The Experts below are selected from a list of 285 Experts worldwide ranked by ideXlab platform
Marc G Caron - One of the best experts on this subject based on the ideXlab platform.
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akt gsk3 signaling in the action of psychotropic drugs
Annual Review of Pharmacology and Toxicology, 2009Co-Authors: Jeanmartin Beaulieu, Marc G CaronAbstract:Psychotropic drugs acting on monoamine neurotransmission are major pharmacological treatments for neuropsychiatric conditions such as schizophrenia, depression, bipolar disorder, Tourette syndrome, ADHD, and Alzheimer disease. Independent lines of research involving biochemical and behavioral approaches in normal and/or genetically modified mice provide converging evidence for an involvement of the signaling molecules Akt and glycogen synthase kinase-3 (GSK3) in the regulation of behavior by dopamine and serotonin (5-HT). These signaling molecules have also received attention for their role in the actions of psychoactive drugs such as antidepressants, antipsychotics, lithium, and other mood stabilizers. Furthermore, investigations of the mechanism by which D2 dopamine receptors regulate Akt/GSK3 signaling strongly support the physiological relevance of a new modality of G protein–coupled receptor (GPCR) signaling involving the multifunctional scaffolding protein Beta-Arrestin 2. Elucidation of the contrib...
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akt gsk3 signaling in the action of psychotropic drugs
Annual Review of Pharmacology and Toxicology, 2009Co-Authors: Jeanmartin Beaulieu, Raul R. Gainetdinov, Marc G CaronAbstract:Psychotropic drugs acting on monoamine neurotransmission are major pharmacological treatments for neuropsychiatric conditions such as schizophrenia, depression, bipolar disorder, Tourette syndrome, ADHD, and Alzheimer disease. Independent lines of research involving biochemical and behavioral approaches in normal and/or genetically modified mice provide converging evidence for an involvement of the signaling molecules Akt and glycogen synthase kinase-3 (GSK3) in the regulation of behavior by dopamine and serotonin (5-HT). These signaling molecules have also received attention for their role in the actions of psychoactive drugs such as antidepressants, antipsychotics, lithium, and other mood stabilizers. Furthermore, investigations of the mechanism by which D2 dopamine receptors regulate Akt/GSK3 signaling strongly support the physiological relevance of a new modality of G protein-coupled receptor (GPCR) signaling involving the multifunctional scaffolding protein Beta-Arrestin 2. Elucidation of the contribution of multiple signaling pathways to the action of psychotropic drugs may provide a better biological understanding of psychiatric disorders and lead to more efficient therapeutics.
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an akt β Arrestin 2 pp2a signaling complex mediates dopaminergic neurotransmission and behavior
Cell, 2005Co-Authors: Jeanmartin Beaulieu, Robert J Lefkowitz, Raul R. Gainetdinov, Tatyana D Sotnikova, Sebastien Marion, Marc G CaronAbstract:Dopamine plays an important role in the etiology of schizophrenia, and D2 class dopamine receptors are the best-established target of antipsychotic drugs. Here we show that D2 class-receptor-mediated Akt regulation involves the formation of signaling complexes containing Beta-Arrestin 2, PP2A, and Akt. Beta-Arrestin 2 deficiency in mice results in reduction of dopamine-dependent behaviors, loss of Akt regulation by dopamine in the striatum, and disruption of the dopamine-dependent interaction of Akt with its negative regulator, protein phosphatase 2A. Importantly, canonical cAMP-mediated dopamine-receptor signaling is not inhibited in the absence of Beta-Arrestin 2. These results demonstrate that, apart from its classical function in receptor desensitization, Beta-Arrestin 2 also acts as a signaling intermediate through a kinase/phosphatase scaffold. Furthermore, this function of Beta-Arrestin 2 is important for the expression of dopamine-associated behaviors, thus implicating Beta-Arrestin 2 as a positive mediator of dopaminergic synaptic transmission and a potential pharmacological target for dopamine-related psychiatric disorders.
