Serine-727 Phosphorylation Activates Hypothalamic STAT-3 Independently From Tyrosine-705 Phosphorylation

Breit, Andreas; Besik, Valeria; Solinski, Hans Jürgen; Muehlich, Susanne; Glas, Evi; Yarwood, Stephen J.; Gudermann, Thomas

doi:10.1210/me.2014-1300

Cited by 21 publications

(34 citation statements)

References 78 publications

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“…mHypoA-2/10 cells express a wide range of hypothalamic markers and, as with to PVN neurons, have been shown to respond to MSH and NPY (Belsham et al, 2009;Dalvi et al, 2012;Nazarians-Armavil et al, 2013;Breit et al, 2015). In line with these previous reports, mHypoA-2/10 cells used herein strongly reacted to MSH with cAMP accumulation, indicating expression of G s -coupled MSH receptors (Fig.…”

Section: Discussionsupporting

confidence: 89%

“…In this work we took advantage of the recently introduced mHypoA-2/10 cell line that has been shown to be sensitive toward MSH, NPY, and T3 hormone and thus resembles 5-HT 1B R-and 5-HT 2C R-expressing neurons of the hypothalamic PVN (Belsham et al, 2009;Dalvi et al, 2012;NazariansArmavil et al, 2013;Breit et al, 2015). We provide compelling pharmacological evidence on the basis of selective 5-HTR antagonists and agonists that mHypoA-2/10 cells express functional 5-HT 1B R and, of particular interest, 5-HT 2C R. Thus mHypoA-2/10 cells represent the first CNS-derived cell model known so far that allows investigation of 5-HT 2C R biology in its natural environment.…”

Section: Discussionmentioning

confidence: 99%

“…To obtain mHypoA-2/10 cells, hypothalamic neurons from 2-month-old male C57Bl/6 mice were isolated, ciliary neurotrophic factor was used to trigger neurogenesis of hypothalamic primary cultures, and immortalization was induced by retroviral transfer of SV40 T-Ag (Belsham et al, 2009). To obtain mHypoA-2/10 cells stably expressing the CRE reporter, cells were cotransfected with the CREreporter construct and an empty pcDNA4 vector containing the resistance gene for zeocin (Breit et al, 2015). Transfected cells were then selected with 600 mg/ml zeocin for 4 weeks and positively tested cell pools were used for further studies.…”

Section: Cell Culture and Transfectionsmentioning

confidence: 99%

“…The hypothalamic cell line mHypoA-2/10 has recently been introduced and characterized as a suitable PVN-derived cell model with regard to sensitivity toward MSH and NPY, but its sensitivity toward 5-HT has not been investigated yet (Belsham et al, 2009;Dalvi et al, 2012;Nazarians-Armavil et al, 2013;Breit et al, 2015). In the present study we dissected the effects of 5-HT, WAY-161,503, and CP-94,253 on classic G protein-signaling cascades that involve cAMP, calcium, extracellular-regulated kinases-1/2 (ERK-1/2), and CRE binding protein (CREB)/cAMP response element (CRE).…”

Section: Introductionmentioning

confidence: 99%

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Endogenous 5-HT2C Receptors Phosphorylate the cAMP Response Element Binding Protein via Protein Kinase C-Promoted Activation of Extracellular-Regulated Kinases-1/2 in Hypothalamic mHypoA-2/10 Cells

Lauffer

Glas

Gudermann

et al. 2016

Journal of Pharmacology and Experimental Therapeutics

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Serotonin 5-HT 2C receptors (5-HT 2C R) activate G q proteins and are expressed in the central nervous system (CNS). 5-HT 2C R regulate emotion, feeding, reward, or cognition and may serve as promising drug targets to treat psychiatric disorders or obesity. Owing to technical difficulties in isolating cells from the CNS and the lack of suitable cell lines endogenously expressing 5-HT 2C R, our knowledge about this receptor subtype in native environments is rather limited. The hypothalamic mHypoA-2/10 cell line was recently established and resembles appetite-regulating hypothalamic neurons of the paraventricular nucleus (PVN), where 5-HT 2C R have been detected in vivo. Therefore, we tested mHypoA-2/10 cells for endogenous 5-HT 2C R expression. Serotonin or the 5-HT 2C R preferential agonist WAY-161,503 initiated cAMP response element (CRE)-dependent gene transcription with EC 50 values of 15.5 6 9.8 and 1.1 6 0.9 nM, respectively. Both responses were blocked by two unrelated 5-HT 2C R-selective antagonists (SB-242,084, RS-102,221) but not by a 5-HT 2A R (EMD-281,014) or 5-HT 2B R (RS-127,455) antagonists. By singlecell calcium imaging, we found that serotonin and WAY-161,503 induced robust calcium transients, which were also blunted by both 5-HT 2C R antagonists. Additionally we revealed, first, that 5-HT 2C R induced CRE activation via protein kinase C (PKC)-mediated engagement of extracellular-regulated kinases-1/2 and, second, that intrinsic activity of WAY-161,503 was in the range of 0.3-0.5 compared with serotonin, defining the frequently used 5-HT 2C R agonist as a partial agonist of endogenous 5-HT 2C R. In conclusion, we have shown that hypothalamic mHypoA-2/10 cells endogenously express 5-HT 2C R and thus are the first cell line in which to analyze 5-HT 2C R pharmacology, signaling, and regulation in its natural environment.

