TY - JOUR
T1 - Intracellular endothelin type B receptor-driven Ca2+ signal elicits nitric oxide production in endothelial cells
AU - Deliu, Elena
AU - Brailoiu, G. Cristina
AU - Mallilankaraman, Karthik
AU - Wang, Hong
AU - Madesh, Muniswamy
AU - Undieh, Ashiwel S.
AU - Koch, Walter J.
AU - Brailoiu, Eugen
PY - 2012/11/30
Y1 - 2012/11/30
N2 - Endothelin-1 exerts its actions via activation of ETA and ETBGq/11 protein-coupled receptors, located in the plasmalemma, cytoplasm, and nucleus. Although the autocrine/paracrine nature of endothelin-1 signaling has been extensively studied, its intracrine role has been largely attributed to interaction with receptors located on nuclear membranes and the nucleoplasm. Because ETB receptors have been shown to be targeted to endolysosomes, we used intracellular microinjection and concurrent imaging methods to test their involvement in Ca2+ signaling and subsequential NO production. We provide evidence that microinjected endothelin-1 produces a dose-dependent elevation in cytosolic calcium concentration in ETB-transfected cells and endothelial cells; this response is sensitive to ETB but not ETA receptor blockade. In endothelial cells, the endothelin-1-induced Ca2+ response is abolished upon endolysosomal but not Golgi disruption. Moreover, the effect is prevented by inhibition of microautophagy and is sensitive to inhibitors of the phospholipase C and inositol 1,4,5-trisphosphate receptor. Furthermore, intracellular endothelin-1 increases nitric oxide via an ET B-dependent mechanism. Our results indicate for the first time that intracellular endothelin-1 activates endolysosomal ETB receptors and increase cytosolic Ca2+ and nitric oxide production. Endothelin-1 acts in an intracrine fashion on endolysosomal ETB to induce nitric oxide formation, thus modulating endothelial function.
AB - Endothelin-1 exerts its actions via activation of ETA and ETBGq/11 protein-coupled receptors, located in the plasmalemma, cytoplasm, and nucleus. Although the autocrine/paracrine nature of endothelin-1 signaling has been extensively studied, its intracrine role has been largely attributed to interaction with receptors located on nuclear membranes and the nucleoplasm. Because ETB receptors have been shown to be targeted to endolysosomes, we used intracellular microinjection and concurrent imaging methods to test their involvement in Ca2+ signaling and subsequential NO production. We provide evidence that microinjected endothelin-1 produces a dose-dependent elevation in cytosolic calcium concentration in ETB-transfected cells and endothelial cells; this response is sensitive to ETB but not ETA receptor blockade. In endothelial cells, the endothelin-1-induced Ca2+ response is abolished upon endolysosomal but not Golgi disruption. Moreover, the effect is prevented by inhibition of microautophagy and is sensitive to inhibitors of the phospholipase C and inositol 1,4,5-trisphosphate receptor. Furthermore, intracellular endothelin-1 increases nitric oxide via an ET B-dependent mechanism. Our results indicate for the first time that intracellular endothelin-1 activates endolysosomal ETB receptors and increase cytosolic Ca2+ and nitric oxide production. Endothelin-1 acts in an intracrine fashion on endolysosomal ETB to induce nitric oxide formation, thus modulating endothelial function.
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U2 - 10.1074/jbc.M112.418533
DO - 10.1074/jbc.M112.418533
M3 - Article
C2 - 23086942
AN - SCOPUS:84870324879
SN - 0021-9258
VL - 287
SP - 41023
EP - 41031
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 49
ER -