STIM1-Regulated Ca2+ Influx across the Apical and the Basolateral Membrane in Colonic Epithelium

STIM1-Regulated Ca2+ Influx across the Apical and the Basolateral Membrane in Colonic Epithelium In nonexcitable cells, store-operated Ca2+ entry is the most important pathway for influx of extracellular Ca2+ serving as a second messenger in the cytoplasm. The present study investigated the expression, localization and polar distribution of two key components of store-operated Ca2+ entry identified, e.g., in lymphocytes or epithelial cell lines—STIM1 (stromal interacting molecule 1), working as a Ca2+ sensor in the endoplasmic reticulum, and Orai1, working as the (or part of the) store-operated Ca2+ channel in the plasma membrane—in a native intestinal epithelium, i.e., rat colon. Immunohistochemical investigations revealed expression of STIM1 and Orai1 in the rat colonic epithelium. Ca2+ store depletion led to a translocation of STIM1 both to the basolateral as well as to the apical cell pole as observed by confocal microscopy. A Ca2+ depletion/repletion protocol was used in Ussing chamber experiments to investigate the contribution of basolateral and apical store-operated Ca2+ entry to the induction of anion secretion. These experiments revealed that Ca2+-dependent anion secretion was induced not only by basolateral Ca2+ repletion but also, to a lesser extent, by apical Ca2+ repletion. Both responses were suppressed by La3+. The effect of basolateral Ca2+ repletion was significantly inhibited by brefeldin A, a blocker of vesicular transport from the endoplasmic reticulum to the Golgi apparatus. In a final series of experiments, fura-2-loaded HT29/B6 cells were used. A carbachol-induced increase in the cytosolic Ca2+ concentration was significantly reduced when cells were pretreated with siRNA against STIM1. In conclusion, these results demonstrate that STIM1 as a key component of intracellular Ca2+ signaling is expressed by rat colonic epithelium and is involved in the regulation not only of basolateral but also of apical Ca2+ influx. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png The Journal of Membrane Biology Springer Journals

STIM1-Regulated Ca2+ Influx across the Apical and the Basolateral Membrane in Colonic Epithelium

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Publisher
Springer-Verlag
Copyright
Copyright © 2013 by Springer Science+Business Media New York
Subject
Life Sciences; Biochemistry, general; Human Physiology
ISSN
0022-2631
eISSN
1432-1424
D.O.I.
10.1007/s00232-013-9528-9
Publisher site
See Article on Publisher Site

Abstract

In nonexcitable cells, store-operated Ca2+ entry is the most important pathway for influx of extracellular Ca2+ serving as a second messenger in the cytoplasm. The present study investigated the expression, localization and polar distribution of two key components of store-operated Ca2+ entry identified, e.g., in lymphocytes or epithelial cell lines—STIM1 (stromal interacting molecule 1), working as a Ca2+ sensor in the endoplasmic reticulum, and Orai1, working as the (or part of the) store-operated Ca2+ channel in the plasma membrane—in a native intestinal epithelium, i.e., rat colon. Immunohistochemical investigations revealed expression of STIM1 and Orai1 in the rat colonic epithelium. Ca2+ store depletion led to a translocation of STIM1 both to the basolateral as well as to the apical cell pole as observed by confocal microscopy. A Ca2+ depletion/repletion protocol was used in Ussing chamber experiments to investigate the contribution of basolateral and apical store-operated Ca2+ entry to the induction of anion secretion. These experiments revealed that Ca2+-dependent anion secretion was induced not only by basolateral Ca2+ repletion but also, to a lesser extent, by apical Ca2+ repletion. Both responses were suppressed by La3+. The effect of basolateral Ca2+ repletion was significantly inhibited by brefeldin A, a blocker of vesicular transport from the endoplasmic reticulum to the Golgi apparatus. In a final series of experiments, fura-2-loaded HT29/B6 cells were used. A carbachol-induced increase in the cytosolic Ca2+ concentration was significantly reduced when cells were pretreated with siRNA against STIM1. In conclusion, these results demonstrate that STIM1 as a key component of intracellular Ca2+ signaling is expressed by rat colonic epithelium and is involved in the regulation not only of basolateral but also of apical Ca2+ influx.

Journal

The Journal of Membrane BiologySpringer Journals

Published: Feb 9, 2013

References

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