Showing papers by "Robert J. Lefkowitz published in 1997"
TL;DR: A mechanism previously shown to mediate uncoupling of the β2-adrenergic receptor from Gs and thus heterologous desensitization (PKA-mediated receptor phosphorylation), also serves to ‘switch’ coupling of this receptor fromGs to Gi and initiate a new set of signalling events.
Abstract: Many of the G-protein-coupled receptors for hormones that bind to the cell surface can signal to the interior of the cell through several different classes of G protein1,2,3,4. For example, although most of the actions of the prototype β2-adrenergic receptor are mediated through Gs proteins and the cyclic-AMP-dependent protein kinase (PKA) system5,6, β-adrenergic receptors can also couple to Gi proteins1,2. Here we investigate the mechanism that controls the specificity of this coupling. We show that in HEK293 cells, stimulation of mitogen-activated protein (MAP) kinase by the β2-adrenergic receptor is mediated by the βγ subunits of pertussis-toxin-sensitive G proteins through a pathway involving the non-receptor tyrosine kinase c-Src and the G protein Ras. Activation of this pathway by the β2-adrenergic receptor requires that the receptor be phosphorylated by PKA because it is blocked by H-89, an inhibitor of PKA. Additionally, a mutant of the receptor, which lacks the sites normally phosphorylated by PKA, can activate adenylyl cyclase5, the enzyme that generates cAMP, but not MAP kinase. Our results demonstrate that a mechanism previously shown to mediate uncoupling of the β2-adrenergic receptor from Gs and thus heterologous desensitization7 (PKA-mediated receptor phosphorylation), also serves to ‘switch’ coupling of this receptor from Gs to Gi and initiate a new set of signalling events.
1,284 citations
TL;DR: Data suggest that Gbetagamma subunit-mediated activation of Src family nonreceptor tyrosine kinases can account for the Gi-coupled receptor-mediated tyosine phosphorylation events that direct recruitment of the Shc and Grb2 adapter proteins to the membrane.
466 citations
TL;DR: Results indicate that calcium-calmodulin plays a central role in the calcium-dependent regulation of tyrosine phosphorylation by G protein-coupled receptors in some systems, and indicate that in HEK-293 cells, the Gβγ subunit-mediated α2A-AR- and the Gαq/11-mediated βARK1ct-mediated Erk1/2 activation pathways converge at the level of phospholipase C.
465 citations
TL;DR: It is suggested that a conformational change in the receptor induced by acidification of the endosomal vesicles is the key determinant regulating receptor dephosphorylation and resensitization.
379 citations
TL;DR: It is reported that phosphorylation of a carboxyl-terminal serine (Ser-412) in β-arrestin1 regulates its endocytotic but not its desensitization function, as with the classical endocytic adaptor protein complex AP2, which functions as a clathrin adaptor in receptor endocyTosis which is regulated by dephosphorylation at the plasma membrane.
241 citations
TL;DR: It is shown that reduction in or removal of the ability of the β2AR to be phosphorylated by βARK or to interact normally with β-arrestin substantially reduces agonist-mediated sequestration, and that the complement of cellular βARK and arrestin proteins synergistically regulateβ2AR sequestration.
