacts with Nrv2.2, a Drosophila homolog of the Na,K-ATPase beta subunits. We also show that NKAIN1 can form a complex with another beta subunit-binding protein, MONaKA, when binding to the beta1 subunit of the Na,K-ATPase. Our results suggest that a complex between mammalian NKAIN1 and MONaKA is required for NKAIN function, which is carried out by a single protein, dNKAIN, in Drosophila. This hypothesis is supported by the fact that dNKAIN, but not NKAIN1, induces voltage-independent amiloride-insensitive Na(+)-specific conductance that can be blocked by lanthanum. Drosophila mutants with decreased dNKAIN expression due to a P-element insertion in the dNKAIN gene exhibit temperature-sensitive paralysis, a phenotype also caused by mutations in the Na,K-ATPase alpha subunit and several ion channels. The neuronal expression of NKAIN proteins, their membrane localization and the temperature-sensitive paralysis of NKAIN Drosophila mutants strongly suggest that this novel protein family may be critical for neuronal function

Chronic Nicotine Modifies Skeletal Muscle Na,K-ATPase Activity through Its Interaction with the Nicotinic Acetylcholine Receptor and Phospholemman

Our previous finding that the muscle nicotinic acetylcholine receptor (nAChR) and the Na,K-ATPase interact as a regulatory complex to modulate Na,K-ATPase activity suggested that chronic, circulating nicotine may alter this interaction, with long-term changes in the membrane potential. To test this hypothesis, we chronically exposed rats to nicotine delivered orally for 21–31 days. Chronic nicotine produced a steady membrane depolarization of ∼3 mV in the diaphragm muscle, which resulted from a net change in electrogenic transport by the Na,K-ATPase α2 and α1 isoforms. Electrogenic transport by the α2 isoform increased (+1.8 mV) while the activity of the α1 isoform decreased (−4.4 mV). Protein expression of Na,K-ATPase α1 or α2 isoforms and the nAChR did not change; however, the content of α2 subunit in the plasma membrane decreased by 25%, indicating that its stimulated electrogenic transport is due to an increase in specific activity. The physical association between the nAChR, the Na,K-ATPase α1 or α2 subunits, and the regulatory subunit of the Na,K-ATPase, phospholemman (PLM), measured by co-immuno precipitation, was stable and unchanged. Chronic nicotine treatment activated PKCα/β2 and PKCδ and was accompanied by parallel increases in PLM phosphorylation at Ser63 and Ser68. Collectively, these results demonstrate that nicotine at chronic doses, acting through the nAChR-Na,K-ATPase complex, is able to modulate Na,K-ATPase activity in an isoform-specific manner and that the regulatory range includes both stimulation and inhibition of enzyme activity. Cholinergic modulation of Na,K-ATPase activity is achieved, in part, through activation of PKC and phosphorylation of PLM

Regulation of the Cardiac Na+/K+ ATPase by Phospholemman

Hansraj Dhayan, Rajender Kumar, Andreas Kukol, ‘Regulation of the Cardiac Na+/K+ ATPase by Phospholemman’, in Sajal Chakraborti, Naranjan Dhalla, eds., Regulation of Membrane Na+-K+ ATPase, (Switzerland: Springer, 2016), ISBN 978-3-319-24748-9, eISBN 978-3-319-24750-2.Peer reviewe

Geometric Approach to Pontryagin's Maximum Principle

Since the second half of the 20th century, Pontryagin's Maximum Principle has been widely discussed and used as a method to solve optimal control problems in medicine, robotics, finance, engineering, astronomy. Here, we focus on the proof and on the understanding of this Principle, using as much geometric ideas and geometric tools as possible. This approach provides a better and clearer understanding of the Principle and, in particular, of the role of the abnormal extremals. These extremals are interesting because they do not depend on the cost function, but only on the control system. Moreover, they were discarded as solutions until the nineties, when examples of strict abnormal optimal curves were found. In order to give a detailed exposition of the proof, the paper is mostly self\textendash{}contained, which forces us to consider different areas in mathematics such as algebra, analysis, geometry.Comment: Final version. Minors changes have been made. 56 page