G Protein-coupled Receptor Kinase 5 in Cultured Vascular Smooth Muscle Cells and Rat Aorta

Abstract

GRK5, a recently cloned member of the G proteincoupled receptor kinase family, has been shown to phosphorylate and participate in the desensitization of angiotensin II (Ang II) type 1A (AT 1A ) receptors. In this study, the effect of angiotensin II on GRK5 expression was examined in cultured vascular smooth muscle cells and aortas of Ang II-infused hypertensive rats. In vascular smooth muscle cells, Ang II (100 nM) up-regulated GRK5 mRNA as early as 1 h, with a peak at 16 h. This up-regulation was dose-and calcium-dependent. The increase in GRK5 mRNA was reflected in a smaller increase in protein expression, which nonetheless had functional significance since AT 1 receptor phosphorylation was increased and phospholipase C activation was decreased following prolonged incubation with Ang II. In aortas of Ang II-infused hypertensive rats, both GRK5 mRNA and protein levels increased ϳ3-fold compared with sham-operated rats at 5 and 7 days, respectively. This up-regulation was blocked either by losartan or by the nonspecific vasodilator hydralazine. Since a subpressor dose of Ang II did not increase GRK5 mRNA levels and norepinephrine infusion also increased GRK5 mRNA expression, we conclude that Ang II-induced GRK5 up-regulation in rat aortas may be due to hypertension per se. Hormone-and hemodynamic stress-induced GRK5 regulation may provide a novel molecular basis for long-term regulation of agonist sensitivity of vascular cells. G protein-coupled receptor kinases (GRKs)1 are a recently identified family of enzymes that phosphorylate seven-transmembrane receptors. Receptor phosphorylation by GRKs subsequently promotes the binding of the regulatory arrestin protein, which results in uncoupling of the receptor and G protein (1). Six different mammalian GRKs have so far been cloned and characterized. Although GRK1 (rhodopsin kinase) and GRK4 are expressed exclusively in retinal photoreceptor cells and testis, respectively, ␤-adrenergic receptor kinase 1 (GRK2) and ␤-adrenergic receptor kinase 2 (GRK3), as well as GRK5 and GRK6, are widely distributed throughout the body. The range of substrates on which they may act and the mechanisms that regulate their expression are largely unknown.GRK5 is expressed in heart, placenta, lung, and skeletal muscle. GRK5 phosphorylates ␤ 1 -adrenergic, muscarinic, ␦-opioid, and angiotensin II (Ang II) type 1A (AT 1A ) receptors (1-4). In the same tissue or cell, two or more GRKs may be present and may target the same G protein-coupled receptors. For example, both GRK2 and GRK5 are expressed in heart, and both GRKs can target the ␤-adrenergic receptor (5). The relative importance of each GRK expressed in the same tissue may be tested by antisense transfection (6, 7), treatment with monoclonal antibodies (4), or a transgene technique (8, 9).Vascular smooth muscle cells (VSMC) express high levels of AT 1A receptors that rapidly desensitize coupling to phospholipase C upon agonist stimulation (10). These receptors also couple to phospholipase D, a response that continues for as long as th...

The G-protein-coupled receptor kinase 5 inhibits NFκB transcriptional activity by inducing nuclear accumulation of IκBα

“…Our in vivo and in vitro data are therefore in agreement showing the regulatory effect of GRK5-NT on NFB. It is interesting to note that upregulation of cellular GRK5 levels can be found in several conditions: Hypertension, for instance, presents increased GRK5 levels at the vasculature and impaired neoangiogenesis to chronic ischemia (45). Acute GRK5 upregulation to levels comparable with what we achieved in our setups and impairment of NFB-dependent responses, such as cytokine production and chemotaxis, have been observed in neutrophils from patients with sepsis (46).…”

Angiotensin II cell signaling: physiological and pathological effects in the cardiovascular system

“…For example, when AT 1 Rs are phosphorylated by GRK 5/6, ␤-arrestin-mediated ERK signaling is activated, independent of G protein signaling (92). Defects in the desensitization process have been implicated in vascular disease; indeed, ANG II-induced hypertensive rats appear to overexpress GRK-5, altering ANG II responsiveness (82).…”

Novel gp91 phox Homologues in Vascular Smooth Muscle Cells

“…Many proteins that mediate the cellular responses to Ang II are themselves regulated by this agonist, including the AT 1 receptor, 30 G␣q, 31 GRK-5, 32 and RGS-2. 33 To gain insight into a possible involvement of nox1 and nox4 in signaling, we measured the regulation of their messages by Ang II.…”