Structural Interactions between FXYD Proteins and Na+,K+-ATPase

Lindzen, Moshit; Gottschalk, Kay‐Eberhard; Füzesi, Maria; Garty, Haim; Karlish, Steven J. D.

doi:10.1074/jbc.m512063200

Cited by 29 publications

(17 citation statements)

References 57 publications

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“…Molecular and biochemical studies indicate that PLM and other FXYD proteins interact with TM2 and TM9 of Na + -K + -ATPase (31,45). Molecular modeling suggests that PLM docks into the groove between TM2, TM6 and TM9 of Na + -K + -ATPase.…”

Section: Discussionmentioning

confidence: 99%

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Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na⁺/Ca²⁺Exchanger

Wang¹,

Zhang²,

Ahlers³

et al. 2006

J. Biol. Chem.

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Phospholemman (PLM), a member of the FXYD family of small ion transport regulators, inhibits cardiac Na + /Ca 2+ exchanger (NCX1). NCX1 is made up of N-terminal domain consisting of the 1 st five transmembrane segments (residues 1-217), a large intracellular loop (residues 218-764), and a C-terminal domain comprising the last four transmembrane segments (residues 765-938). Using GST pulldown assay, we demonstrated that the intracellular loop, but not the N-or C-terminal transmembrane domains of NCX1, was associated with PLM. Further analysis using protein constructs of GST fused to various segments of the intracellular loop of NCX1 suggest that PLM bound to residues 218-371 and 508-764 but not 371-508. Split Na + /Ca 2+ exchangers consisting of N-or C-terminal domains with different lengths of the intracellular loop were co-expressed with PLM in HEK293 cells that are devoid of endogenous PLM and NCX1. While expression of Nterminal but not C-terminal domain alone resulted in correct membrane targeting, co-expression of both N-and C-terminal domains was required for correct membrane targeting and functional exchange activity. NCX1 current measurements indicate that PLM decreased NCX1 current only when the split exchangers contained residues 218-358 of the intracellular loop. Coimmunoprecipitation experiments with PLM and split exchangers suggest that PLM associated with the N-terminal domain of NCX1 when it contained intracellular loop residues 218-358. TM43, a PLM mutant with its cytoplasmic tail truncated, did not co-immunoprecipitate with wild-type NCX1 when co-expressed in HEK293 cells, confirming little to no interaction between the transmembrane domains of PLM and NCX1. We conclude that PLM interacted with the intracellular loop of NCX1, most likely at residues 218-358.

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Section: Discussionmentioning

confidence: 99%

“…It is known that the TM domain of PLM interacts with the TM domains of Na + -K + -ATPase (31). It is therefore of interest to determine if the TM domain of PLM is necessary for its interaction with NCX1.…”

Section: Association Of Ncx1 With Plm Requires the Cytoplasmic Domainmentioning

confidence: 99%

Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na⁺/Ca²⁺Exchanger

Wang¹,

Zhang²,

Ahlers³

et al. 2006

J. Biol. Chem.

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“…The Na ϩ /K ϩ -ATPase consists of a catalytic ␣-subunit with 10 transmembrane segments and a ␤-subunit involved in membrane insertion and as a modulator of transport properties (5). In particular, cross-linking and modeling studies revealed the direct interaction between the Na ϩ /K ϩ -ATPase ␣-subunit and members of the FXYD family (␥ and CHIF), which involves the binding of a single transmembrane domain into a groove formed by M2, M6, and M9 transmembrane helices of the Na ϩ /K ϩ -ATPase (6,7). PLM appears to be a major control point in the function of heart cells, although its specific physiological role is unclear, complicated by the possibility that its regulatory effect on the Na ϩ /K ϩ -ATPase may depend on the phosphorylation state of PLM (8) as well as other less well established functional roles (e.g.…”

Section: Human Phospholemman (Plm)mentioning

confidence: 99%

Phospholemman Transmembrane Structure Reveals Potential Interactions with Na+/K+-ATPase