Abstract: Two of the common mechanisms regulating G protein-coupled receptor (GPCR) signal transduction are phosphorylation and sequestration (internalization). Agonist-mediated receptor phosphorylation by the beta-adrenergic receptor kinase (betaARK) facilitates subsequent interaction with an arrestin protein, resulting in receptor desensitization. Studies of the beta2-adrenergic receptor (beta2AR) receptor in human embryonic kidney (HEK) 293 cells indicate that betaARK and arrestin proteins (beta-arrestins) also regulate sequestration. Consistent with this notion, we show in HEK 293 cells that reduction in or removal of the ability of the beta2AR to be phosphorylated by betaARK or to interact normally with beta-arrestin substantially reduces agonist-mediated sequestration. To evaluate betaARK and beta-arrestin regulation of beta2AR sequestration, we examined the relationship between betaARK and/or beta-arrestin expression and beta2AR sequestration in a variety of cultured cells, including HEK 293, COS 7, CHO, A431, and CHW. COS cells had both the lowest levels of endogenous beta-arrestin expression and beta2AR sequestration, whereas HEK 293 had the highest. Overexpression of beta-arrestin, but not betaARK, in COS cells increased the extent of wild-type beta2AR sequestration to levels observed in HEK 293 cells. However, a betaARK phosphorylation-impaired beta2AR mutant (Y326A) required the simultaneous overexpression of both betaARK and beta-arrestin for this to occur. Among all cell lines, sequestration correlated best with the product of betaARK and beta-arrestin expression. Moreover, an agonist-mediated translocation of wild-type beta2AR and endogenous beta-arrestin 2 to endocytic vesicles prepared from CHO fibroblasts was observed. These data suggest not only that the complement of cellular betaARK and arrestin proteins synergistically regulate beta2AR sequestration but also that beta-arrestins directly regulate beta2AR trafficking as well as desensitization.
238 citations
TL;DR: It is proposed that an additional step involving vesicle-mediated endocytosis is required for the rapid, Ras-dependent activation of Erk kinases in fibroblasts.
238 citations
TL;DR: This study provides a direct demonstration of a role for G betagamma in mediating the agonist-stimulated translocation of GRK2 and GRK3 in an intact cellular system and demonstrates isoform specificity in the interaction of these components.
Abstract: The G protein-coupled receptor (GPCR) kinases (GRKs) phosphorylate and desensitize agonist-occupied GPCRs. GRK2-mediated receptor phosphorylation is preceded by the agonist-dependent membrane association of this enzyme. Previous in vitro studies with purified proteins have suggested that this translocation may be mediated by the recruitment of GRK2 to the plasma membrane by its interaction with the free βγ subunits of heterotrimeric G proteins (Gβγ). Here we demonstrate that this mechanism operates in intact cells and that specificity is imparted by the selective interaction of discrete pools of Gβγ with receptors and GRKs. Treatment of Cos-7 cells transiently overexpressing GRK2 with a β-receptor agonist promotes a 3-fold increase in plasma membrane-associated GRK2. This translocation of GRK2 is inhibited by the carboxyl terminus of GRK2, a known Gβγ sequestrant. Furthermore, in cells overexpressing both GRK2 and Gβ1γ2, activation of lysophosphatidic acid receptors leads to the rapid and transient formation of a GRK/Gβγ complex. That Gβγ specificity exists at the level of the GPCR and the GRK is indicated by the observation that a GRK2/Gβγ complex is formed after agonist occupancy of the lysophosphatidic acid and β-adrenergic but not thrombin receptors. In contrast to GRK2, GRK3 forms a Gβγ complex after stimulation of all three GPCRs. This Gβγ binding specificity of the GRKs is also reflected at the level of the purified proteins. Thus the GRK2 carboxyl terminus binds Gβ1 and Gβ2 but not Gβ3, while the GRK3 fusion protein binds all three Gβ isoforms. This study provides a direct demonstration of a role for Gβγ in mediating the agonist-stimulated translocation of GRK2 and GRK3 in an intact cellular system and demonstrates isoform specificity in the interaction of these components.
198 citations
TL;DR: It is demonstrated that defects present in this critical myocardial signaling pathway can be corrected in vitro using genetic modification and raise the possibility of novel inotropic therapies for CHF including the inhibition of betaARK1 activity in the heart.
