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The Na+, K+-ATPase ?1 subunit regulates epithelial tight junctions via MRCK?.

ABSTRACT: An intact lung epithelial barrier is essential for lung homeostasis. The Na+, K+-ATPase (NKA), primarily serving as an ion transporter, also regulates epithelial barrier function via modulation of tight junctions. However, the underlying mechanism is not well understood. Here, we show that overexpression of the NKA ?1 subunit upregulates the expression of tight junction proteins, leading to increased alveolar epithelial barrier function by an ion transport-independent mechanism. Using IP and mass spectrometry, we identified a number of unknown protein interactions of the ?1 subunit, including a top candidate, myotonic dystrophy kinase-related cdc42-binding kinase ? (MRCK?), which is a protein kinase known to regulate peripheral actin formation. Using a doxycycline-inducible gene expression system, we demonstrated that MRCK? and its downstream activation of myosin light chain is required for the regulation of alveolar barrier function by the NKA ?1 subunit. Importantly, MRCK? is expressed in both human airways and alveoli and has reduced expression in patients with acute respiratory distress syndrome (ARDS), a lung illness that can be caused by multiple direct and indirect insults, including the infection of influenza virus and SARS-CoV-2. Our results have elucidated a potentially novel mechanism by which NKA regulates epithelial tight junctions and have identified potential drug targets for treating ARDS and other pulmonary diseases that are caused by barrier dysfunction.

SUBMITTER: Bai H

PROVIDER: S-EPMC7934944 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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The Na+, K+-ATPase β1 subunit regulates epithelial tight junctions via MRCKα.

Bai Haiqing H Zhou Rui R Barravecchia Michael M Norman Rosemary R Friedman Alan A Yu Deborah D Lin Xin X Young Jennifer L JL Dean David A DA

JCI insight 20210222 4

An intact lung epithelial barrier is essential for lung homeostasis. The Na+, K+-ATPase (NKA), primarily serving as an ion transporter, also regulates epithelial barrier function via modulation of tight junctions. However, the underlying mechanism is not well understood. Here, we show that overexpression of the NKA β1 subunit upregulates the expression of tight junction proteins, leading to increased alveolar epithelial barrier function by an ion transport-independent mechanism. Using IP and mas ...[more]

PMID: 33507884

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Project description:BackgroundBenign prostatic hyperplasia (BPH) is arguably the most common disease in aging men. Although the etiology is not well understood, chronic prostatic inflammation is thought to play an important role in BPH initiation and progression. Our recent studies suggest that the prostatic epithelial barrier is compromised in glandular BPH tissues. The proinflammatory cytokine transforming growth factor beta 1 (TGF-β1) impacts tight junction formation, enhances epithelial barrier permeability, and suppresses claudin-1 messenger RNA expression in prostatic epithelial cells. However, the role of claudin-1 in the prostatic epithelial barrier and its regulation by TGF-β1 in prostatic epithelial cells are not clear.MethodsThe expression of claudin-1 was analyzed in 22 clinical BPH specimens by immunohistochemistry. Human benign prostate epithelial cell lines BPH-1 and BHPrE1 were treated with TGF-β1 and transfected with small interfering RNAs specific to claudin-1. Epithelial monolayer permeability changes in the treated cells were measured using trans-epithelial electrical resistance (TEER). The expression of claudin-1, E-cadherin, N-cadherin, snail, slug, and activation of mitogen-activated proteins kinases (MAPKs) and AKT was assessed following TGF-β1 treatment using Western blot analysis.ResultsClaudin-1 expression was decreased in glandular BPH tissue compared with adjacent normal prostatic tissue in patient specimens. TGF-β1 treatment or claudin-1 knockdown in prostatic epithelial cell lines increased monolayer permeability. TGF-β1 decreased levels of claudin-1 and increased levels of snail and slug as well as increased phosphorylation of the MAPK extracellular signal-regulated kinase-1/2 (ERK-1/2) in both BPH-1 and BHPrE1 cells. Overexpression of snail or slug had no effect on claudin-1 expression. In contrast, PD98059 and U0126, inhibitors of the upstream activator of ERK-1/2 (ie, MEK-1/2) restored claudin-1 expression level as well as the epithelial barrier.ConclusionOur findings suggest that downregulation of claudin-1 by TGF-β1 acting through the noncanonical MEK-1/2/ERK-1/2 pathway triggers increased prostatic epithelial monolayer permeability in vitro. These findings also suggest that elevated TGF-β1 may contribute to claudin-1 downregulation and compromised epithelial barrier in clinical BPH specimens.

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The Na+, K+-ATPase ?1 subunit regulates epithelial tight junctions via MRCK?.

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The Na+, K+-ATPase β1 subunit regulates epithelial tight junctions via MRCKα.

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