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Tadevosyan A.,University of Montréal | Tadevosyan A.,Montreal Heart Institute | Villeneuve L.R.,Montreal Heart Institute | Fournier A.,University of Québec | And 9 more authors.
Methods | Year: 2016

In addition to cell surface membranes, numerous G protein-coupled receptors (GPCRs) are located on intracellular membranes including the nuclear envelope. Although the role of numerous GPCRs at the cell surface has been well characterized, the physiological function of these same receptors located on intracellular membranes remains to be determined. Here, we employ a novel caged Ang-II analog, cAng-II, to compare the effects of the activation of cell surface versus intracellular angiotensin receptors in intact cardiomyocytes. When added extracellularly to HEK 293 cells, Ang-II and photolysed cAng-II increased ERK1/2 phosphorylation (via AT1R) and cGMP production (AT2R). In contrast unphotolysed cAng-II did not. Cellular uptake of cAng-II was 6-fold greater than that of Ang-II and comparable to the HIV TAT(48-60) peptide. Intracellular photolysis of cAng-II induced an increase in nucleoplasmic Ca2+ ([Ca2+]n) that was greater than that induced by extracellular application of Ang-II. We conclude that cell-permeable ligands that can access intracellular GPCRs may evoke responses distinct from those with access restricted to the same receptor located on the cell surface. © 2015 Elsevier Inc.

Nguyen T.-T.M.,University of Québec | Nguyen T.-T.M.,Laboratoire International Associe Samuel Of Champlain | Letourneau M.,University of Québec | Letourneau M.,Laboratoire International Associe Samuel Of Champlain | And 4 more authors.
International Journal of Biochemistry and Cell Biology | Year: 2012

Urotensin II (UII) and its receptor UT, are widely expressed in the cardiovascular and central nervous system, where they exert regulatory actions under both physiological and pathological conditions. Our study, aimed at investigating the presence of functional nuclear UT in various rat and monkey tissues as well as in human cell lines, demonstrated for the first time by Western blot analysis and confocal immunofluorescence a tissue-specific nuclear expression of this receptor (heart and central nervous system). This nuclear UT was further characterized pharmacologically through radioligand binding studies using specific ligands of the urotensinergic system, as well as somatostatin. In 2D-gel experiments, we observed the presence of different post-translational modifications between membrane and nuclear UT receptors in brain extracts. Transcription initiation assays showed de novo RNA synthesis caused by UII and Urotensin-related peptide (URP) which were inhibited by an UT antagonist urantide. In hypoxic/ischemic conditions, UT receptors were differentially modulated in regard to subcellular localization. Thus, the unique regiospecificity of the nuclear UT receptor along with its particular modulation under hypoxic conditions could indicate a specific and complementary physiological role that could be correlated with pro-angiogenic and/or neuromodulatory actions of UII, both in the cardiovascular and central nervous system. © 2012 Elsevier Ltd. All rights reserved.

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