Beevers

Kukol

2007

Journal of Biological Chemistry

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Phospholemman (PLM) is a 72-residue bitopic cardiac transmembrane protein, which acts as a modulator of the Na ؉ /K ؉ -ATPase and the Na ؉ /Ca 2؉ exchanger and possibly forms taurine channels in nonheart tissue. This work presents a high resolution structural model obtained from a combination of site-specific infrared spectroscopy and experimentally constrained high throughput molecular dynamics (MD) simulations. Altogether, 37 experimental constraints, including nine long range orientational constraints, have been used during MD simulations in an explicit lipid bilayer/water system. The resulting tetrameric ␣-helical bundle has an average helix tilt of 7.3°a nd a crossing angle close to 0°. It does not reveal a hydrophilic pore, but instead strong interactions between various residues occlude any pore. The helix-helix packing is unusual, with Gly Human phospholemman (PLM)2 is a member of a family of single-span transmembrane proteins characterized by the invariant extracellular motif FXYD (1) and is also known as FXYD1. It is found in liver, skeletal muscle, and most abundantly in the cardiac sarcolemma (2). Over recent years, conclusive experimental evidence has been presented that PLM acts as a tissue-specific modulator of the Na ϩ /K ϩ -ATPase similar to the other members of the FXYD family, each of which is prevalent in a different tissue, summarized in several recent reviews (3, 4). The Na ϩ /K ϩ -ATPase consists of a catalytic ␣-subunit with 10 transmembrane segments and a ␤-subunit involved in membrane insertion and as a modulator of transport properties (5). In particular, cross-linking and modeling studies revealed the direct interaction between the Na ϩ /K ϩ -ATPase ␣-subunit and members of the FXYD family (␥ and CHIF), which involves the binding of a single transmembrane domain into a groove formed by M2, M6, and M9 transmembrane helices of the Na ϩ /K ϩ -ATPase (6, 7). PLM appears to be a major control point in the function of heart cells, although its specific physiological role is unclear, complicated by the possibility that its regulatory effect on the Na ϩ /K ϩ -ATPase may depend on the phosphorylation state of PLM (8) as well as other less well established functional roles (e.g. interaction with the Na/Ca 2ϩ exchanger (9, 10) or independent channel formation). PLM knock-out mice expressed a complex response, including increased cardiac mass, larger cardiomyocytes, and ejection fractions in the absence of hypertension, whereas the overall Na ϩ /K ϩ -ATPase activity was reduced by 50% (11). There is growing evidence that PLM regulates the activity of Na/Ca 2ϩ exchanger 1 (NCX1) from heterologous expression studies of PLM and NCX1 in HEK239 cells (10) and overexpression of PLM in rat cardiomyocytes (12, 13) using different approaches to measure NCX1 activity. It appears that phosphorylation of PLM abolishes its inhibitory effect on the Na ϩ /K ϩ -ATPase (14, 15), whereas the phosphorylated form of PLM inhibits NCX1 (9, 12).Electrical measurements of PLM in artificial lipid bilayers and oocytes sh...

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“…In some experiments cells were maintained in a K ϩ -free RPMI 1640 medium (Biological Industries). Rat ␣1-Na ϩ /K ϩ -ATPase-expressing cell clones were produced by transfecting the above YFP-␣1 cells with rat ␣1 tagged in its N terminus by the hemagglutinin A (HA) epitope subcloned into pcDNA-3 (21). Transfected cells were cultivated in the presence of 10 M ouabain, and stably transfected ouabain-resistant cell clones were isolated.…”

Section: Methodsmentioning

confidence: 99%

Ouabain-induced Internalization and Lysosomal Degradation of the Na+/K+-ATPase

Cherniavsky-Lev

Golani

Karlish

et al. 2014

Journal of Biological Chemistry

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Structural Interactions between FXYD Proteins and Na+,K+-ATPase

Abstract: Interactions of rat FXYD4 (corticosteroid hormone-induced factor (CHIF)), FXYD2 (␥), or FXYD1 (phospholemman (PLM))

Cited by 29 publications

References 57 publications

Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na⁺/Ca²⁺Exchanger

Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na⁺/Ca²⁺Exchanger

Phospholemman Transmembrane Structure Reveals Potential Interactions with Na+/K+-ATPase

Ouabain-induced Internalization and Lysosomal Degradation of the Na+/K+-ATPase

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Structural Interactions between FXYD Proteins and Na+,K+-ATPase

Abstract: Interactions of rat FXYD4 (corticosteroid hormone-induced factor (CHIF)), FXYD2 (␥), or FXYD1 (phospholemman (PLM))

Cited by 29 publications

References 57 publications

Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na+/Ca2+Exchanger

Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na+/Ca2+Exchanger

Phospholemman Transmembrane Structure Reveals Potential Interactions with Na+/K+-ATPase

Ouabain-induced Internalization and Lysosomal Degradation of the Na+/K+-ATPase

Contact Info

Product

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Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na⁺/Ca²⁺Exchanger

Cytoplasmic Tail of Phospholemman Interacts with the Intracellular Loop of the Cardiac Na⁺/Ca²⁺Exchanger