Abstract: Cardiovascular gene therapy is a novel approach to the treatment of diseases such as congestive heart failure (CHF). Gene transfer to the heart would allow for the replacement of defective or missing cellular proteins that may improve cardiac performance. Our laboratory has been focusing on the feasibility of restoring beta-adrenergic signaling deficiencies that are a characteristic of chronic CHF. We have now studied isolated ventricular myocytes from rabbits that have been chronically paced to produce hemodynamic failure. We document molecular beta-adrenergic signaling defects including down-regulation of myocardial beta-adrenergic receptors (beta-ARs), functional beta-AR uncoupling, and an up-regulation of the beta-AR kinase (betaARK1). Adenoviral-mediated gene transfer of the human beta2-AR or an inhibitor of betaARK1 to these failing myocytes led to the restoration of beta-AR signaling. These results demonstrate that defects present in this critical myocardial signaling pathway can be corrected in vitro using genetic modification and raise the possibility of novel inotropic therapies for CHF including the inhibition of betaARK1 activity in the heart.
197 citations
TL;DR: The phosphatases and regulatory mechanisms involved in this resensitization process have only recently begun to be elucidated and are included in this chapter.
Abstract: Publisher Summary Desensitization represents the summation of several different processes, including receptor phosphorylation, receptor sequestration (defined as the agonist-induced translocation of receptor away from the plasma membrane), enhanced degradation of intracellular messengers, and degradation of receptor protein. Rapid receptor desensitization, however, appears to be mediated by uncoupling of the receptor from its respective G protein, a consequence of receptor phosphorylation. In the case of the β 2 -adrenergic receptor ( β 2 AR), phosphorylation by two distinct classes of serine-threonine kinases leads to receptor desensitization. The first class, the second-messenger—dependent kinases—cyclic adenosine monophosphate (CAMP)-dependent protein kinase (PKA) and protein kinase C—phosphorylate and directly uncouple β 2 AR from G s . Because these kinases phosphorylate receptors in an agonist-independent manner, this process permits cross-talk between receptor families. The second class of enzymes, the G protein-coupled receptor kinases (GRKs), plays a highly specialized role in receptor desensitization because only agonist-occupied receptors serve as substrates for these enzymes. In the case of GRKs, the very signal that promotes activation of the G protein and the effector (that is, ligand binding) also promotes the desensitization of that specific receptor. Once uncoupled from the G protein, receptor function can be restored only by receptor dephosphorylation, a process termed resensitization . The phosphatases and regulatory mechanisms involved in this resensitization process have only recently begun to be elucidated and are included in this chapter.
194 citations
TL;DR: It is proposed that because of its affinity for ET-Rs demonstrated by co-immunoprecipitation, GRK2 is the most likely of the GRKs to initiate ET-R desensitization, and both the ETA-R and ETB-R can be regulated indistinguishably by GRK-initiated desensItization.
TL;DR: It is reported that cilia preparations derived from GRK3-deficient mice lack the fast agonist-induced desensitization normally seen after odorant stimulation, and total second messenger (cAMP) generation in these cilia Preparation following odorant stimulating is markedly reduced when compared with preparations from wild-type littermates.
TL;DR: These mice serve as a unique experimental model to study the in vivo interactions between α- and β-ARs in the heart and Interestingly, thesein vitro changes in signaling were reversed after treating the mice with pertussis toxin, suggesting that the extraordinarily high levels of WT α1B- ARs can lead to coupling to pertussi toxin-sensitive G proteins.
TL;DR: It is concluded that recombinant adenoviral gene transfer of the beta2AR or an inhibitor of betaARK-mediated desensitization can potentiate beta-adrenergic signaling.
