Ca v1.3 channels and intracellular calcium mediate osmotic stress-induced N-terminal c-Jun kinase activation and disruption of tight junctions in Caco-2 cell monolayers

Geetha Samak, Damodaran Narayanan, Jonathan Jaggar, Radhakrishna Rao

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Abstract

We investigated the role of a Ca 2+ channel and intracellular calcium concentration ([Ca 2+] i) in osmotic stress-induced JNK activation and tight junction disruption in Caco-2 cell monolayers. Osmotic stress-induced tight junction disruption was attenuated by 1,2-bis(2- aminophenoxyl)ethane-N, N, N′, N′-tetraacetic acid (BAPTA)-mediated intracellular Ca 2+ depletion. Depletion of extracellular Ca 2+ at the apical surface, but not basolateral surface, also prevented tight junction disruption. Similarly, thapsigargin-mediated endoplasmic reticulum (ER) Ca 2+ depletion attenuated tight junction disruption. Thapsigargin or extracellular Ca 2+ depletion partially reduced osmotic stress-induced rise in [Ca 2+] i, whereas thapsigargin and extracellular Ca 2+ depletion together resulted in almost complete loss of rise in [Ca 2+] i. L-type Ca 2+ channel blockers (isradipine and diltiazem) or knockdown of the Ca V1.3 channel abrogated [Ca 2+] i rise and disruption of tight junction. Osmotic stress-induced JNK2 activation was abolished by BAPTA and isradipine, and partially reduced by extracellular Ca 2+ depletion, thapsigargin, or Ca V1.3 knockdown. Osmotic stress rapidly induced c-Src activation, which was significantly attenuated by BAPTA, isradipine, or extracellular Ca 2+ depletion. Tight junction disruption by osmotic stress was blocked by tyrosine kinase inhibitors (genistein and PP2) or siRNA-mediated knockdown of c-Src. Osmotic stress induced a robust increase in tyrosine phosphorylation of occludin, which was attenuated by BAPTA, SP600125 (JNK inhibitor), or PP2. These results demonstrate that Ca V1.3 and rise in [Ca 2+] i play a role in the mechanism of osmotic stress-induced tight junction disruption in an intestinal epithelial monolayer. [Ca 2+] i mediate osmotic stress-induced JNK activation and subsequent c-Src activation and tyrosine phosphorylation of tight junction proteins. Additionally, inositol 1,4,5-trisphosphate receptor-mediated release of ER Ca 2+ also contributes to osmotic stress-induced tight junction disruption.

Original languageEnglish (US)
Pages (from-to)30232-30243
Number of pages12
JournalJournal of Biological Chemistry
Volume286
Issue number34
DOIs
StatePublished - Aug 26 2011

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Caco-2 Cells
JNK Mitogen-Activated Protein Kinases
Tight Junctions
Osmotic Pressure
Calcium Channels
Monolayers
Chemical activation
Calcium
Thapsigargin
Isradipine
Phosphorylation
Endoplasmic Reticulum
Tyrosine
Occludin
Tight Junction Proteins
Inositol 1,4,5-Trisphosphate Receptors
Ethane
Diltiazem
Genistein
Protein-Tyrosine Kinases

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Medicine(all)

Cite this

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title = "Ca v1.3 channels and intracellular calcium mediate osmotic stress-induced N-terminal c-Jun kinase activation and disruption of tight junctions in Caco-2 cell monolayers",
abstract = "We investigated the role of a Ca 2+ channel and intracellular calcium concentration ([Ca 2+] i) in osmotic stress-induced JNK activation and tight junction disruption in Caco-2 cell monolayers. Osmotic stress-induced tight junction disruption was attenuated by 1,2-bis(2- aminophenoxyl)ethane-N, N, N′, N′-tetraacetic acid (BAPTA)-mediated intracellular Ca 2+ depletion. Depletion of extracellular Ca 2+ at the apical surface, but not basolateral surface, also prevented tight junction disruption. Similarly, thapsigargin-mediated endoplasmic reticulum (ER) Ca 2+ depletion attenuated tight junction disruption. Thapsigargin or extracellular Ca 2+ depletion partially reduced osmotic stress-induced rise in [Ca 2+] i, whereas thapsigargin and extracellular Ca 2+ depletion together resulted in almost complete loss of rise in [Ca 2+] i. L-type Ca 2+ channel blockers (isradipine and diltiazem) or knockdown of the Ca V1.3 channel abrogated [Ca 2+] i rise and disruption of tight junction. Osmotic stress-induced JNK2 activation was abolished by BAPTA and isradipine, and partially reduced by extracellular Ca 2+ depletion, thapsigargin, or Ca V1.3 knockdown. Osmotic stress rapidly induced c-Src activation, which was significantly attenuated by BAPTA, isradipine, or extracellular Ca 2+ depletion. Tight junction disruption by osmotic stress was blocked by tyrosine kinase inhibitors (genistein and PP2) or siRNA-mediated knockdown of c-Src. Osmotic stress induced a robust increase in tyrosine phosphorylation of occludin, which was attenuated by BAPTA, SP600125 (JNK inhibitor), or PP2. These results demonstrate that Ca V1.3 and rise in [Ca 2+] i play a role in the mechanism of osmotic stress-induced tight junction disruption in an intestinal epithelial monolayer. [Ca 2+] i mediate osmotic stress-induced JNK activation and subsequent c-Src activation and tyrosine phosphorylation of tight junction proteins. Additionally, inositol 1,4,5-trisphosphate receptor-mediated release of ER Ca 2+ also contributes to osmotic stress-induced tight junction disruption.",
author = "Geetha Samak and Damodaran Narayanan and Jonathan Jaggar and Radhakrishna Rao",
year = "2011",
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language = "English (US)",
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journal = "Journal of Biological Chemistry",
issn = "0021-9258",
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T1 - Ca v1.3 channels and intracellular calcium mediate osmotic stress-induced N-terminal c-Jun kinase activation and disruption of tight junctions in Caco-2 cell monolayers

