Caveolae compartmentalise β2-adrenoceptor signals by curtailing cAMP production and maintaining phosphatase activity in the sarcoplasmic reticulum of the adult ventricular myocyte.
Caveolae compartmentalise β2-adrenoceptor signals by curtailing cAMP production and maintaining phosphatase activity in the sarcoplasmic reticulum of the adult ventricular myocyte.
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DOI：
10.1016/j.yjmcc.2011.06.014
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发表时间：
2012-02
影响因子：
5
通讯作者：
Calaghan S
中科院分区：
文献类型：
--
作者：
Macdougall DA;Agarwal SR;Stopford EA;Chu H;Collins JA;Longster AL;Colyer J;Harvey RD;Calaghan S
激活肾上腺素能受体(β1-，β2-，AR)可有效地诱导心肌细胞的正性和负性。β2-AR衍生的cAMP依赖信号的区隔是这种功能差异的基础。在这里，我们研究了小凹(富含胆固醇和小窝蛋白-3的特殊肌膜内陷)对成年大鼠心室肌细胞分室的作用机制。选择性地激活β2-AR(与Zinterol/CGP20712A一起)在对照细胞中几乎不产生收缩反应，而在胆固醇去除剂甲基-β-环糊精(MBCD)处理的细胞中却有明显的正性和向左收缩反应。这与L型钙电流的调节无关，而是与蛋白激酶A介导的Ser16位磷蛋白的磷酸化有关。在β2-AR刺激过程中，应用一种细胞通透性的小凹-3支架相互作用抑制剂，模拟了MBCD对磷酸化磷蛋白的影响，这与MBCD通过小窝发挥作用一致。生物传感器实验显示，只有在β处理后，cAMP 2-AR才能在完整细胞的PKA II信号域中动员cAMP，这提供了cAMP从小窝约束中解脱的实时演示。在对照组和MBCD处理组中，比较了磷酸二酯酶(PDE)、蛋白磷酸酶(PP)和磷脂酰肌醇-3-激酶(PI3K)抑制剂对pPLB和收缩的影响。仅PP抑制在显示对照细胞中β2-AR衍生信号的强健分解和胆固醇耗竭后相对较小的影响方面是显著的。综合所有证据，我们提出了一个新的概念，即小窝通过提供一个平台来限制β2-AR-cAMP信号传递，该平台不仅可以减少cAMP的产生，还可以防止PPS在肌浆网的抑制性调制。本文是题为“心肌细胞中的局部信号”的特刊的一部分。►我们研究了小凹将β2-AR信号区间化的机制(S)。►破坏小窝揭示了β2-AR cAMP信号位于PKA-RII区。►破坏小窝可促进pKA介导的原核蛋白选择性磷酸化。►磷酸酶(PP)1/2a(而不是PDE 2/3/4)参与了空泡β2-AR的区划。►我们认为完整的小窝减弱了cAMP的产生，并限制了PP对SR的抑制调制。
Inotropy and lusitropy in the ventricular myocyte can be efficiently induced by activation of β1-, but not β2-, adrenoceptors (ARs). Compartmentation of β2-AR-derived cAMP-dependent signalling underlies this functional discrepancy. Here we investigate the mechanism by which caveolae (specialised sarcolemmal invaginations rich in cholesterol and caveolin-3) contribute to compartmentation in the adult rat ventricular myocyte. Selective activation of β2-ARs (with zinterol/CGP20712A) produced little contractile response in control cells but pronounced inotropic and lusitropic responses in cells treated with the cholesterol-depleting agent methyl-β-cyclodextrin (MBCD). This was not linked to modulation of L-type Ca2+ current, but instead to a discrete PKA-mediated phosphorylation of phospholamban at Ser16. Application of a cell-permeable inhibitor of caveolin-3 scaffolding interactions mimicked the effect of MBCD on phosphorylated phospholamban (pPLB) during β2-AR stimulation, consistent with MBCD acting via caveolae. Biosensor experiments revealed β2-AR mobilisation of cAMP in PKA II signalling domains of intact cells only after MBCD treatment, providing a real-time demonstration of cAMP freed from caveolar constraint. Other proteins have roles in compartmentation, so the effects of phosphodiesterase (PDE), protein phosphatase (PP) and phosphoinositide-3-kinase (PI3K) inhibitors on pPLB and contraction were compared in control and MBCD treated cells. PP inhibition alone was conspicuous in showing robust de-compartmentation of β2-AR-derived signalling in control cells and a comparatively diminutive effect after cholesterol depletion. Collating all evidence, we promote the novel concept that caveolae limit β2-AR-cAMP signalling by providing a platform that not only attenuates production of cAMP but also prevents inhibitory modulation of PPs at the sarcoplasmic reticulum. This article is part of a Special Issue entitled “Local Signaling in Myocytes”. ► We investigate the mechanism(s) by which caveolae compartmentalise β2-AR signalling. ► Disrupting caveolae reveals β2-AR cAMP signals in the PKA-RII compartment. ► Disrupting caveolae promotes a selective PKA-mediated phosphorylation of PLB. ► Phosphatases (PP) 1/2a (but not PDE 2/3/4) contribute to caveolar β2-AR compartmentation. ► We propose that intact caveolae attenuate cAMP production and limit inhibitory modulation of PP at the SR.
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影响因子：
5
作者：
Calaghan, Sarah;Kozera, Lukasz;White, Ed
通讯作者：
White, Ed
影响因子：
20.1
作者：
Jo, SH;Leblais, V;Xiao, RP
通讯作者：
Xiao, RP
影响因子：
4.8
作者：
Couet, J;Li, SW;Lisanti, MP
通讯作者：
Lisanti, MP
DOI：
10.1073/pnas.0607294103
发表时间：
2006-10-31
影响因子：
11.1
作者：
Hulme, Joanne T.;Westenbroek, Ruth E.;Catterall, William A.
通讯作者：
Catterall, William A.
影响因子：
4.5
作者：
Calaghan, SC;White, E;Colyer, J
通讯作者：
Colyer, J