TRPC1 functions as a store-operated Ca2+ channel in intestinal epithelial cells and regulates early mucosal restitution after wounding

2006 ◽  
Vol 290 (4) ◽  
pp. G782-G792 ◽  
Author(s):  
Jaladanki N. Rao ◽  
Oleksandr Platoshyn ◽  
Vera A. Golovina ◽  
Lan Liu ◽  
Tongtong Zou ◽  
...  
Keyword(s):  
Cell Migration ◽  
Epithelial Cells ◽  
Transient Receptor Potential ◽  
Intestinal Epithelial Cells ◽  
Critical Role ◽  
Receptor Potential ◽  
Mucosal Injury ◽  
Intestinal Epithelial ◽  
Epithelial Restitution ◽  
Transient Receptor

An increase in cytosolic free Ca2+ concentration ([Ca2+]cyt) results from Ca2+ release from intracellular stores and extracellular Ca2+ influx through Ca2+-permeable ion channels and is crucial for initiating intestinal epithelial restitution to reseal superficial wounds after mucosal injury. Capacitative Ca2+ entry (CCE) induced by Ca2+ store depletion represents a major Ca2+ influx mechanism, but the exact molecular components constituting this process remain elusive. This study determined whether canonical transient receptor potential (TRPC)1 served as a candidate protein for Ca2+-permeable channels mediating CCE in intestinal epithelial cells and played an important role in early epithelial restitution. Normal intestinal epithelial cells (the IEC-6 cell line) expressed TRPC1 and TPRC5 and displayed typical records of whole cell store-operated Ca2+ currents and CCE generated by Ca2+ influx after depletion of intracellular stores. Induced TRPC1 expression by stable transfection with the TRPC1 gene increased CCE and enhanced cell migration during restitution. Differentiated IEC-Cdx2L1 cells induced by forced expression of the Cdx2 gene highly expressed endogenous TRPC1 and TRPC5 and exhibited increased CCE and cell migration. Inhibition of TRPC1 expression by small interfering RNA specially targeting TRPC1 not only reduced CCE but also inhibited cell migration after wounding. These findings strongly suggest that TRPC1 functions as store-operated Ca2+ channels and plays a critical role in intestinal epithelial restitution by regulating CCE and intracellular [Ca2+]cyt.

scholarly journals RhoA enhances store-operated Ca2+ entry and intestinal epithelial restitution by interacting with TRPC1 after wounding

2015 ◽  
Vol 309 (9) ◽  
pp. G759-G767 ◽  
Author(s):  
Hee Kyoung Chung ◽  
Navneeta Rathor ◽  
Shelley R. Wang ◽  
Jian-Ying Wang ◽  
Jaladanki N. Rao
Keyword(s):  
Cell Migration ◽  
Transient Receptor Potential ◽  
Ectopic Expression ◽  
Receptor Potential ◽  
Dominant Negative ◽  
Intestinal Epithelial ◽  
Epithelial Restitution ◽  
Epithelial Cell Migration ◽  
Transient Receptor ◽  
Wounded Area

Early mucosal restitution occurs as a consequence of epithelial cell migration to resealing of superficial wounds after injury. Our previous studies show that canonical transient receptor potential-1 (TRPC1) functions as a store-operated Ca2+ channel (SOC) in intestinal epithelial cells (IECs) and plays an important role in early epithelial restitution by increasing Ca2+ influx. Here we further reported that RhoA, a small GTP-binding protein, interacts with and regulates TRPC1, thus enhancing SOC-mediated Ca2+ entry (SOCE) and epithelial restitution after wounding. RhoA physically associated with TRPC1 and formed the RhoA/TRPC1 complexes, and this interaction increased in stable TRPC1-transfected IEC-6 cells (IEC-TRPC1). Inactivation of RhoA by treating IEC-TRPC1 cells with exoenzyme C3 transferase (C3) or ectopic expression of dominant negative RhoA (DNMRhoA) reduced RhoA/TRPC1 complexes and inhibited Ca2+ influx after store depletion, which was paralleled by an inhibition of cell migration over the wounded area. In contrast, ectopic expression of wild-type (WT)-RhoA increased the levels of RhoA/TRPC1 complexes, induced Ca2+ influx through activation of SOCE, and promoted cell migration after wounding. TRPC1 silencing by transfecting stable WT RhoA-transfected cells with siRNA targeting TRPC1 (siTRPC1) reduced SOCE and repressed epithelial restitution. Moreover, ectopic overexpression of WT-RhoA in polyamine-deficient cells rescued the inhibition of Ca2+ influx and cell migration induced by polyamine depletion. These findings indicate that RhoA interacts with and activates TRPC1 and thus stimulates rapid epithelial restitution after injury by inducing Ca2+ signaling.