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Beta Arrestin 2 regulates zebrafish development through the hedgehog signaling pathway
Science, 2004Co-Authors: Larry S Barak, Marc G Caron, Alyson Wilbanks, Gregory B Fralish, Margaret L KirbyAbstract:β-Arrestins are multifunctional proteins that act as scaffolds and transducers of intracellular signals from heptahelical transmembrane-spanning receptors (7TMR). Hedgehog (Hh) signaling, which uses the putative 7TMR, Smoothened, is established as a fundamental pathway in development, and unregulated Hh signaling is associated with certain malignancies. Here, we show that the functional knockdown of β-Arrestin 2 in zebrafish embryos recapitulates the many phenotypes of Hh pathway mutants. Expression of wild-type β-Arrestin 2, or constitutive activation of the Hh pathway downstream of Smoothened, rescues the phenotypes caused by β-Arrestin 2 deficiency. These results suggest that a functional interaction between β-Arrestin 2 and Smoothened may be critical to regulate Hh signaling in zebrafish development.
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dishevelled 2 recruits Beta Arrestin 2 to mediate wnt5a stimulated endocytosis of frizzled 4
Science, 2003Co-Authors: Wei Chen, Roel Nusse, Marc G Caron, Derk Ten Berge, J M Brown, Liaoyuan A Hu, Robert J LefkowitzAbstract:Wnt proteins, regulators of development in many organisms, bind to seven transmembrane-spanning (7TMS) receptors called frizzleds, thereby recruiting the cytoplasmic molecule dishevelled (Dvl) to the plasma membrane.Frizzled-mediated endocytosis of Wg (a Drosophila Wnt protein) and lysosomal degradation may regulate the formation of morphogen gradients. Endocytosis of Frizzled 4 (Fz4) in human embryonic kidney 293 cells was dependent on added Wnt5A protein and was accomplished by the multifunctional adaptor protein Beta-Arrestin 2 (Betaarr2), which was recruited to Fz4 by binding to phosphorylated Dvl2. These findings provide a previously unrecognized mechanism for receptor recruitment of Beta-Arrestin and demonstrate that Dvl plays an important role in the endocytosis of frizzled, as well as in promoting signaling.
Robert J Lefkowitz - One of the best experts on this subject based on the ideXlab platform.
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an akt β Arrestin 2 pp2a signaling complex mediates dopaminergic neurotransmission and behavior
Cell, 2005Co-Authors: Jeanmartin Beaulieu, Robert J Lefkowitz, Raul R. Gainetdinov, Tatyana D Sotnikova, Sebastien Marion, Marc G CaronAbstract:Dopamine plays an important role in the etiology of schizophrenia, and D2 class dopamine receptors are the best-established target of antipsychotic drugs. Here we show that D2 class-receptor-mediated Akt regulation involves the formation of signaling complexes containing Beta-Arrestin 2, PP2A, and Akt. Beta-Arrestin 2 deficiency in mice results in reduction of dopamine-dependent behaviors, loss of Akt regulation by dopamine in the striatum, and disruption of the dopamine-dependent interaction of Akt with its negative regulator, protein phosphatase 2A. Importantly, canonical cAMP-mediated dopamine-receptor signaling is not inhibited in the absence of Beta-Arrestin 2. These results demonstrate that, apart from its classical function in receptor desensitization, Beta-Arrestin 2 also acts as a signaling intermediate through a kinase/phosphatase scaffold. Furthermore, this function of Beta-Arrestin 2 is important for the expression of dopamine-associated behaviors, thus implicating Beta-Arrestin 2 as a positive mediator of dopaminergic synaptic transmission and a potential pharmacological target for dopamine-related psychiatric disorders.
