scholarly journals Cyclic AMP-Rap1A signaling activates RhoA to induce α2c-adrenoceptor translocation to the cell surface of microvascular smooth muscle cells

2012 ◽  
Vol 303 (5) ◽  
pp. C499-C511 ◽  
Author(s):  
Selvi C. Jeyaraj ◽  
Nicholas T. Unger ◽  
Ali H. Eid ◽  
Srabani Mitra ◽  
N. Paul El-Dahdah ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Cyclic Amp ◽  
Cell Surface ◽  
Smooth Muscle Cells ◽  
Intracellular Signaling ◽  
Muscle Cells ◽  
Small Gtpase ◽  
Hek293 Cells ◽  
Intracellular Camp

Intracellular signaling by the second messenger cyclic AMP (cAMP) activates the Ras-related small GTPase Rap1 through the guanine exchange factor Epac. This activation leads to effector protein interactions, activation, and biological responses in the vasculature, including vasorelaxation. In vascular smooth muscle cells derived from human dermal arterioles (microVSM), Rap1 selectively regulates expression of G protein-coupled α2C-adrenoceptors (α2C-ARs) through JNK-c-jun nuclear signaling. The α2C-ARs are generally retained in the trans-Golgi compartment and mobilize to the cell surface and elicit vasoconstriction in response to cellular stress. The present study used human microVSM to examine the role of Rap1 in receptor localization. Complementary approaches included murine microVSM derived from tail arteries of C57BL6 mice that express functional α2C-ARs and mice deficient in Rap1A (Rap1A-null). In human microVSM, increasing intracellular cAMP by direct activation of adenylyl cyclase by forskolin (10 μM) or selectively activating Epac-Rap signaling by the cAMP analog 8-pCPT-2′- O-Me-cAMP (100 μM) activated RhoA, increased α2C-AR expression, and reorganized the actin cytoskeleton, increasing F-actin. The α2C-ARs mobilized from the perinuclear region to intracellular filamentous structures and to the plasma membrane. Similar results were obtained in murine wild-type microVSM, coupling Rap1-Rho-actin dynamics to receptor relocalization. This signaling was impaired in Rap1A-null murine microVSM and was rescued by delivery of constitutively active (CA) mutant of Rap1A. When tested in heterologous HEK293 cells, Rap1A-CA or Rho-kinase (ROCK-CA) caused translocation of functional α2C-ARs to the cell surface (∼4- to 6-fold increase, respectively). Together, these studies support vascular bed-specific physiological role of Rap1 and suggest a role in vasoconstriction in microVSM.

scholarly journals Cyclic AMP-Rap1A signaling mediates cell surface translocation of microvascular smooth muscle α2C-adrenoceptors through the actin-binding protein filamin-2

2013 ◽  
Vol 305 (8) ◽  
pp. C829-C845 ◽  
Author(s):  
Hanaa K. B. Motawea ◽  
Selvi C. Jeyaraj ◽  
Ali H. Eid ◽  
Srabani Mitra ◽  
Nicholas T. Unger ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Plasma Membrane ◽  
Cyclic Amp ◽  
Cell Surface ◽  
Smooth Muscle Cells ◽  
Blood Vessels ◽  
Muscle Cells ◽  
Small Gtpase ◽  
Actin Binding ◽  
Surface Translocation

The second messenger cyclic AMP (cAMP) plays a vital role in vascular physiology, including vasodilation of large blood vessels. We recently demonstrated cAMP activation of Epac-Rap1A and RhoA-Rho-associated kinase (ROCK)-F-actin signaling in arteriolar-derived smooth muscle cells increases expression and cell surface translocation of functional α2C-adrenoceptors (α2C-ARs) that mediate vasoconstriction in small blood vessels (arterioles). The Ras-related small GTPAse Rap1A increased expression of α2C-ARs and also increased translocation of perinuclear α2C-ARs to intracellular F-actin and to the plasma membrane. This study examined the mechanism of translocation to better understand the role of these newly discovered mediators of blood flow control, potentially activated in peripheral vascular disorders. We utilized a yeast two-hybrid screen with human microvascular smooth muscle cells (microVSM) cDNA library and the α2C-AR COOH terminus to identify a novel interaction with the actin cross-linker filamin-2. Yeast α-galactosidase assays, site-directed mutagenesis, and coimmunoprecipitation experiments in heterologous human embryonic kidney (HEK) 293 cells and in human microVSM demonstrated that α2C-ARs, but not α2A-AR subtype, interacted with filamin. In Rap1-stimulated human microVSM, α2C-ARs colocalized with filamin on intracellular filaments and at the plasma membrane. Small interfering RNA-mediated knockdown of filamin-2 inhibited Rap1-induced redistribution of α2C-ARs to the cell surface and inhibited receptor function. The studies suggest that cAMP-Rap1-Rho-ROCK signaling facilitates receptor translocation and function via phosphorylation of filamin-2 Ser2113. Together, these studies extend our previous findings to show that functional rescue of α2C-ARs is mediated through Rap1-filamin signaling. Perturbation of this signaling pathway may lead to alterations in α2C-AR trafficking and physiological function.

scholarly journals Inhibition of proliferation, but not of Ca2+ mobilization, by cyclic AMP and GMP in rabbit aortic smooth-muscle cells

1992 ◽  
Vol 288 (2) ◽  
pp. 527-532 ◽  
Author(s):  
J W Assender ◽  
K M Southgate ◽  
M B Hallett ◽  
A C Newby
Keyword(s):  
Smooth Muscle ◽  
Cyclic Amp ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Bovine Serum ◽  
Muscle Cells ◽  
Intracellular Camp ◽  
Camp Concentration ◽  
Atp Concentration ◽  
Foetal Bovine Serum

The effects on cellular proliferation and Ca2+ mobilization of analogues of cyclic AMP (cAMP) and cyclic GMP (cGMP) and of agents that elevate the intracellular concentrations of cyclic nucleotides were compared in closely similar preparations of first-passage rabbit aortic vascular smooth-muscle cells. Proliferation induced by foetal-bovine serum was inhibited by 78% by 1 mM-8-bromo cAMP and by 42% by 1 mM-8-bromo cGMP. In the presence of 100 microM-isobutylmethylxanthine, 100 microM-forskolin increased intracellular cAMP concentration 5-fold and inhibited proliferation by 87%, but did not affect cGMP concentration or cell viability (ATP concentration). Similarly in the presence of 100 microM-isobutylmethylxanthine, 1 mM-SIN-1 (3-morpholinosydnonimine) elevated cGMP concentration 4-fold and inhibited proliferation by 48%, but did not affect cAMP or ATP concentration. Isobutylmethylxanthine (1 mM) elevated cAMP concentration by 3-fold and cGMP concentration by 20-fold and inhibited proliferation by 81%. Concentrations of 8-bromo cAMP, 8-bromo cGMP, forskolin or SIN-1 that inhibited proliferation did not affect the elevation of intracellular free Ca2+ concentration caused by 2% (v/v) foetal-bovine serum, 100 nM-5-hydroxytryptamine or 10 nM-angiotensin II. The results demonstrate that elevation of intracellular cAMP and cGMP concentrations both independently inhibit vascular smooth-muscle cell proliferation, but these effects on proliferation are not mediated by inhibition of Ca2+ mobilization.

The role of WNT/b-catenin signaling in smooth muscle cells during lung development and repair

Pneumologie ◽  
2014 ◽  
Vol 68 (06) ◽  
Author(s):  
A Moiseenko ◽  
E El Agha ◽  
B MacKenzie ◽  
S De Langhe ◽  
S Bellusci
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Lung Development ◽  
Muscle Cells
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