The <i>O</i>-glycosylated ectodomain of FXYD5 impairs adhesion by disrupting cell–cell trans-dimerization of Na,K-ATPase β1 subunits

Tokhtaeva, Elmíra; Sun, He; Deiss-Yehiely, Nimrod; Wen, Yi; Soni, Pritin N.; Gabrielli, Nieves M.; Marcus, Elizabeth A.; Ridge, Karen M.; Sachs, George; Vazquez‐Levin, Mónica H.; Sznajder, Jacob I.; Vagin, Olga; Dada, Laura A.

doi:10.1242/jcs.186148

Cited by 23 publications

(39 citation statements)

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“…A functional NKA requires a catalytic α-subunit and a regulatory β-subunit [2]. Additionally, a γ-subunit has also been identified, which represents a family of single-span transmembrane proteins containing the FXYD motif that is not an integral part of the transporter but rather regulates the activity and membrane abundance of the enzyme [3,4]. In the alveolar epithelium of the lung, the activity of NKA creates an Na + gradient that drives reabsorption of fluid from the alveolar space, which keeps the alveoli relatively "dry," which is essential for an effective gas exchange.…”

Section: Introductionmentioning

confidence: 99%

Hypercapnia Impairs Na,K-ATPase Function by Inducing Endoplasmic Reticulum Retention of the β-Subunit of the Enzyme in Alveolar Epithelial Cells

Kryvenko

Wessendorf

Morty

et al. 2020

IJMS

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Alveolar edema, impaired alveolar fluid clearance, and elevated CO2 levels (hypercapnia) are hallmarks of the acute respiratory distress syndrome (ARDS). This study investigated how hypercapnia affects maturation of the Na,K-ATPase (NKA), a key membrane transporter, and a cell adhesion molecule involved in the resolution of alveolar edema in the endoplasmic reticulum (ER). Exposure of human alveolar epithelial cells to elevated CO2 concentrations caused a significant retention of NKA-β in the ER and, thus, decreased levels of the transporter in the Golgi apparatus. These effects were associated with a marked reduction of the plasma membrane (PM) abundance of the NKA-α/β complex as well as a decreased total and ouabain-sensitive ATPase activity. Furthermore, our study revealed that the ER-retained NKA-β subunits were only partially assembled with NKA α-subunits, which suggests that hypercapnia modifies the ER folding environment. Moreover, we observed that elevated CO2 levels decreased intracellular ATP production and increased ER protein and, particularly, NKA-β oxidation. Treatment with α-ketoglutaric acid (α-KG), which is a metabolite that has been shown to increase ATP levels and rescue mitochondrial function in hypercapnia-exposed cells, attenuated the deleterious effects of elevated CO2 concentrations and restored NKA PM abundance and function. Taken together, our findings provide new insights into the regulation of NKA in alveolar epithelial cells by elevated CO2 levels, which may lead to the development of new therapeutic approaches for patients with ARDS and hypercapnia.

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Section: Introductionmentioning

confidence: 99%

Hypercapnia Impairs Na,K-ATPase Function by Inducing Endoplasmic Reticulum Retention of the β-Subunit of the Enzyme in Alveolar Epithelial Cells

Kryvenko

Wessendorf

Morty

et al. 2020

IJMS

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“…Despite this essential task, Na/K‐ATPases also mediate intercellular adhesion and induce activation of intracellular signalling pathways upon binding of glycoside hormones such as ouabain . Members of the FXYD family, a class of Na/K‐ATPase‐binding proteins , were reported to be important regulators of the Na/K‐ATPase, modulating its pump activity and mediation of intercellular adhesion . Similar to FXYD proteins, one could consider retinoschisin to exert a role as a modulator of Na/K‐ATPase activity.…”

Section: Introductionmentioning

confidence: 99%

The X‐linked juvenile retinoschisis protein retinoschisin is a novel regulator of mitogen‐activated protein kinase signalling and apoptosis in the retina