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Beta Arrestin 1 and galphaq 11 coordinately activate rhoa and stress fiber formation following receptor stimulation
Journal of Biological Chemistry, 2005Co-Authors: William G Barnes, Graeme Milligan, Eric Reiter, Jonathan D Violin, Xiurong Ren, Robert J LefkowitzAbstract:Beta-Arrestins were initially shown, in conjunction with G protein-coupled receptor kinases, to be involved in the desensitization and internalization of activated seven-transmembrane receptors. Recently, Beta-Arrestin 2 has been shown to act as a signal mediator in mitogen-activated protein kinase cascades and to play a positive regulatory role in chemotaxis. We now show that Beta-Arrestin 1 is required to activate the small GTPase RhoA leading to the re-organization of stress fibers following the activation of the angiotensin II type 1A receptor. This angiotensin II type 1A receptor-directed RhoA activation and stress fiber formation also require the activation of the heterotrimeric G protein G(alphaq/11). Whereas neither Beta-Arrestin 1 nor G(alphaq/11) activation alone is sufficient to robustly activate RhoA, the concurrent recruitment of Beta-Arrestin 1 and activation of G(alphaq/11) leads to full activation of RhoA and to the subsequent formation of stress fibers.
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dishevelled 2 recruits Beta Arrestin 2 to mediate wnt5a stimulated endocytosis of frizzled 4
Science, 2003Co-Authors: Wei Chen, Roel Nusse, Marc G Caron, Derk Ten Berge, J M Brown, Liaoyuan A Hu, Robert J LefkowitzAbstract:Wnt proteins, regulators of development in many organisms, bind to seven transmembrane-spanning (7TMS) receptors called frizzleds, thereby recruiting the cytoplasmic molecule dishevelled (Dvl) to the plasma membrane.Frizzled-mediated endocytosis of Wg (a Drosophila Wnt protein) and lysosomal degradation may regulate the formation of morphogen gradients. Endocytosis of Frizzled 4 (Fz4) in human embryonic kidney 293 cells was dependent on added Wnt5A protein and was accomplished by the multifunctional adaptor protein Beta-Arrestin 2 (Betaarr2), which was recruited to Fz4 by binding to phosphorylated Dvl2. These findings provide a previously unrecognized mechanism for receptor recruitment of Beta-Arrestin and demonstrate that Dvl plays an important role in the endocytosis of frizzled, as well as in promoting signaling.
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differential mechanisms of morphine antinociceptive tolerance revealed in Beta Arrestin 2 knock out mice
The Journal of Neuroscience, 2002Co-Authors: Laura M. Bohn, Robert J Lefkowitz, Marc G CaronAbstract:Morphine induces antinociception by activating μ opioid receptors (μORs) in spinal and supraspinal regions of the CNS. βArrestin-2 (βarr2), a G-protein-coupled receptor-regulating protein, regulates the μOR in vivo . We have shown previously that mice lacking βarr2 experience enhanced morphine-induced analgesia and do not become tolerant to morphine as determined in the hot-plate test, a paradigm that primarily assesses supraspinal pain responsiveness. To determine the general applicability of the βarr2-μOR interaction in other neuronal systems, we have, in the present study, tested βarr2 knock-out (βarr2-KO) mice using the warm water tail-immersion paradigm, which primarily assesses spinal reflexes to painful thermal stimuli. In this test, the βarr2-KO mice have greater basal nociceptive thresholds and markedly enhanced sensitivity to morphine. Interestingly, however, after a delayed onset, they do ultimately develop morphine tolerance, although to a lesser degree than the wild-type (WT) controls. In the βarr2-KO but not WT mice, morphine tolerance can be completely reversed with a low dose of the classical protein kinase C (PKC) inhibitor chelerythrine. These findings provide in vivo evidence that the μOR is differentially regulated in diverse regions of the CNS. Furthermore, although βarr2 appears to be the most prominent and proximal determinant of μOR desensitization and morphine tolerance, in the absence of this mechanism, the contributions of a PKC-dependent regulatory system become readily apparent.