Abstract: Our laboratory has been testing the hypothesis that genetic modulation of the beta-adrenergic signaling cascade can enhance cardiac function. We have previously shown that transgenic mice with cardiac overexpression of either the human beta2-adrenergic receptor (beta2AR) or an inhibitor of the beta-adrenergic receptor kinase (betaARK), an enzyme that phosphorylates and uncouples agonist-bound receptors, have increased myocardial inotropy. We now have created recombinant adenoviruses encoding either the beta2AR (Adeno-beta2AR) or a peptide betaARK inhibitor (consisting of the carboxyl terminus of betaARK1, Adeno-betaARKct) and tested their ability to potentiate beta-adrenergic signaling in cultured adult rabbit ventricular myocytes. As assessed by radioligand binding, Adeno-beta2AR infection led to approximately 20-fold overexpression of beta-adrenergic receptors. Protein immunoblots demonstrated the presence of the Adeno-betaARKct transgene. Both transgenes significantly increased isoproterenol-stimulated cAMP as compared to myocytes infected with an adenovirus encoding beta-galactosidase (Adeno-betaGal) but did not affect the sarcolemmal adenylyl cyclase response to Forskolin or NaF. beta-Adrenergic agonist-induced desensitization was significantly inhibited in Adeno-betaARKct-infected myocytes (16+/-2%) as compared to Adeno-betaGal-infected myocytes (37+/-1%, P < 0.001). We conclude that recombinant adenoviral gene transfer of the beta2AR or an inhibitor of betaARK-mediated desensitization can potentiate beta-adrenergic signaling.
Journal Article•
TL;DR: Studying on the specificity of beta-arrestin functions reveal a multiplicity of G protein-coupled receptor endocytic pathways and suggest that beta-Arrestins might serve as adaptors specifically targeting receptors for dynamin-dependent clathrin-mediated endocytosis.
Abstract: Dynamic regulation of G protein-coupled receptor signaling demands a coordinated balance between mechanisms leading to the generation, turning off and re-establishment of agonist-mediated signals. G protein-coupled receptor kinases (GRKs) and arrestin proteins not only mediate agonist-dependent G protein-coupled receptor desensitization, but also initiate the internalization (sequestration) of activated receptors, a process leading to receptor resensitization. Studies on the specificity of beta-arrestin functions reveal a multiplicity of G protein-coupled receptor endocytic pathways and suggest that beta-arrestins might serve as adaptors specifically targeting receptors for dynamin-dependent clathrin-mediated endocytosis. Moreover, inactivation of the GRK2 gene in mice has lead to the discovery of an unexpected role of GRK2 in cardiac development, further emphasizing the pleiotropic function of GRKs and arrestins.
Journal Article•
TL;DR: G-protein-coupled receptors that mediate cellular responses to a variety of humoral, endothelial-, or platelet-derived substances are able to stimulate MAP kinase activity, and the means by which the mitogenic signaling complex is assembled at the plasma membrane are subjects of further study.
Abstract: G-protein-coupled receptors that mediate cellular responses to a variety of humoral, endothelial-, or platelet-derived substances are able to stimulate MAP kinase activity. In transfected model systems, G-protein-coupled receptors that couple to pertussis toxin-insensitive G proteins of the Gq/11 family mediate this activation predominantly via a PKC-dependent mechanism. In contrast, activation of MAP kinase by receptors that couple to pertussis toxin-sensitive Gi proteins is PKC-independent and requires downstream activation of the low-molecular-weight G protein, Ras. This pathway can be inhibited by coexpression of peptides that sequester Gbetagamma subunits, and is mimicked by overexpression of Gbetagamma subunits. This Ras-dependent MAP kinase activation requires tyrosine phosphorylation of "docking proteins," including the shc adapter protein, and depends upon recruitment of Grb2/Sos1 complexes to the plasma membrane, thus resembling the pathway of MAP kinase activation employed by the receptor tyrosine kinases. Other molecules, including PI-3-kinases and phosphotyrosine phosphatases, probably also contribute to Gbetagamma-subunit-mediated assembly of a mitogenic signaling complex. Identification of the G-protein-coupled, receptor-regulated tyrosine kinase(s), and the means by which the mitogenic signaling complex is assembled at the plasma membrane, remain subjects of further study.
TL;DR: It is reported that transgenic mice expressing a mutated constitutively active form of the receptor (CAM) show no such phenotype, owing to its modest expression, and treatment of the animals with a variety of beta-adrenergic receptor ligands leads to a 50-fold increase in CAM beta2-adRenergic receptor expression, by stabilizing the CAM beta 2-ad Renergic receptor protein.
