Persistent signaling by thyrotropin‐releasing hormone receptors correlates with G‐protein and receptor levels

Boutin, Alisa; Allen, Michael D.; Neumann, Susanne; Gershengorn, Marvin C.

doi:10.1096/fj.12-207860

Cited by 10 publications

(6 citation statements)

References 37 publications

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“…Similarly, PVNGq/11KO mice also showed hyper phagia at 12 to 16 weeks of age (Supplemental Figure 2, B and C), at a time when obesity was established (Supplemental Figure 2A), while energy expenditure As PLC is the major effector of G q α and G 11 α, we examined the effect of knocking out these G proteins on PLC signaling in the PVN. PVNs from control and PVNGq/11KO mice were isolated and incubated with either PBS or MTII, and accumulation of inositol monophosphate (IP 1 ), a metabolite of inositol triphosphate (IP 3 ), was examined in the presence of LiCl, which inhibits the metabolism of IP 1 (19). Baseline IP 1 levels (after PBS administration) in the PVNs from PVNGq/11KO mice were only 21% of those present in controls ( Figure 1C), consistent with G q α and G 11 α being lost in the PVN.…”

Section: Resultsmentioning

confidence: 99%

Gq/11α and Gsα mediate distinct physiological responses to central melanocortins

Li¹,

Shrestha²,

Pandey³

et al. 2015

Journal of Clinical Investigation

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Activation of brain melanocortin 4 receptors (MC4Rs) leads to reduced food intake, increased energy expenditure, increased insulin sensitivity, and reduced linear growth. MC4R effects on energy expenditure and glucose metabolism are primarily mediated by the G protein G(s)α in brain regions outside of the paraventricular nucleus of the hypothalamus (PVN). However, the G protein(s) that is involved in MC4R-mediated suppression of food intake and linear growth, which are believed to be regulated primarily though action in the PVN, is unknown. Here, we show that PVN-specific loss of G(q)α and G11α, which stimulate PLC, leads to severe hyperphagic obesity, increased linear growth, and inactivation of the hypothalamic-pituitary-adrenal axis, without affecting energy expenditure or glucose metabolism. Moreover, we demonstrate that the ability of an MC4R agonist delivered to PVN to inhibit food intake is lost in mice lacking G(q/11)α in the PVN but not in animals deficient for G(s)α. The blood pressure response to the same MC4R agonist was only lost in animals lacking G(s)α specifically in the PVN. Together, our results exemplify how different physiological effects of GPCRs may be mediated by different G proteins and identify a pathway for appetite regulation that could be selectively targeted by G(q/11)α-biased MC4R agonists as a potential treatment for obesity.

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Section: Resultsmentioning

confidence: 99%

Gq/11α and Gsα mediate distinct physiological responses to central melanocortins

Li¹,

Shrestha²,

Pandey³

et al. 2015

Journal of Clinical Investigation

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“…Together these results confirm the importance of arrestin for both desensitization and internalization and prove that internalization is not the cause of desensitization. In highly sensitive assays, persistent IP1 production can be demonstrated for up to an hour after TRH is washed out in HEK293 cells expressing TRH receptors (Boutin et al, 2012). This sustained response does not depend on internalization.…”

Section: Role Of Arrestin In Trh Receptor Signaling Desensitization mentioning

confidence: 99%

Desensitization, Trafficking, and Resensitization of the Pituitary Thyrotropin-Releasing Hormone Receptor

Hinkle¹,

Gehret²,

Jones³

2012

Front. Neurosci.

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The pituitary receptor for thyrotropin-releasing hormone (TRH) is a calcium-mobilizing G protein-coupled receptor (GPCR) that signals through Gq/11, elevating calcium, and activating protein kinase C. TRH receptor signaling is quickly desensitized as a consequence of receptor phosphorylation, arrestin binding, and internalization. Following activation, TRH receptors are phosphorylated at multiple Ser/Thr residues in the cytoplasmic tail. Phosphorylation catalyzed by GPCR kinase 2 (GRK2) takes place rapidly, reaching a maximum within seconds. Arrestins bind to two phosphorylated regions, but only arrestin bound to the proximal region causes desensitization and internalization. Phosphorylation at Thr365 is critical for these responses. TRH receptors internalize in clathrin-coated vesicles with bound arrestin. Following endocytosis, vesicles containing phosphorylated TRH receptors soon merge with rab5-positive vesicles. Over approximately 20 min these form larger endosomes rich in rab4 and rab5, early sorting endosomes. After TRH is removed from the medium, dephosphorylated receptors start to accumulate in rab4-positive, rab5-negative recycling endosomes. The mechanisms responsible for sorting dephosphorylated receptors to recycling endosomes are unknown. TRH receptors from internal pools help repopulate the plasma membrane. Dephosphorylation of TRH receptors begins when TRH is removed from the medium regardless of receptor localization, although dephosphorylation is fastest when the receptor is on the plasma membrane. Protein phosphatase 1 is involved in dephosphorylation but the details of how the enzyme is targeted to the receptor remain obscure. It is likely that future studies will identify biased ligands for the TRH receptor, novel arrestin-dependent signaling pathways, mechanisms responsible for targeting kinases and phosphatases to the receptor, and principles governing receptor trafficking.