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Beta Arrestin 1 and 2 differentially regulate heptahelical receptor signaling and trafficking
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Trudy A Kohout, Fang-tsyr Lin, Stephen J. Perry, David A Conner, Robert J LefkowitzAbstract:The two widely coexpressed isoforms of Beta-Arrestin (termed Beta Arrestin 1 and 2) are highly similar in amino acid sequence. The Beta-Arrestins bind phosphorylated heptahelical receptors to desensitize and target them to clathrin-coated pits for endocytosis. To better define differences in the roles of Beta-Arrestin 1 and 2, we prepared mouse embryonic fibroblasts from knockout mice that lack one of the Beta-Arrestins (Beta arr1-KO and Beta arr2-KO) or both (Beta arr1/2-KO), as well as their wild-type (WT) littermate controls. These cells were analyzed for their ability to support desensitization and sequestration of the Beta(2)-adrenergic receptor (Beta(2)-AR) and the angiotensin II type 1A receptor (AT(1A)-R). Both Beta arr1-KO and Beta arr2-KO cells showed similar impairment in agonist-stimulated Beta(2)-AR and AT(1A)-R desensitization, when compared with their WT control cells, and the Beta arr1/2-KO cells were even further impaired. Sequestration of the Beta(2)-AR in the Beta arr2-KO cells was compromised significantly (87% reduction), whereas in the Beta arr1-KO cells it was not. Agonist-stimulated internalization of the AT(1A)-R was only slightly reduced in the Beta arr1-KO but was unaffected in the Beta arr2-KO cells. In the Beta arr1/2-KO cells, the sequestration of both receptors was dramatically reduced. Comparison of the ability of the two Beta-Arrestins to sequester the Beta(2)-AR revealed Beta-Arrestin 2 to be 100-fold more potent than Beta-Arrestin 1. Down-regulation of the Beta(2)-AR was also prevented in the Beta arr1/2-KO cells, whereas no change was observed in the single knockout cells. These findings suggest that sequestration of various heptahelical receptors is regulated differently by the two Beta-Arrestins, whereas both isoforms are capable of supporting receptor desensitization and down-regulation.
Laura M. Bohn - One of the best experts on this subject based on the ideXlab platform.
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morphine side effects in β Arrestin 2 knockout mice
Journal of Pharmacology and Experimental Therapeutics, 2005Co-Authors: Kirsten M Raehal, Julia K L Walker, Laura M. BohnAbstract:Morphine is a potent analgesic, yet, like most opioid narcotics, it exerts unwanted side effects such as constipation and respiratory suppression, thereby limiting its clinical utility. Pharmacological approaches taken to preserve the analgesic properties, while eliminating the unwanted side effects, have met with very limited success. Here, we provide evidence that altering mu opioid receptor regulation may provide a novel approach to discriminate morphine's beneficial and deleterious effects in vivo. We have previously reported that mice lacking the G protein-coupled receptor regulatory protein, Beta-Arrestin 2, display profoundly altered morphine responses. Beta-Arrestin 2 knockout mice have enhanced and prolonged morphine analgesia with very little morphine tolerance. In this report, we examine whether the side effects of morphine treatment are also augmented in this animal model. Surprisingly, the genetic disruption of opioid receptor regulation, while enhancing and prolonging analgesia, dramatically attenuates the respiratory suppression and acute constipation caused by morphine.