Abstract: (A) β2-adrenergic receptor density in the myocardium. NT, nontransgenic littermates (n = 8); CAM vehicle, vehicle-treated CAM β2-adrenergic receptor transgenic animals (n = 12); CAM ICI, ICI 118,551-treated transgenic animals (n = 8). Shown are means ± SEM. ∗∗, P < 0.01, CAM ICI vs. CAM vehicle; ∗, P < 0.05, CAM vehicle vs. NT (Student’s t test). (B) Adenylate cyclase activity in myocardial particulate fractions. Basal, no added drug; Iso, 100 μM isoproterenol. NT, n = 6; CAM vehicle, n = 6; CAM ICI, n = 5. ∗, P < 0.05, Iso vs. Basal; †, P < 0.05, Iso CAM vehicle vs. Iso NT; ††, P < 0.05, Iso CAM ICI vs. Iso CAM vehicle; ‡, P < 0.05, Basal CAM ICI vs. Basal CAM vehicle; Iso CAM vehicle vs. Basal CAM ICI, P = n.s.
TL;DR: It is concluded that Ang II-induced GRK5 up-regulation in rat aortas may be due to hypertension per se and may provide a novel molecular basis for long-term regulation of agonist sensitivity of vascular cells.
TL;DR: Current knowledge about the regulation of cardiovascular homeostasis involving signaling pathways through stimulation of adrenergic receptors is summarized.
Abstract: In disease states such as heart failure, catecholamines released from sympathetic nerve endings and the adrenal medulla play a central role in the adaptive and maladaptive physiological response to altered tissue perfusion. G protein-coupled receptors are importantly involved in myocardial growth and the regulation of contractility. The adrenergic receptors themselves are regulated by a set of specific kinases, termed the G protein-coupled receptor kinases. The study of complex systems in vivo has recently been advanced by the development of transgenic and gene-targeted "knockout" mouse models. Combining transgenic technology with sophisticated physiological measurements of cardiac function is an extremely powerful strategy for studying the regulation of myocardial contractility in normal animals and in models of disease states. The purpose of this review is to summarize current knowledge about the regulation of cardiovascular homeostasis involving signaling pathways through stimulation of adrenergic receptors.
TL;DR: It is within these two domains that the GRKs are most variable at the amino acid level, and the GRKS exhibit an absolute requirement for phospholipids not only for association with the membrane but also for activity.
Abstract: Membrane association is essential for GRK function and because of this the GRKs have evolved complex regulatory mechanisms for associating with the membrane. Although the GRKs are highly homologous, each kinase utilizes a distinct mechanism for associating with the membrane, which makes it unique within the family. Initially, the carboxyl terminus of the GRKs was identified as the “membrane association domain” but recent evidence suggests that the amino terminus may also play a critical role in localizing the kinases to the membrane (Murga et al., 1996; Pitcher et al, 1996). It is within these two domains that the GRKs are most variable at the amino acid level. The GRKS exhibit an absolute requirement for phospholipids not only for association with the membrane but also for activity. There are differences in preference and binding sites for the phospholipids within the GRK family, which may reflect differential targeting of the GRKs to G protein-coupled receptors situated in different lipid environments. There are hundreds of G protein-coupled receptors and only six known GRKs. All the GRKs appear to phosphorylate the same receptor substrates in vitro (Sterne-Marr & Benovic, 1995; Premont et al., 1995). Receptor specificity, in a cellular
TL;DR: The results suggest that phosducin may act as a phosphorylation-dependent switch in second messenger signaling cascades, regulating the kinetics of desensitization processes by controlling the activity of Gβγ-dependent GRKs.
TL;DR: Observations demonstrate that the cAMP and IP3/Ca2+/DAG cascades can cooperate in vivo toward the development of thyroid follicular cell malignancies.
