Abstract:The transient receptor potential V4 channel (TRPV4) is responsive to a variety of physical and chemical stimuli, including a synthetic agonist GSK1016790A (GSK). Here, we show that TRPV4 is functionally expressed in, and that GSK induces the reversible opening of tight junctions (TJs) in epithelial Madin–Darby canine kidney II monolayers. Stimulation of TRPV4 by GSK induces an increase in fluorescein isothiocyanate‐conjugated dextran (4 kDa) permeability and a reduction in transepithelial resistance, and these… Show more
“…1 A ) and a doublet in MDCK cells and choroid plexus tissue ( Fig. 1 , B and C ) consistent with previous studies ( 39 , 44 , 45 , 46 , 47 ). Figure 1 TRPV4 is monoubiquitinated/multiubiquitinated.…”
Section: Resultssupporting
confidence: 91%
“…Our data showed a single TRPV4 band in 50B11 cells (Fig 1A) and a doublet in MDCK cells and choroid plexus tissue (Fig. 1B, C) consistent with previous studies (39,(44)(45)(46)(47).…”
Section: Trpv4 Is Mono-/multi-ubiquitinated In Multiple Cell Typessupporting
confidence: 92%
“…To determine whether endogenous TRPV4 protein is ubiquitinated in cells and whole tissue, we assessed an immortalized rat sensory neuron cell line (50B11 cells), the Madin-Darby canine kidney (MDCK) epithelial cell line, and mouse choroid plexus tissue; all of which express robust levels of TRPV4 (32,38,39). To immunopurify endogenous TRPV4, we used an anti-TRPV4 antibody that detects TRPV4 protein in cultured primary choroid plexus epithelial cells (40), kidney, and choroid plexus derived from wild type (WT), but not Trpv4 null mice (40)(41)(42) (Supplementary Fig 1A-B).…”
Section: Trpv4 Is Mono-/multi-ubiquitinated In Multiple Cell Typesmentioning
“…1 A ) and a doublet in MDCK cells and choroid plexus tissue ( Fig. 1 , B and C ) consistent with previous studies ( 39 , 44 , 45 , 46 , 47 ). Figure 1 TRPV4 is monoubiquitinated/multiubiquitinated.…”
Section: Resultssupporting
confidence: 91%
“…Our data showed a single TRPV4 band in 50B11 cells (Fig 1A) and a doublet in MDCK cells and choroid plexus tissue (Fig. 1B, C) consistent with previous studies (39,(44)(45)(46)(47).…”
Section: Trpv4 Is Mono-/multi-ubiquitinated In Multiple Cell Typessupporting
confidence: 92%
“…To determine whether endogenous TRPV4 protein is ubiquitinated in cells and whole tissue, we assessed an immortalized rat sensory neuron cell line (50B11 cells), the Madin-Darby canine kidney (MDCK) epithelial cell line, and mouse choroid plexus tissue; all of which express robust levels of TRPV4 (32,38,39). To immunopurify endogenous TRPV4, we used an anti-TRPV4 antibody that detects TRPV4 protein in cultured primary choroid plexus epithelial cells (40), kidney, and choroid plexus derived from wild type (WT), but not Trpv4 null mice (40)(41)(42) (Supplementary Fig 1A-B).…”
Section: Trpv4 Is Mono-/multi-ubiquitinated In Multiple Cell Typesmentioning
“…Based on this background, we have now tested the influence of ouabain on the expression of transient receptor potential vanilloid 4 (TRPV4), a type of ion channel which has been shown to be endogenously expressed in MDCK epithelial cells [19,20]. TRPV4 channels are expressed by various cells across the body, including the nervous system, skin, and vasculature.…”
Ouabain, a substance originally obtained from plants, is now classified as a hormone because it is produced endogenously in certain animals, including humans. However, its precise effects on the body remain largely unknown. Previous studies have shown that ouabain can influence the phenotype of epithelial cells by affecting the expression of cell–cell molecular components and voltage-gated potassium channels. In this study, we conducted whole-cell clamp assays to determine whether ouabain affects the activity and/or expression of TRPV4 channels. Our findings indicate that ouabain has a statistically significant effect on the density of TRPV4 currents (dITRPV4), with an EC50 of 1.89 nM. Regarding treatment duration, dITRPV4 reaches its peak at around 1 h, followed by a subsequent decline and then a resurgence after 6 h, suggesting a short-term modulatory effect related to on TRPV4 channel activity and a long-term effect related to the promotion of synthesis of new TRPV4 channel units. The enhancement of dITRPV4 induced by ouabain was significantly lower in cells seeded at low density than in cells in a confluent monolayer, indicating that the action of ouabain depends on intercellular contacts. Furthermore, the fact that U73122 and neomycin suppress the effect caused by ouabain in the short term suggests that the short-term induced enhancement of dITRPV4 is due to the depletion of PIP2 stores. In contrast, the fact that the long-term effect is inhibited by PP2, wortmannin, PD, FR18, and IKK16 suggests that cSrc, PI3K, Erk1/2, and NF-kB are among the components included in the signaling pathways.
“…Additionally, it plays a central role in cell volume regulation by acting as a membrane-stretch-sensing channel upon osmotic changes 26,27 . Besides these physiological functions, TRPV4 seems to be important for the maintenance of epithelial integrity through its role in the regulation of on adherens and tight junction proteins [28][29][30][31][32][33] . Further, TRPV4-mediated calcium influx seems to be essential for the integrity of cell-cell junctions in the skin keratinocytes 34,35 .…”
Biophysical cues from the cell microenvironment are detected by mechanosensitive components at the cell surface. Such machineries convert physical information into biochemical signaling cascades within cells, subsequently leading to various cellular responses in a stimulus-dependent manner. At the surface of extracellular environment and cell cytoplasm exist several ion channel families that are activated by mechanical signals to direct intracellular events. One of such channel is formed by transient receptor potential cation channel subfamily V member, TRPV4 that is known to act as a mechanosensor in wide variaty of tissues and control ion-influx in a spatio-temporal way. Here we report that TRPV4 is prominently expressed in the stem/progenitor cell populations of the mammary epithelium and seems important for the lineage-specific differentiation, consequently affecting mechanical features of the mature mammary epithelium. This was evident by the lack of several markers for mature myoepithelial and luminal epithelial cells in TRPV4-depleted cell lines. Interestingly, TRPV4 expression is controlled in a tension-dependent manner and it also impacts differentation process dependently on the stiffness of the microenvironment. Furthermore, such cells in a 3D compartment were disabled to maintain normal mammosphere structures and displayed abnormal lumen formation, size of the structures and disrupted cellular junctions. Mechanosensitive TRPV4 channel therefore act as critical player in the homeostasis of normal mammary epithelium through sensing the physical environment and guiding accordingly differentiation and structural organization of the bilayered mammary epithelium.
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