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differential mechanisms of morphine antinociceptive tolerance revealed in Beta Arrestin 2 knock out mice
The Journal of Neuroscience, 2002Co-Authors: Laura M. Bohn, Robert J Lefkowitz, Marc G CaronAbstract:Morphine induces antinociception by activating μ opioid receptors (μORs) in spinal and supraspinal regions of the CNS. βArrestin-2 (βarr2), a G-protein-coupled receptor-regulating protein, regulates the μOR in vivo . We have shown previously that mice lacking βarr2 experience enhanced morphine-induced analgesia and do not become tolerant to morphine as determined in the hot-plate test, a paradigm that primarily assesses supraspinal pain responsiveness. To determine the general applicability of the βarr2-μOR interaction in other neuronal systems, we have, in the present study, tested βarr2 knock-out (βarr2-KO) mice using the warm water tail-immersion paradigm, which primarily assesses spinal reflexes to painful thermal stimuli. In this test, the βarr2-KO mice have greater basal nociceptive thresholds and markedly enhanced sensitivity to morphine. Interestingly, however, after a delayed onset, they do ultimately develop morphine tolerance, although to a lesser degree than the wild-type (WT) controls. In the βarr2-KO but not WT mice, morphine tolerance can be completely reversed with a low dose of the classical protein kinase C (PKC) inhibitor chelerythrine. These findings provide in vivo evidence that the μOR is differentially regulated in diverse regions of the CNS. Furthermore, although βarr2 appears to be the most prominent and proximal determinant of μOR desensitization and morphine tolerance, in the absence of this mechanism, the contributions of a PKC-dependent regulatory system become readily apparent.
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mu opioid receptor desensitization by Beta Arrestin 2 determines morphine tolerance but not dependence
Nature, 2000Co-Authors: Laura M. Bohn, Robert J Lefkowitz, Raul R. Gainetdinov, Fang-tsyr Lin, Marc G CaronAbstract:μ-Opioid receptor desensitization by β-Arrestin-2 determines morphine tolerance but not dependence
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Enhanced morphine analgesia in mice lacking Beta-Arrestin 2.
Science (New York N.Y.), 1999Co-Authors: Laura M. Bohn, Robert J Lefkowitz, Marc G Caron, Raul R. Gainetdinov, Karsten Peppel, Fang-tsyr LinAbstract:The ability of morphine to alleviate pain is mediated through a heterotrimeric guanine nucleotide binding protein (G protein)-coupled heptahelical receptor (GPCR), the mu opioid receptor (muOR). The efficiency of GPCR signaling is tightly regulated and ultimately limited by the coordinated phosphorylation of the receptors by specific GPCR kinases and the subsequent interaction of the phosphorylated receptors with Beta-Arrestin 1 and Beta-Arrestin 2. Functional deletion of the Beta-Arrestin 2 gene in mice resulted in remarkable potentiation and prolongation of the analgesic effect of morphine, suggesting that muOR desensitization was impaired. These results provide evidence in vivo for the physiological importance of Beta-Arrestin 2 in regulating the function of a specific GPCR, the muOR. Moreover, they suggest that inhibition of Beta-Arrestin 2 function might lead to enhanced analgesic effectiveness of morphine and provide potential new avenues for the study and treatment of pain, narcotic tolerance, and dependence.
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enhanced morphine analgesia in mice lacking Beta Arrestin 2
Science, 1999Co-Authors: Laura M. Bohn, Robert J Lefkowitz, Raul R. Gainetdinov, Karsten Peppel, Marc G CaronAbstract:The ability of morphine to alleviate pain is mediated through a heterotrimeric guanine nucleotide binding protein (G protein)–coupled heptahelical receptor (GPCR), the μ opioid receptor (μOR). The efficiency of GPCR signaling is tightly regulated and ultimately limited by the coordinated phosphorylation of the receptors by specific GPCR kinases and the subsequent interaction of the phosphorylated receptors with β-Arrestin 1 and β-Arrestin 2. Functional deletion of the β-Arrestin 2 gene in mice resulted in remarkable potentiation and prolongation of the analgesic effect of morphine, suggesting that μOR desensitization was impaired. These results provide evidence in vivo for the physiological importance of β-Arrestin 2 in regulating the function of a specific GPCR, the μOR. Moreover, they suggest that inhibition of β-Arrestin 2 function might lead to enhanced analgesic effectiveness of morphine and provide potential new avenues for the study and treatment of pain, narcotic tolerance, and dependence.