Abstract: Proliferation of thyroid follicular cells is controlled by three intra-cellular cascades [cAMP, inositol 1,4,5-triphosphate (IP3)/Ca2+/diacylglycerol (DAG), and tyrosine kinases] that are activated by distinct extracellular signals and receptors. We had previously generated a transgenic mouse model in which the cAMP cascade was permanently stimulated in thyroid cells by an adenosine A2a receptor (Tg-A2aR model). In the present work, we have generated a transgenic model characterized by the chronic stimulation of both adenylyl cyclase and phospholipase C in thyroid follicular cells. The bovine thyroglobulin gene promoter was used to direct the expression of a constitutively active mutant of the alpha 1B adrenergic receptor, which is known to couple to both cascades in transfected cell lines. The expression of the transgene resulted, as expected, in the activation of phospholipase C and adenylyl cyclase, as demonstrated by the direct measurement of IP3 and cAMP in thyroid tissue. The phenotype resulting from this dual stimulation included growth stimulation, hyperfunction, cell degeneracy attributed to the overproduction of free radicals, and the development of malignant nodules invading the capsule, muscles, and blood vessels. Differentiated metastases were found occasionally in old animals. The development of malignant lesions was more frequent and of earlier onset than in our previous Tg-A2aR model, in which only the cAMP cascade was stimulated. These observations demonstrate that the cAMP and IP3/Ca2+/DAG cascades can cooperate in vivo toward the development of thyroid follicular cell malignancies.
TL;DR: In the absence of circulating catecholamines or basal sympathetic tone, beta ARK1 actions in single myocytes are minimal, and substantial functional beta ARk1 modulation of beta 1-AR signaling occurs in cardiac myocytes even during short-term beta 2-AR stimulation.
Abstract: To determine the direct functional significance of the beta-adrenergic receptor (AR) kinase 1 (beta ARK1) on myocardial performance in the absence of tonic sympathoadrenal neural activation and mechanical loading, we measured the contractile responses to acute beta 1-AR stimulation in left ventricular myocytes isolated from nontransgenic control (NTG) and transgenic mice overexpressing either beta ARK1 (TG beta K12) or a beta ARK1 inhibitor (TGMini27). Contractile response to five concentrations (10(-8)-10(-7) M) of the beta 1-AR agonist norepinephrine (NE) plus prazosin (10(-6) M) was measured after a 60-s rest, i.e., rested-state contraction (RSC), and during steady-state contraction (SSC) stimulation at 0.5 Hz (23 degrees C). At baseline, resting cell length was significantly greater in TG beta K12 myocytes (P < 0.05); however, there were no significant differences in RSC or SSC among NTG, TG beta K12, or TG Mini27 mice. On the other hand, both the dose-response curve and kinetics for the NE-induced SSC response normalized to RSC (SSC/RSC) were significantly different among experimental groups (P < 0.001). Specifically, maximal SSC induced by NE in myocytes isolated from TG beta K12 was only 70% of the response observed in NTG cells and 50% of the response measured in TGMini27. These data suggest that 1) in the absence of circulating catecholamines or basal sympathetic tone, beta ARK1 actions in single myocytes are minimal, and 2) substantial functional beta ARK1 modulation of beta 1-AR signaling occurs in cardiac myocytes even during short-term beta 1-AR stimulation. These results are consistent with a role for agonist-induced phosphorylation and desensitization of cardiac beta 1-ARs by beta ARK1 in single myocytes and highlight the potential functional importance of beta ARK1 as a critical determinant of the cardiac beta 1-AR contractile response.
TL;DR: An increase in myocardial adrenergic receptor density leads to significantly higher indices of cardiac function in transgenic mice compared to control mice, suggesting a possible alternative method of establishing a positive chronotropic and inotropic state without the use of pharmacological agents.
TL;DR: Current efforts to achieve myocardial gene transfer in several model systems are described, with particular emphasis placed on adenovirus-mediated gene delivery, its possibilities, and current limitations.