AU - Samak, Geetha

AU - Narayanan, Damodaran

AU - Jaggar, Jonathan

AU - Rao, Radhakrishna

PY - 2011/8/26

Y1 - 2011/8/26

N2 - We investigated the role of a Ca 2+ channel and intracellular calcium concentration ([Ca 2+] i) in osmotic stress-induced JNK activation and tight junction disruption in Caco-2 cell monolayers. Osmotic stress-induced tight junction disruption was attenuated by 1,2-bis(2- aminophenoxyl)ethane-N, N, N′, N′-tetraacetic acid (BAPTA)-mediated intracellular Ca 2+ depletion. Depletion of extracellular Ca 2+ at the apical surface, but not basolateral surface, also prevented tight junction disruption. Similarly, thapsigargin-mediated endoplasmic reticulum (ER) Ca 2+ depletion attenuated tight junction disruption. Thapsigargin or extracellular Ca 2+ depletion partially reduced osmotic stress-induced rise in [Ca 2+] i, whereas thapsigargin and extracellular Ca 2+ depletion together resulted in almost complete loss of rise in [Ca 2+] i. L-type Ca 2+ channel blockers (isradipine and diltiazem) or knockdown of the Ca V1.3 channel abrogated [Ca 2+] i rise and disruption of tight junction. Osmotic stress-induced JNK2 activation was abolished by BAPTA and isradipine, and partially reduced by extracellular Ca 2+ depletion, thapsigargin, or Ca V1.3 knockdown. Osmotic stress rapidly induced c-Src activation, which was significantly attenuated by BAPTA, isradipine, or extracellular Ca 2+ depletion. Tight junction disruption by osmotic stress was blocked by tyrosine kinase inhibitors (genistein and PP2) or siRNA-mediated knockdown of c-Src. Osmotic stress induced a robust increase in tyrosine phosphorylation of occludin, which was attenuated by BAPTA, SP600125 (JNK inhibitor), or PP2. These results demonstrate that Ca V1.3 and rise in [Ca 2+] i play a role in the mechanism of osmotic stress-induced tight junction disruption in an intestinal epithelial monolayer. [Ca 2+] i mediate osmotic stress-induced JNK activation and subsequent c-Src activation and tyrosine phosphorylation of tight junction proteins. Additionally, inositol 1,4,5-trisphosphate receptor-mediated release of ER Ca 2+ also contributes to osmotic stress-induced tight junction disruption.

AB - We investigated the role of a Ca 2+ channel and intracellular calcium concentration ([Ca 2+] i) in osmotic stress-induced JNK activation and tight junction disruption in Caco-2 cell monolayers. Osmotic stress-induced tight junction disruption was attenuated by 1,2-bis(2- aminophenoxyl)ethane-N, N, N′, N′-tetraacetic acid (BAPTA)-mediated intracellular Ca 2+ depletion. Depletion of extracellular Ca 2+ at the apical surface, but not basolateral surface, also prevented tight junction disruption. Similarly, thapsigargin-mediated endoplasmic reticulum (ER) Ca 2+ depletion attenuated tight junction disruption. Thapsigargin or extracellular Ca 2+ depletion partially reduced osmotic stress-induced rise in [Ca 2+] i, whereas thapsigargin and extracellular Ca 2+ depletion together resulted in almost complete loss of rise in [Ca 2+] i. L-type Ca 2+ channel blockers (isradipine and diltiazem) or knockdown of the Ca V1.3 channel abrogated [Ca 2+] i rise and disruption of tight junction. Osmotic stress-induced JNK2 activation was abolished by BAPTA and isradipine, and partially reduced by extracellular Ca 2+ depletion, thapsigargin, or Ca V1.3 knockdown. Osmotic stress rapidly induced c-Src activation, which was significantly attenuated by BAPTA, isradipine, or extracellular Ca 2+ depletion. Tight junction disruption by osmotic stress was blocked by tyrosine kinase inhibitors (genistein and PP2) or siRNA-mediated knockdown of c-Src. Osmotic stress induced a robust increase in tyrosine phosphorylation of occludin, which was attenuated by BAPTA, SP600125 (JNK inhibitor), or PP2. These results demonstrate that Ca V1.3 and rise in [Ca 2+] i play a role in the mechanism of osmotic stress-induced tight junction disruption in an intestinal epithelial monolayer. [Ca 2+] i mediate osmotic stress-induced JNK activation and subsequent c-Src activation and tyrosine phosphorylation of tight junction proteins. Additionally, inositol 1,4,5-trisphosphate receptor-mediated release of ER Ca 2+ also contributes to osmotic stress-induced tight junction disruption.

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U2 - 10.1074/jbc.M111.240358

DO - 10.1074/jbc.M111.240358

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