Gang Pei - One of the best experts on this subject based on the ideXlab platform.
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deficiency of a Beta Arrestin 2 signal complex contributes to insulin resistance
Nature, 2009Co-Authors: Bing Luan, Jian Zhao, Baoyu Duan, Guangwen Shu, Xiaoying Wang, Weiping Jia, Jiuhong Kang, Gang PeiAbstract:The insulin resistance characteristic of type 2 diabetes and obesity is caused by the failure of insulin to stimulate receptor signalling. Defining the cellular mechanisms of this defect is critical to understanding these disorders. Experiments in type 2 diabetes clinical samples and mouse models now show that the scaffold protein β-Arrestin-2 is necessary for efficient insulin signalling, linking the downstream kinases Akt and Src to the insulin receptor. β-Arrestin-2 is downregulated both in diabetic mice and in patients. Without β-Arrestin-2, insulin resistance develops, and reinstating its expression restores insulin sensitivity in mice. This suggests possible new therapeutic targets in insulin resistance and its related disorders. Beta-Arrestin-2, an adaptor protein, is necessary for efficient insulin signalling by scaffolding downstream kinases, Akt and Src, to the insulin receptor. Without Beta-Arrestin-2 insulin resistance develops. Insulin resistance, a hallmark of type 2 diabetes, is a defect of insulin in stimulating insulin receptor signalling1,2, which has become one of the most serious public health threats. Upon stimulation by insulin, insulin receptor recruits and phosphorylates insulin receptor substrate proteins3, leading to activation of the phosphatidylinositol-3-OH kinase (PI(3)K)–Akt pathway. Activated Akt phosphorylates downstream kinases and transcription factors, thus mediating most of the metabolic actions of insulin4,5,6. β-Arrestins mediate biological functions of G-protein-coupled receptors by linking activated receptors with distinct sets of accessory and effecter proteins, thereby determining the specificity, efficiency and capacity of signals7,8,9,10,11. Here we show that in diabetic mouse models, β-Arrestin-2 is severely downregulated. Knockdown of β-Arrestin-2 exacerbates insulin resistance, whereas administration of β-Arrestin-2 restores insulin sensitivity in mice. Further investigation reveals that insulin stimulates the formation of a new β-Arrestin-2 signal complex, in which β-Arrestin-2 scaffolds Akt and Src to insulin receptor. Loss or dysfunction of β-Arrestin-2 results in deficiency of this signal complex and disturbance of insulin signalling in vivo, thereby contributing to the development of insulin resistance and progression of type 2 diabetes. Our findings provide new insight into the molecular pathogenesis of insulin resistance, and implicate new preventive and therapeutic strategies against insulin resistance and type 2 diabetes.
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deficiency of a β Arrestin 2 signal complex contributes to insulin resistance
Nature, 2009Co-Authors: Bing Luan, Jian Zhao, Baoyu Duan, Guangwen Shu, Xiaoying Wang, Weiping Jia, Jiuhong Kang, Gang PeiAbstract:The insulin resistance characteristic of type 2 diabetes and obesity is caused by the failure of insulin to stimulate receptor signalling. Defining the cellular mechanisms of this defect is critical to understanding these disorders. Experiments in type 2 diabetes clinical samples and mouse models now show that the scaffold protein β-Arrestin-2 is necessary for efficient insulin signalling, linking the downstream kinases Akt and Src to the insulin receptor. β-Arrestin-2 is downregulated both in diabetic mice and in patients. Without β-Arrestin-2, insulin resistance develops, and reinstating its expression restores insulin sensitivity in mice. This suggests possible new therapeutic targets in insulin resistance and its related disorders. Beta-Arrestin-2, an adaptor protein, is necessary for efficient insulin signalling by scaffolding downstream kinases, Akt and Src, to the insulin