Plössl

Weber

Friedrich

2016

J Cellular Molecular Medi

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X‐linked juvenile retinoschisis (XLRS) is a hereditary retinal dystrophy in young males, caused by mutations in the RS1 gene. The function of the encoded protein, termed retinoschisin, and the molecular mechanisms underlying XLRS pathogenesis are still unresolved, although a direct interaction partner of the secreted retinoschisin, the retinal Na/K‐ATPase, was recently identified. Earlier gene expression studies in retinoschisin‐deficient (Rs1h −/Y) mice provided a first indication of pathological up‐regulation of mitogen‐activated protein (MAP) kinase signalling in disease pathogenesis. To further investigate the role for retinoschisin in MAP kinase regulation, we exposed Y‐79 cells and murine Rs1h −/Y retinae to recombinant retinoschisin and the XLRS‐associated mutant RS1‐C59S. Although normal retinoschisin stably bound to retinal cells, RS1‐C59S exhibited a strongly reduced binding affinity. Simultaneously, exposure to normal retinoschisin significantly reduced phosphorylation of C‐RAF and MAP kinases ERK1/2 in Y‐79 cells and murine Rs1h −/Y retinae. Expression of MAP kinase target genes C‐FOS and EGR1 was also down‐regulated in both model systems. Finally, retinoschisin treatment decreased pro‐apoptotic BAX‐2 transcript levels in Y‐79 cells and Rs1h −/Y retinae. Upon retinoschisin treatment, these cells showed increased resistance against apoptosis, reflected by decreased caspase‐3 activity (in Y‐79 cells) and increased photoreceptor survival (in Rs1h −/Y retinal explants). RS1‐C59S did not influence C‐RAF or ERK1/2 activation, C‐FOS or EGR1 expression, or apoptosis. Our data imply that retinoschisin is a novel regulator of MAP kinase signalling and exerts an anti‐apoptotic effect on retinal cells. We therefore discuss that disturbances of MAP kinase signalling by retinoschisin deficiency could be an initial step in XLRS pathogenesis.

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“…To determine whether LPS modulates FXYD5 levels in AEC, MLE-12 cells were treated with LPS for up to 24 h and cell culture media, cell lysates, and surface biotinylated plasma membrane (PM) proteins were collected. In the PM fraction of MLE-12 cells, FXYD5 was detected as a 60–70 kDa band (Figure 1A), suggesting that the plasmalemma-located FXYD5 is heavily O -glycosylated in these cells similar to that found in A549 cells (26). An additional 25 kDa band was seen in MLE-12 cell lysates (not shown) that represents the intracellular immature unglycosylated or less glycosylated fraction of FXYD5.…”

Section: Resultsmentioning

confidence: 65%

FXYD5 Is an Essential Mediator of the Inflammatory Response during Lung Injury

et al. 2017

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The alveolar epithelium secretes cytokines and chemokines that recruit immune cells to the lungs, which is essential for fighting infections but in excess can promote lung injury. Overexpression of FXYD5, a tissue-specific regulator of the Na,K-ATPase, in mice, impairs the alveolo-epithelial barrier, and FXYD5 overexpression in renal cells increases C-C chemokine ligand-2 (CCL2) secretion in response to lipopolysaccharide (LPS). The aim of this study was to determine whether FXYD5 contributes to the lung inflammation and injury. Exposure of alveolar epithelial cells (AEC) to LPS increased FXYD5 levels at the plasma membrane, and FXYD5 silencing prevented both the activation of NF-κB and the secretion of cytokines in response to LPS. Intratracheal instillation of LPS into mice increased FXYD5 levels in the lung. FXYD5 overexpression increased the recruitment of interstitial macrophages and classical monocytes to the lung in response to LPS. FXYD5 silencing decreased CCL2 levels, number of cells, and protein concentration in bronchoalveolar lavage fluid (BALF) after LPS treatment, indicating that FXYD5 is required for the NF-κB-stimulated epithelial production of CCL2, the influx of immune cells, and the increase in alveolo-epithelial permeability in response to LPS. Silencing of FXYD5 also prevented the activation of NF-κB and cytokine secretion in response to interferon α and TNF-α, suggesting that pro-inflammatory effects of FXYD5 are not limited to the LPS-induced pathway. Furthermore, in the absence of other stimuli, FXYD5 overexpression in AEC activated NF-κB and increased cytokine production, while FXYD5 overexpression in mice increased cytokine levels in BALF, indicating that FXYD5 is sufficient to induce the NF-κB-stimulated cytokine secretion by the alveolar epithelium. The FXYD5 overexpression also increased cell counts in BALF, which was prevented by silencing the CCL2 receptor (CCR2), or by treating mice with a CCR2-blocking antibody, confirming that FXYD5-induced CCL2 production leads to the recruitment of monocytes to the lung. Taken together, the data demonstrate that FXYD5 is a key contributor to inflammatory lung injury.

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The O-glycosylated ectodomain of FXYD5 impairs adhesion by disrupting cell–cell trans-dimerization of Na,K-ATPase β1 subunits

Cited by 23 publications

References 65 publications

Hypercapnia Impairs Na,K-ATPase Function by Inducing Endoplasmic Reticulum Retention of the β-Subunit of the Enzyme in Alveolar Epithelial Cells

Hypercapnia Impairs Na,K-ATPase Function by Inducing Endoplasmic Reticulum Retention of the β-Subunit of the Enzyme in Alveolar Epithelial Cells

The X‐linked juvenile retinoschisis protein retinoschisin is a novel regulator of mitogen‐activated protein kinase signalling and apoptosis in the retina

FXYD5 Is an Essential Mediator of the Inflammatory Response during Lung Injury

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