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Section: Discussionsupporting

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Section: Cell Culture and Transfectionsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Endogenous 5-HT2C Receptors Phosphorylate the cAMP Response Element Binding Protein via Protein Kinase C-Promoted Activation of Extracellular-Regulated Kinases-1/2 in Hypothalamic mHypoA-2/10 Cells

Lauffer

Glas

Gudermann

et al. 2016

Journal of Pharmacology and Experimental Therapeutics

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“…In the present study we investigated mHypoA-2/10-CRE cells that express a wide range of hypothalamic markers and resemble PVN neurons with regard to melanocortin sensitivity and TRH production and in addition, stable express a CRE-dependent reporter construct (27)(28)(29)(30)(31)(32)(33). We dissected effects of glucose on basal and MSHinduced CRE activation in terms of kinetics, affinity and desensitization.…”

mentioning

confidence: 99%

Glucose Enhances Basal or Melanocortin-Induced cAMP-Response Element Activity in Hypothalamic Cells

Breit

Wicht

Boekhoff

et al. 2016

Molecular Endocrinology

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Melanocyte-stimulating hormone (MSH)-induced activation of the cAMP-response element (CRE) via the CRE-binding protein in hypothalamic cells promotes expression of TRH and thereby restricts food intake and increases energy expenditure. Glucose also induces central anorexigenic effects by acting on hypothalamic neurons, but the underlying mechanisms are not completely understood. It has been proposed that glucose activates the CRE-binding protein-regulated transcriptional coactivator 2 (CRTC-2) in hypothalamic neurons by inhibition of AMP-activated protein kinases (AMPKs), but whether glucose directly affects hypothalamic CRE activity has not yet been shown. Hence, we dissected effects of glucose on basal and MSH-induced CRE activation in terms of kinetics, affinity, and desensitization in murine, hypothalamic mHypoA-2/10-CRE cells that stably express a CRE-dependent reporter gene construct. Physiologically relevant increases in extracellular glucose enhanced basal or MSH-induced CRE-dependent gene transcription, whereas prolonged elevated glucose concentrations reduced the sensitivity of mHypoA-2/10-CRE cells towards glucose. Glucose also induced CRCT-2 translocation into the nucleus and the AMPK activator metformin decreased basal and glucose-induced CRE activity, suggesting a role for AMPK/CRTC-2 in glucose-induced CRE activation. Accordingly, small interfering RNA-induced down-regulation of CRTC-2 expression decreased glucose-induced CRE-dependent reporter activation. Of note, glucose also induced expression of TRH, suggesting that glucose might affect the hypothalamic-pituitary-thyroid axis via the regulation of hypothalamic CRE activity. These findings significantly advance our knowledge about the impact of glucose on hypothalamic signaling and suggest that TRH release might account for the central anorexigenic effects of glucose and could represent a new molecular link between hyperglycaemia and thyroid dysfunction.

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Pathological cyclic strain promotes proliferation of vascular smooth muscle cells via the ACTH/ERK/STAT3 pathway

Tang

Liu

Xiao

et al. 2018

J of Cellular Biochemistry

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Abnormal proliferation of vascular smooth muscle cells (VSMCs) is closely related to hyperplasia in hypertension. Our previous study suggested that adrenocorticotropic hormone (ACTH) is mechano-responsive and may regulate VSMC proliferation. However, the molecular mechanism of VSMC abnormal proliferation induced by conditions of high cyclic strain, especially the role of ACTH in this process, is unclear. Our results revealed that ACTH and its specific receptor melanocortin receptor type 2 (MC2R) were highly expressed in hypertensive rat models. Furthermore, it was demonstrated that the expression of ACTH and MC2R was up-regulated when exposed to high cyclic strain in vitro, accompanied by abnormal proliferation of VSMCs. Next, it was proved that ACTH-dependent cell proliferation was related to the phosphorylation of extracellular regulated protein kinases (ERK) and signal transducer and activator of transcription 3 (STAT3). The study also found that ACTH could promote dimerization and glycosylation of melanocortin 2 receptor accessory protein (MRAP), which had a significant effect on MC2R membrane localization and signal activation. When VSMCs were treated with PD98059, a mitogen-activated protein kinase (MAP kinase) cascade antagonist, it was determined that phosphorylation of STAT3 at Ser727 was dependent on ERK phosphorylation. In summary, these data demonstrated that the abnormal proliferation of VSMCs induced by conditions of high cyclic strain is in part attributed to ACTH and its receptor MC2R. Identifying the mechanism of ACTH-dependent proliferation of VSMCs may help to provide new therapeutic targets for hypertension.

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Serine-727 Phosphorylation Activates Hypothalamic STAT-3 Independently From Tyrosine-705 Phosphorylation

Cited by 21 publications

References 78 publications

Endogenous 5-HT2C Receptors Phosphorylate the cAMP Response Element Binding Protein via Protein Kinase C-Promoted Activation of Extracellular-Regulated Kinases-1/2 in Hypothalamic mHypoA-2/10 Cells

Endogenous 5-HT2C Receptors Phosphorylate the cAMP Response Element Binding Protein via Protein Kinase C-Promoted Activation of Extracellular-Regulated Kinases-1/2 in Hypothalamic mHypoA-2/10 Cells

Glucose Enhances Basal or Melanocortin-Induced cAMP-Response Element Activity in Hypothalamic Cells

Pathological cyclic strain promotes proliferation of vascular smooth muscle cells via the ACTH/ERK/STAT3 pathway

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