receptor. Without Beta-Arrestin-2 insulin resistance develops. Insulin resistance, a hallmark of type 2 diabetes, is a defect of insulin in stimulating insulin receptor signalling1,2, which has become one of the most serious public health threats. Upon stimulation by insulin, insulin receptor recruits and phosphorylates insulin receptor substrate proteins3, leading to activation of the phosphatidylinositol-3-OH kinase (PI(3)K)–Akt pathway. Activated Akt phosphorylates downstream kinases and transcription factors, thus mediating most of the metabolic actions of insulin4,5,6. β-Arrestins mediate biological functions of G-protein-coupled receptors by linking activated receptors with distinct sets of accessory and effecter proteins, thereby determining the specificity, efficiency and capacity of signals7,8,9,10,11. Here we show that in diabetic mouse models, β-Arrestin-2 is severely downregulated. Knockdown of β-Arrestin-2 exacerbates insulin resistance, whereas administration of β-Arrestin-2 restores insulin sensitivity in mice. Further investigation reveals that insulin stimulates the formation of a new β-Arrestin-2 signal complex, in which β-Arrestin-2 scaffolds Akt and Src to insulin receptor. Loss or dysfunction of β-Arrestin-2 results in deficiency of this signal complex and disturbance of insulin signalling in vivo, thereby contributing to the development of insulin resistance and progression of type 2 diabetes. Our findings provide new insight into the molecular pathogenesis of insulin resistance, and implicate new preventive and therapeutic strategies against insulin resistance and type 2 diabetes.
Trudy A Kohout - One of the best experts on this subject based on the ideXlab platform.
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differential desensitization receptor phosphorylation Beta Arrestin recruitment and erk1 2 activation by the two endogenous ligands for the cc chemokine receptor 7
Journal of Biological Chemistry, 2004Co-Authors: Trudy A Kohout, Shelby Nicholas, Stephen J. Perry, Greg J. Reinhart, Sachiko Junger, Scott R StruthersAbstract:Many members of the chemokine receptor family of G protein-coupled receptors utilize multiple endogenous ligands. However, differences between the signaling properties of multiple chemokines through a single receptor have yet to be well characterized. In this study we investigated the early signaling events of CCR7 initiated by its two endogenous ligands, CCL19 and CCL21. Both CCL19 and CCL21 induce G protein activation and calcium mobilization with equal potency. However, only activation by CCL19, not CCL21, promotes robust desensitization of endogenous CCR7 in the human T cell lymphoma cell line H9. Desensitization occurs through the induction of receptor phosphorylation and Beta-Arrestin recruitment (shown in HEK293 cells expressing CCR7-FLAG). The sites of CCL19-induced phosphorylation were mapped by mutating to alanines the serines and threonines found within kinase phosphorylation consensus sequences in the carboxyl terminus of CCR7. A cluster of sites, including Thr-373-376 and Ser-378 is important for CCL19-mediated phosphorylation of the receptor, whereas residues serine 356, 357, 364, and 365 are important for basal receptor phosphorylation by protein kinase C. Activation of CCR7 by both ligands leads to signaling to the ERK1/2 mitogen-activated protein kinase pathway. However, CCL19 promotes 4-fold more ERK1/2 phosphorylation than does CCL21. The mechanism by which CCL19 activates ERK1/2 was determined to be Beta-Arrestin-dependent, because it is reduced both by depletion of Beta-Arrestin-2 with small interfering RNA and by elimination of the phosphorylation sites in the tail of the receptor. Taken together, these findings demonstrate that CCL19 and CCL21 place CCR7 in functionally distinct conformations that are independent of their G protein-coupling potency: one that allows the efficient desensitization of the receptor and activation of ERK1/2, and another that is impaired in these functions.