scholarly journals Polyamines regulate intestinal epithelial restitution through TRPC1-mediated Ca2+ signaling by differentially modulating STIM1 and STIM2

2012 ◽  
Vol 303 (3) ◽  
pp. C308-C317 ◽  
Author(s):  
Jaladanki N. Rao ◽  
Navneeta Rathor ◽  
Ran Zhuang ◽  
Tongtong Zou ◽  
Lan Liu ◽  
...  
Keyword(s):  
Cell Migration ◽  
Intestinal Epithelial Cell ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Intestinal Epithelial ◽  
Canonical Transient Receptor Potential ◽  
Pathological Conditions ◽  
Epithelial Restitution ◽  
Transient Receptor ◽  
Stimulation Of

Early epithelial restitution occurs as a consequence of intestinal epithelial cell (IEC) migration after wounding, and its defective regulation is implicated in various critical pathological conditions. Polyamines stimulate intestinal epithelial restitution, but their exact mechanism remains unclear. Canonical transient receptor potential-1 (TRPC1)-mediated Ca2+ signaling is crucial for stimulation of IEC migration after wounding, and induced translocation of stromal interaction molecule 1 (STIM1) to the plasma membrane activates TRPC1-mediated Ca2+ influx and thus enhanced restitution. Here, we show that polyamines regulate intestinal epithelial restitution through TRPC1-mediated Ca2+ signaling by altering the ratio of STIM1 to STIM2. Increasing cellular polyamines by ectopic overexpression of the ornithine decarboxylase (ODC) gene stimulated STIM1 but inhibited STIM2 expression, whereas depletion of cellular polyamines by inhibiting ODC activity decreased STIM1 but increased STIM2 levels. Induced STIM1/TRPC1 association by increasing polyamines enhanced Ca2+ influx and stimulated epithelial restitution, while decreased formation of the STIM1/TRPC1 complex by polyamine depletion decreased Ca2+ influx and repressed cell migration. Induced STIM1/STIM2 heteromers by polyamine depletion or STIM2 overexpression suppressed STIM1 membrane translocation and inhibited Ca2+ influx and epithelial restitution. These results indicate that polyamines differentially modulate cellular STIM1 and STIM2 levels in IECs, in turn controlling TRPC1-mediated Ca2+ signaling and influencing cell migration after wounding.

scholarly journals Induced TRPC1 expression sensitizes intestinal epithelial cells to apoptosis by inhibiting NF-κB activation through Ca2+ influx

2006 ◽  
Vol 397 (1) ◽  
pp. 77-87 ◽  
Author(s):  
Bernard S. Marasa ◽  
Jaladanki N. Rao ◽  
Tongtong Zou ◽  
Lan Liu ◽  
Kaspar M. Keledjian ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Transient Receptor Potential ◽  
Intestinal Epithelial Cells ◽  
Critical Role ◽  
Ectopic Expression ◽  
Receptor Potential ◽  
Nuclear Factor Κb ◽  
Intestinal Epithelial ◽  
Store Depletion ◽  
Increased Susceptibility

Apoptosis occurs within crypts and at the intestinal luminal surface and plays a critical role in mucosal homoeostasis. NF-κB (nuclear factor-κB) is the central regulator of the transcription of genes involved in apoptosis, and its activity is highly regulated in the intestinal mucosa. We have recently demonstrated that TRPC1 (transient receptor potential canonical-1) is expressed in IECs (intestinal epithelial cells) and functions as a Ca2+ permeable channel activated by Ca2+ store depletion. The present study tests the hypothesis that TRPC1 channels are implicated in the regulation of apoptosis by inhibiting NF-κB through the induction of TRPC1-mediated Ca2+ influx in the IEC-6 line. The expression of TRPC1 induced by stable transfection of IEC-6 cells with the wild-type TRPC1 gene (IEC-TRPC1 cells) increased Ca2+ influx after Ca2+ store depletion and repressed NF-κB transactivation, which was associated with an increase in susceptibility to apoptosis induced by exposure to TNFα (tumour necrosis factor-α) plus CHX (cycloheximide) (TNF-α/CHX), or STS (staurosporine). By contrast, the induction of endogenous NF-κB activity, by the depletion of cellular polyamines, promoted resistance to apoptosis, which was prevented by the ectopic expression of the IκBα super-repressor. Furthermore, inhibition of TRPC1 expression by transfection with siRNA (small interfering RNA) targeting TRPC1 (siTRPC1) decreased Ca2+ influx, increased NF-κB transactivation, and prevented the increased susceptibility of IEC-TRPC1 cells to apoptosis. Decreasing Ca2+ influx by exposure to a Ca2+-free medium also induced NF-κB activity and blocked the increased susceptibility to apoptosis of stable IEC-TRPC1 cells. These results indicate that induced TRPC1 expression sensitizes IECs to apoptosis by inhibiting NF-κB activity as a result of the stimulation of Ca2+ influx.

