The International Journal of Biochemistry & Cell Biology
Presence of urotensin-II receptors at the cell nucleus: Specific tissue distribution and hypoxia-induced modulation
Introduction
In the classical view, peptide hormones bind to cell surface receptors, mostly G protein-coupled receptors (GPCR), and subsequently activate intracellular mediators of signal transduction. Over the last three decades, evidence has accumulated to indicate that at least some extracellular signalling molecules also bind and act within cells either after internalization into target cells or retention inside the producing cell (Re and Cook, 2006, Duverger et al., 1995, Gobeil et al., 2006). Defined as intracrines, such factors are quite diverse in structure and function (Re and Cook, 2008). Demonstration of an intracellular localization and trafficking of GPCRs both in vivo and in vitro was achieved (Koenig and Edwardson, 1997, Roth et al., 1981). However, only a few reports demonstrated the presence of functional GPCRs at the nucleus. Indeed, angiotensin II receptor (AT1), endothelin receptors (ETA) and (ETB) within the large family 1 of rhodopsin-like GPCRs, have been characterized in cell nuclei. In addition, the parathyroid hormone receptor and the metabotropic glutamate mGluR5 receptor, belonging respectively to family 2 and 3 of GPCRs, were also observed in nuclei (O’Malley et al., 2003). Interestingly, it was demonstrated that AT1 and endothelin-1 receptors present in the nuclei were able to induce intranuclear calcium mobilization and mRNA synthesis (Boivin et al., 2003, Eggena et al., 1993). These results suggested that nuclear receptors might represent a complementary system that could participate to the regulation of specific physiological action.
Urotensin II (UII), a disulfide bridged peptide originally isolated from the caudal neurosecretory system of a teleost fish (Pearson et al., 1980), is currently considered as the most potent mammalian vasoconstrictor identified so far (Douglas and Ohlstein, 2000). The urotensinergic system, composed of the urotensin II receptor (UT), UII and a paralogue peptide named urotensin II-related peptide (URP) (Vaudry et al., 2010), is not restricted to cardiac tissues. In fact, UT is widely expressed in the central nervous system as well as in various peripheral organs including kidney, liver, lung, skeletal muscle, pancreas, and adrenal gland (Jegou et al., 2006, Onan et al., 2004). UT activation is associated with multiple physiological actions such as vasoconstriction, vasodilation, osmoregulation, inhibition of glucose-induced insulin release, and cell proliferation (Vaudry et al., 2010). The multiple effects of UII and the broad expression pattern of its receptor indicate that the urotensinergic system could be involved in physiopathological processes. Indeed, correlations between different pathological states, i.e. chronic renal failure (Totsune et al., 2001), congestive heart failure (Gruson et al., 2006), hypertension (Cheung et al., 2004), diabetes mellitus (Totsune et al., 2003), and elevated concentration of plasmatic UII have been demonstrated.
The similarities existing between UII, endothelin-1 and angiotensin II, such as a strong contractile activity and receptors belonging to subclass 1A of GPCRs, prompted us to investigate the presence of UT at the surface of heart isolated nuclei. As we had hypothesized, our investigation confirmed the ectopic localization of functional nuclear UT receptors in heart (Doan et al., 2011). Consequently, based on our previous findings and the broad expression pattern of the urotensinergic system, we hypothesized that the presence of nuclear UT would not be limited to cardiac tissues.
In this paper, we investigated the presence of specific urotensin II binding sites both at the cellular and nuclear level in various rat and monkey tissues. Surprisingly, from a variety of tissues, nuclear UT receptors, as observed by Western blot and characterized by competitive binding assays, were only found in the central nervous system (CNS). Moreover, these nuclear receptors were functional as evaluated by a transcription initiation assay. Finally, we observed that cellular UT distribution was modulated under hypoxic conditions. Altogether, these results suggest a precise and complementary role for nuclear UT receptors.
Section snippets
Materials and methods
Materials, Total protein isolation, Isolation of membrane and nuclei from heart, Electrophoresis and Immunoblotting, Peptide Iodination, Nuclear Receptor Binding Assay, Confocal Microscopy, Transcription Initiation Assay, Effect of Cobalt chloride on UT receptor expression, Data Analysis are described in Supplemental Information.
Biochemical characterization of nuclear UII binding sites in rat tissues
A few years ago, the presence at the nuclear level of functional neuropeptide Y, endothelin-1 (ET-1), angiotensin II (AT), apelin and beta-adrenergic receptors suggested that nuclei were in fact a cell inside a cell (Boivin et al., 2008). The similarity between UII, AT and ET-1 in terms of physiological function led our group to investigate the presence of UII receptors at the nuclear surface. As such, we recently reported the presence of functional UT receptors in isolated rat heart nuclei (
Discussion
Hundreds of neurotransmitters regulate cellular development and function through G protein-coupled receptor signalling. GPCRs exert their effects by regulating ion channels, second messenger production, and protein kinase cascades, which in turn control cellular activity, gene expression, plasticity, differentiation, morphogenesis, and migration. In the recent years, the presence of endosomal receptors that could regulate complementary physiological action has almost become “a classic GPCR
Acknowledgements
This research was supported by the Canadian Institutes of Health Research. N.T.T.M is the recipient of a studentship from the Fondation Armand-Frappier.
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Novel insights into the role of urotensin II in cardiovascular disease
2019, Drug Discovery TodayCitation Excerpt :It was previously accepted that GPCRs would have a localization restricted to the cell membrane, nevertheless these receptors could also be found in the nuclear membrane, as happens with the UT receptor. The nuclear localization of this receptor, as occurs in the heart and CNS of rat and monkey tissues, implies ligand internalization through a receptor-independent mediated endocytosis, which could partially explain the pseudo-irreversible binding of UII [82,83]. Because higher levels of hUII were found in the cytoplasm compared with those of URP, it seems that variations in the N-terminal region could also explain the higher propensity of hUII to cross the plasma membrane.
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2016, Archives of Biochemistry and BiophysicsCitation Excerpt :The nuclear fractions does not contain endosomes as demonstrated by western blot experiments performed with an anti-RAB7 antibody. Following stimulation with the appropriate agonist, several GPCRs have been detected in the perinuclear or nuclear regions [49–51], whereas a perinuclear localization of the rat UT receptor has been observed without stimulation by its endogenous ligand [52]. The nuclear (or perinuclear) localization can be attributed to translocation of a GPCR from the cell surface or de novo synthesis of the receptor.
Intracrine endothelin signaling evokes IP3-dependent increases in nucleoplasmic Ca<sup>2+</sup> in adult cardiac myocytes
2013, Journal of Molecular and Cellular CardiologyCitation Excerpt :Similarly, dynorphin B, an agonist of the κ opioid receptor, increases opioid peptide transcription in isolated cardiac nuclei [19]. In nuclei isolated from non-cardiac cells, Ang II [20], bradykinin B2 [21], prostaglandin E2 [22–24], lysophosphatidic acid type-1 [25,26], metabotropic glutamate type-5 [27,28], thromboxane A2 [29,30], and urotensin-II [31] receptor activation also altered gene expression. Hence, endothelin receptors couple to effectors within the nuclear membrane and may be involved in stimulus–transcription coupling.