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differential desensitization receptor phosphorylation β Arrestin recruitment and erk1 2 activation by the two endogenous ligands for the cc chemokine receptor 7
Journal of Biological Chemistry, 2004Co-Authors: Trudy A Kohout, Stephen J. Perry, Greg J. Reinhart, Sachiko Junger, Shelby L Nicholas, Scott R StruthersAbstract:Many members of the chemokine receptor family of G protein-coupled receptors utilize multiple endogenous ligands. However, differences between the signaling properties of multiple chemokines through a single receptor have yet to be well characterized. In this study we investigated the early signaling events of CCR7 initiated by its two endogenous ligands, CCL19 and CCL21. Both CCL19 and CCL21 induce G protein activation and calcium mobilization with equal potency. However, only activation by CCL19, not CCL21, promotes robust desensitization of endogenous CCR7 in the human T cell lymphoma cell line H9. Desensitization occurs through the induction of receptor phosphorylation and Beta-Arrestin recruitment (shown in HEK293 cells expressing CCR7-FLAG). The sites of CCL19-induced phosphorylation were mapped by mutating to alanines the serines and threonines found within kinase phosphorylation consensus sequences in the carboxyl terminus of CCR7. A cluster of sites, including Thr-373-376 and Ser-378 is important for CCL19-mediated phosphorylation of the receptor, whereas residues serine 356, 357, 364, and 365 are important for basal receptor phosphorylation by protein kinase C. Activation of CCR7 by both ligands leads to signaling to the ERK1/2 mitogen-activated protein kinase pathway. However, CCL19 promotes 4-fold more ERK1/2 phosphorylation than does CCL21. The mechanism by which CCL19 activates ERK1/2 was determined to be Beta-Arrestin-dependent, because it is reduced both by depletion of Beta-Arrestin-2 with small interfering RNA and by elimination of the phosphorylation sites in the tail of the receptor. Taken together, these findings demonstrate that CCL19 and CCL21 place CCR7 in functionally distinct conformations that are independent of their G protein-coupling potency: one that allows the efficient desensitization of the receptor and activation of ERK1/2, and another that is impaired in these functions.
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Beta Arrestin 1 and 2 differentially regulate heptahelical receptor signaling and trafficking
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Trudy A Kohout, Fang-tsyr Lin, Stephen J. Perry, David A Conner, Robert J LefkowitzAbstract:The two widely coexpressed isoforms of Beta-Arrestin (termed Beta Arrestin 1 and 2) are highly similar in amino acid sequence. The Beta-Arrestins bind phosphorylated heptahelical receptors to desensitize and target them to clathrin-coated pits for endocytosis. To better define differences in the roles of Beta-Arrestin 1 and 2, we prepared mouse embryonic fibroblasts from knockout mice that lack one of the Beta-Arrestins (Beta arr1-KO and Beta arr2-KO) or both (Beta arr1/2-KO), as well as their wild-type (WT) littermate controls. These cells were analyzed for their ability to support desensitization and sequestration of the Beta(2)-adrenergic receptor (Beta(2)-AR) and the angiotensin II type 1A receptor (AT(1A)-R). Both Beta arr1-KO and Beta arr2-KO cells showed similar impairment in agonist-stimulated Beta(2)-AR and AT(1A)-R desensitization, when compared with their WT control cells, and the Beta arr1/2-KO cells were even further impaired. Sequestration of the Beta(2)-AR in the Beta arr2-KO cells was compromised significantly (87% reduction), whereas in the Beta arr1-KO cells it was not. Agonist-stimulated internalization of the AT(1A)-R was only slightly reduced in the Beta arr1-KO but was unaffected in the Beta arr2-KO cells. In the Beta arr1/2-KO cells, the sequestration of both receptors was dramatically reduced. Comparison of the ability of the two Beta-Arrestins to sequester the Beta(2)-AR revealed Beta-Arrestin 2 to be 100-fold more potent than Beta-Arrestin 1. Down-regulation of the Beta(2)-AR was also prevented in the Beta arr1/2-KO cells, whereas no change was observed in the single knockout cells. These findings suggest that sequestration of various heptahelical receptors is regulated differently by the two Beta-Arrestins, whereas both isoforms are capable of supporting receptor desensitization and down-regulation.