01 May 1997
TL;DR: Supporting the hypothesis that reversal of these beta-adrenergic defects may be able to restore cardiac inotropy to normal in patients with depressed systolic function is demonstrated, the demonstration that transgenic mice with cardiac overexpression of Gs alpha and enhanced myocardial responsiveness to isoproterenol develop myocardIAL fibrosis is demonstrated.
Abstract: The beta-adrenergic signaling cascade is an important regulator of myocardial function. Numerous abnormalities occur in this pathway and are associated with impaired cardiac contractility in patients with congestive heart failure (CHF). These signaling defects include downregulation of beta-adrenergic receptors (beta ARs) and increased levels of beta-adrenergic receptor kinase (beta ARK), an enzyme that phosphorylates and uncouples only agonist-bound receptors. Our laboratory has been testing the hypothesis that reversal of these beta-adrenergic defects may be able to restore cardiac inotropy to normal in patients with depressed systolic function. Transgenic mice with cardiac overexpression of beta 2ARs or an inhibitor of beta ARK have enhanced cardiac function as compared to wildtype littermates. Adenoviral vectors encoding the beta 2AR or beta ARK inhibitor potentiate beta AR signaling in cultured adult rabbit ventricular myocytes. However, a controversy has developed in the literature regarding whether increasing beta-adrenergic signaling would be beneficial or detrimental for patients with CHF. Those cautioning against this approach note that increased sympathetic activity is dangerous in CHF. Elevated catecholamine levels predict mortality and beta-agonists are not beneficial for survival, while recent studies suggest that beta-antagonists do improve outcome. Supporting these concerns is the demonstration that transgenic mice with cardiac overexpression of Gs alpha and enhanced myocardial responsiveness to isoproterenol develop myocardial fibrosis. This article summarizes this controversy; highlights important differences between overexpression of beta ARs or a beta ARK inhibitor, overexpression of Gs alpha, and administration of beta-agonists; and develops the hypothesis that these strategies may differ in their therapeutic efficacy in treating CHF.
TL;DR: The mechanisms whereby several GRs, including the α 1 B AR and α 2 A AR, stimulate ERK1/2 activity in transiently transfected COS-7 and Chinese hamster ovary cell model systems has been studied.
Abstract: Publisher Summary Until recently little has been known about the mechanisms of mitogenic signal transduction employed by receptors that couple to heterotrimeric G-proteins (GRs). These receptors participate in the regulation of cell proliferation in both physiological and pathophysiological states, and in cellular transformation in some, mostly neuroendocrine, human tumors. GRs that mediate cellular responses to a variety of humoral, endothelium-, or platelet-derived substances have been found to rapidly stimulate the mitogen-activated protein kinase (ERK1/2) pathway, a major point of convergence for signals regulating cell growth and differentiation. The best understood pathway of ERK1/2 activation is that mediated by growth factor receptors that possess intrinsic ligand-stimulated tyrosine kinase activity (RTK), such as the receptor for epidermal growth factor (EGF). To understand the mechanisms whereby GRs mediate growth regulatory signals, the mechanisms whereby several GRs, including the α 1 B AR and α 2 A AR, stimulate ERK1/2 activity in transiently transfected COS-7 and Chinese hamster ovary (CHO) cell model systems has been studied. In these cells, clear heterogeneity exists between the mechanisms of ERK1/2 activation employed by receptors that signal via pertussis toxin-sensitive G i family proteins, such as α 2 A AR, and receptors that signal via pertussis toxininsensitive G q/11 family proteins, such as α 1 B AR.
Patent•
03 Oct 1997TL;DR: In this article, a method of inhibiting arterial and venous smooth muscle proliferation resulting from arterial injury or vein grafting was proposed, and an expression construct encoding a Gβη inhibitor suitable for use in such a method.
Abstract: The present invention relates, in general, to vascular smooth muscle proliferation and, in particular, to a method of inhibiting arterial and venous smooth muscle proliferation resulting, for example, from arterial injury or vein grafting. The invention also relates to an expression construct encoding a Gβη inhibitor suitable for use in such a method.