scholarly journals STIM1 translocation to the plasma membrane enhances intestinal epithelial restitution by inducing TRPC1-mediated Ca2+ signaling after wounding

2010 ◽  
Vol 299 (3) ◽  
pp. C579-C588 ◽  
Author(s):  
Jaladanki N. Rao ◽  
Navneeta Rathor ◽  
Tongtong Zou ◽  
Lan Liu ◽  
Lan Xiao ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Migration ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Mucosal Injury ◽  
Intestinal Epithelial ◽  
Epithelial Restitution ◽  
Essential Components ◽  
Ef Hand ◽  
Trpc1 Channels

Early epithelial restitution is an important repair modality in the gut mucosa and occurs as a consequence of epithelial cell migration. Canonical transient receptor potential-1 (TRPC1) functions as a store-operated Ca2+ channel (SOCs) in intestinal epithelial cells (IECs) and regulates intestinal restitution, but the exact upstream signals initiating TRPC1 activation after mucosal injury remain elusive. Stromal interaction molecule 1 (STIM1) is a single membrane-spanning protein and is recently identified as essential components of SOC activation. The current study was performed to determine whether STIM1 plays a role in the regulation of intestinal epithelial restitution by activating TRPC1 channels. STIM1 translocation to the plasma membrane increased after wounding, which was followed by an increase in IEC migration to reseal wounds. Increased STIM1 levels at the plasma membrane by overexpressing EF-hand mutant STIM1 enhanced Ca2+ influx through SOCs and stimulated IEC migration after wounding. STIM1 interacted with TRPC1 and formed STIM1/TRPC1 complex, whereas inactivation of STIM1 by STIM1 silencing decreased SOC-mediated Ca2+ influx and inhibited epithelial restitution. In cells overexpressing EF-hand mutant STIM1, TRPC1 silencing also decreased STIM1/TRPC1 complex, reduced SOC-mediated Ca2+ influx, and repressed cell migration after wounding. Our findings demonstrate that induced STIM1 translocation to the plasma membrane promotes IEC migration after wounding by enhancing TRPC1-mediated Ca2+ signaling and provide new insight into the mechanism of intestinal epithelial restitution.

Polyamines regulate β-catenin tyrosine phosphorylation via Ca2+ during intestinal epithelial cell migration

2002 ◽  
Vol 283 (3) ◽  
pp. C722-C734 ◽  
Author(s):  
Xin Guo ◽  
Jaladanki N. Rao ◽  
Lan Liu ◽  
Mort Rizvi ◽  
Douglas J. Turner ◽  
...  
Keyword(s):  
Cell Migration ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Tyrosine Phosphorylation ◽  
Kinase Inhibitor ◽  
Intestinal Epithelial Cells ◽  
Critical Role ◽  
Cytoskeletal Proteins ◽  
Intestinal Epithelial ◽  
Epithelial Cell Migration

Polyamines are essential for early mucosal restitution that occurs by epithelial cell migration to reseal superficial wounds after injury. Normal intestinal epithelial cells are tightly bound in sheets, but they need to be rapidly disassembled during restitution. β-Catenin is involved in cell-cell adhesion, and its tyrosine phosphorylation causes disassembly of adhesion junctions, enhancing the spreading of cells. The current study determined whether polyamines are required for the stimulation of epithelial cell migration by altering β-catenin tyrosine phosphorylation. Migration of intestinal epithelial cells (IEC-6 line) after wounding was associated with an increase in β-catenin tyrosine phosphorylation, which decreased the binding activity of β-catenin to α-catenin. Polyamine depletion by α-difluoromethylornithine reduced cytoplasmic free Ca2+concentration ([Ca2+]cyt), prevented induction of β-catenin phosphorylation, and decreased cell migration. Elevation of [Ca2+]cyt induced by the Ca2+ ionophore ionomycin restored β-catenin phosphorylation and promoted migration in polyamine-deficient cells. Decreased β-catenin phosphorylation through the tyrosine kinase inhibitor herbimycin-A or genistein blocked cell migration, which was accompanied by reorganization of cytoskeletal proteins. These results indicate that β-catenin tyrosine phosphorylation plays a critical role in polyamine-dependent cell migration and that polyamines induce β-catenin tyrosine phosphorylation at least partially through [Ca2+]cyt.

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