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“…Activation of many GPCRs can lead to β-arrestin binding and sequestration of the receptor from the cell surface. The longstanding view that internalization of a GPCR leads to termination of receptor-mediated signalling has recently been challenged with reports demonstrating that, in addition to the role of β-arrestin in scaffolding activation of ERK (Lefkowitz and Shenoy, 2005 ), some receptors can still signal to the cAMP pathway even after receptor internalization (Mullershausen et al ., 2009 ; Boutin et al ., 2012 ; Werthmann et al ., 2012 ). In contrast to its role in G-protein-mediated signalling, the role of the TM6 toggle switch on agonist-mediated receptor internalization has not been widely explored.…”

Section: Introductionmentioning

confidence: 99%

Effect of a toggle switch mutation in TM6 of the human adenosine A₃ receptor on Gi protein‐dependent signalling and Gi‐independent receptor internalization

Stoddart

Kellam

Briddon

et al. 2014

British J Pharmacology

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Background and PurposeThe highly conserved tryptophan (W6.48) in transmembrane domain 6 of GPCRs has been shown to play a central role in forming an active conformation in response to agonist binding. We set out to characterize the effect of this mutation on the efficacy of two agonists at multiple signalling pathways downstream of the adenosine A3 receptor.Experimental ApproachResidue W6.48 in the human adenosine A3 receptor fused to yellow fluorescent protein was mutated to phenylalanine and expressed in CHO-K1 cells containing a cAMP response element reporter gene. The effects on agonist-mediated receptor internalization were monitored by automated confocal microscopy and image analysis. Further experiments were carried out to investigate agonist-mediated ERK1/2 phosphorylation, inhibition of [3H]-cAMP accumulation and β-arrestin2 binding.Key ResultsNECA was able to stimulate agonist-mediated internalization of the W6.48F mutant receptor, while the agonist HEMADO was inactive. Investigation of other downstream signalling pathways indicated that G-protein coupling was impaired for both agonists tested. Mutation of W6.48F therefore resulted in differential effects on agonist efficacy, and introduced signalling pathway bias for HEMADO at the adenosine A3 receptor.Conclusions and ImplicationsInvestigation of the pharmacology of the W6.48F mutant of the adenosine A3 receptor confirms that this region is important in forming the active conformation of the receptor for stimulating a number of different signalling pathways and that mutations in this residue can lead to changes in agonist efficacy and signalling bias.

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Persistent signaling by thyrotropin‐releasing hormone receptors correlates with G‐protein and receptor levels

Cited by 10 publications

References 37 publications

Gq/11α and Gsα mediate distinct physiological responses to central melanocortins

Gq/11α and Gsα mediate distinct physiological responses to central melanocortins

Desensitization, Trafficking, and Resensitization of the Pituitary Thyrotropin-Releasing Hormone Receptor

Effect of a toggle switch mutation in TM6 of the human adenosine A₃ receptor on Gi protein‐dependent signalling and Gi‐independent receptor internalization

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Persistent signaling by thyrotropin‐releasing hormone receptors correlates with G‐protein and receptor levels

Cited by 10 publications

References 37 publications

Gq/11α and Gsα mediate distinct physiological responses to central melanocortins

Gq/11α and Gsα mediate distinct physiological responses to central melanocortins

Desensitization, Trafficking, and Resensitization of the Pituitary Thyrotropin-Releasing Hormone Receptor

Effect of a toggle switch mutation in TM6 of the human adenosine A3 receptor on Gi protein‐dependent signalling and Gi‐independent receptor internalization

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Effect of a toggle switch mutation in TM6 of the human adenosine A₃ receptor on Gi protein‐dependent signalling and Gi‐independent receptor internalization