α1A-Adrenergic Receptor Induces Activation of Extracellular Signal-Regulated Kinase 1/2 through Endocytic Pathway

Liu, Fei; He, Kangmin; Yang, Xinxing; Xu, Ning; Liang, Zhangyi; Xu, Ming; Zhao, Xinsheng; Han, Qinghua; Zhang, Youyi

doi:10.1371/journal.pone.0021520

Cited by 14 publications

(13 citation statements)

References 48 publications

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“…A number of studies have used low experimental temperatures to block GPCR internalization (18,22). In contrast to prior work in which much reduced temperatures (ϳ4°C) were commonly used, the current studies show that reducing temperature to 24°C is sufficient to suppress PAC1 receptor internalization (31).…”

Section: Discussioncontrasting

confidence: 69%

PACAP-induced ERK activation in HEK cells expressing PAC1 receptors involves both receptor internalization and PKC signaling

May

Buttolph

Girard

et al. 2014

American Journal of Physiology-Cell Physiology

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The pituitary adenylate cyclase-activating polypeptide (PACAP)-selective PAC1 receptor (Adcyap1r1) is a G protein-coupled receptor (GPCR) that activates adenylyl cyclase and PLC. Similar to many other GPCRs, our previous studies showed that the PAC1 receptor is internalized after ligand binding to form signaling endosomes, which recruit additional second messenger pathways. Using a human embryonic kidney (HEK 293) PAC1Hop1-EGFP receptor cell line, we have examined how different PAC1 receptor signaling mechanisms contribute to MEK/ERK activation. Unlike PAC1 receptor-stimulated adenylyl cyclase/cAMP production in the plasma membrane, PACAP-mediated ERK phosphorylation was partly dependent on receptor internalization, as determined by treatment with pharmacological inhibitors of endocytosis or temperature reduction, which also suppressed receptor internalization. Stimulation of cAMP generation by forskolin or exposure to the cell-permeable cAMP analogs 8-bromo-cAMP and dibutyryl cAMP had minimal effects on ERK phosphorylation in this system. The ability of reduced temperature (24°C) to consistently suppress ERK activation to a greater extent than the endocytosis inhibitors Pitstop 2 and dynasore indicated that other mechanisms, in addition to PAC1 internalization/endosome activation, were involved. Inhibition of PAC1 receptor-stimulated PLC/diacylglycerol/PKC signaling by bisindoylmaleimide I also attenuated ERK phosphorylation, and direct PKC activation with phorbol ester increased ERK phosphorylation in a temperature-dependent manner. Inhibition of PAC1 receptor endocytosis and PKC activation completely blocked PACAP-stimulated ERK activation. PACAP augmented phosphorylated ERK staining uniformly over the cytoplasm and nucleus, and PKC signaling facilitated nuclear phosphorylated ERK translocation. In sum, our results show that PACAP/PAC1 receptor endocytosis and PLC/diacylglycerol/PKC activation represent two complementary mechanisms contributing to PACAP-induced ERK activation.

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Section: Discussioncontrasting

confidence: 69%

PACAP-induced ERK activation in HEK cells expressing PAC1 receptors involves both receptor internalization and PKC signaling

May

Buttolph

Girard

et al. 2014

American Journal of Physiology-Cell Physiology

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“…This later signal did not disappear over 15 min and we found high levels of pERK1/2 even when the α 1A localization in the deeper endosomes has reached its maximum (Figure 11). The existence of a remaining pERK1/2 signal which depends on endocytosis has previously been proposed for α 1A -ARs and other GPCRs [38], [39], [40], [41]. This late α 1A -mediated pERK1/2, observed after lipid raft disruption by mβCD or caveolae disorganization by filipin, but inhibited by Concanavalin A, corroborates the existence of an ERK1/2 signal independent of the membrane or caveolae integrity but dependent on receptor internalization.…”

Section: Discussionsupporting

confidence: 83%

Differences in the Signaling Pathways of α1A- and α1B-Adrenoceptors Are Related to Different Endosomal Targeting

et al. 2013

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AimsTo compare the constitutive and agonist-dependent endosomal trafficking of α1A- and α1B-adrenoceptors (ARs) and to establish if the internalization pattern determines the signaling pathways of each subtype.MethodsUsing CypHer5 technology and VSV-G epitope tagged α1A- and α1B-ARs stably and transiently expressed in HEK 293 cells, we analyzed by confocal microscopy the constitutive and agonist-induced internalization of each subtype, and the temporal relationship between agonist induced internalization and the increase in intracellular calcium (determined by FLUO-3 flouorescence), or the phosphorylation of ERK1/2 and p38 MAP kinases (determined by Western blot).Results and ConclusionsConstitutive as well as agonist-induced trafficking of α1A and α1B ARs maintain two different endosomal pools of receptors: one located close to the plasma membrane and the other deeper into the cytosol. Each subtype exhibited specific characteristics of internalization and distribution between these pools that determines their signaling pathways: α1A-ARs, when located in the plasma membrane, signal through calcium and ERK1/2 pathways but, when translocated to deeper endosomes, through a mechanism sensitive to β-arrestin and concanavalin A, continue signaling through ERK1/2 and also activate the p38 pathway. α1B-ARs signal through calcium and ERK1/2 only when located in the membrane and the signals disappear after endocytosis and by disruption of the membrane lipid rafts by methyl-β-cyclodextrin

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“…The same pathway is also activated by α‐AR stimulation. Direct activation of Erk1/2 by α‐ARs was reported in several studies . α‐ARs may influence many processes involved in locomotion, pain, and spasms via the Ras‐Raf‐MEK‐ERK signal transduction cascade, and also by other mechanisms.…”

Section: Introduction To the α‐Adrenoceptors: Expression And Multifunmentioning

confidence: 89%

Alpha-Adrenoceptor Modulation in Central Nervous System Trauma: Pain, Spasms, and Paralysis - An Unlucky Triad

et al. 2014

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Many researchers have attempted to pharmacologically modulate the adrenergic system to control locomotion, pain, and spasms after central nervous system (CNS) trauma, although such efforts have led to conflicting results. Despite this, multiple studies highlight that α-adrenoceptors (α-ARs) are promising therapeutic targets because in the CNS, they are involved in reactivity to stressors and regulation of locomotion, pain, and spasms. These functions can be activated by direct modulation of these receptors on neuronal networks in the brain and the spinal cord. In addition, these multifunctional receptors are also broadly expressed on immune cells. This suggests that they might play a key role in modulating immunological responses, which may be crucial in treating spinal cord injury and traumatic brain injury as both diseases are characterized by a strong inflammatory component. Reducing the proinflammatory response will create a more permissive environment for axon regeneration and may support neuromodulation in combination therapies. However, pharmacological interventions are hindered by adrenergic system complexity and the even more complicated anatomical and physiological changes in the CNS after trauma. This review is the first concise overview of the pros and cons of α-AR modulation in the context of CNS trauma.

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α1A-Adrenergic Receptor Induces Activation of Extracellular Signal-Regulated Kinase 1/2 through Endocytic Pathway

Cited by 14 publications

References 48 publications

PACAP-induced ERK activation in HEK cells expressing PAC1 receptors involves both receptor internalization and PKC signaling

PACAP-induced ERK activation in HEK cells expressing PAC1 receptors involves both receptor internalization and PKC signaling

Differences in the Signaling Pathways of α1A- and α1B-Adrenoceptors Are Related to Different Endosomal Targeting

Alpha-Adrenoceptor Modulation in Central Nervous System Trauma: Pain, Spasms, and Paralysis - An Unlucky Triad

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