MyRIP interaction with MyoVa on secretory granules is controlled by the cAMP-PKA pathway.
MyRIP interaction with MyoVa on secretory granules is controlled by the cAMP-PKA pathway.
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DOI：
10.1091/mbc.e12-05-0369
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发表时间：
2012-11
影响因子：
3.3
通讯作者：
Váradi A
中科院分区：
文献类型：
--
作者：
Brozzi F;Lajus S;Diraison F;Rajatileka S;Hayward K;Regazzi R;Molnár E;Váradi A
肌球蛋白和Rab-interacting蛋白不是MyoVa在大的致密核心分泌颗粒（SG）上的经典受体，但它有助于内分泌和神经内分泌细胞中SG上MyoVa相关蛋白的PKA依赖性磷酸化。肌球蛋白和Rab-interacting protein（MyRIP）属于蛋白激酶A（PKA）锚定家族，与激素分泌有关。然而，其作用机制尚未完全阐明。在这里，我们调查的作用MyRIP在肌球蛋白Va（MyoVa）依赖性分泌颗粒（SG）的运输和胰腺β细胞的分泌。这些细胞仅表达MyoVa的脑同种型（BR-MyoVa），其是SG转运中的关键运动蛋白。体外下拉，共免疫沉淀和共定位研究显示，MyRIP不与葡萄糖刺激的胰腺β细胞中的BR-MyoVa相互作用，这表明，与以前的概念相反，MyRIP不将这种运动蛋白与SG联系起来。葡萄糖刺激的胰岛素分泌通过肠促胰岛素激素增强，肠促胰岛素激素增加cAMP水平并导致MyRIP磷酸化、MyRIP与BR-MyoVa的相互作用以及BR-MyoVa受体rabphilin-3A（Rph-3A）的磷酸化。小干扰RNA敲低MyRIP可抑制Ser-234上的Rph-3A磷酸化，这也降低了cAMP介导的激素分泌。证明这种磷酸化的重要性，nonphosphorylatable和拟磷酸化Rph-3A突变体显着改变激素释放时，PKA被激活。这些数据表明，MyRIP仅在cAMP升高时在SG上与BR-MyoVa形成功能性蛋白复合物，并且在这种条件下促进SG相关蛋白的磷酸化，这反过来可以增强分泌。
Myosin- and Rab-interacting protein is not a classic receptor for MyoVa on large, dense-core secretory granules (SGs), but it aids in PKA-dependent phosphorylation of MyoVa-associated proteins on SGs in endocrine and neuroendocrine cells. Myosin- and Rab-interacting protein (MyRIP), which belongs to the protein kinase A (PKA)–anchoring family, is implicated in hormone secretion. However, its mechanism of action is not fully elucidated. Here we investigate the role of MyRIP in myosin Va (MyoVa)-dependent secretory granule (SG) transport and secretion in pancreatic beta cells. These cells solely express the brain isoform of MyoVa (BR-MyoVa), which is a key motor protein in SG transport. In vitro pull-down, coimmunoprecipitation, and colocalization studies revealed that MyRIP does not interact with BR-MyoVa in glucose-stimulated pancreatic beta cells, suggesting that, contrary to previous notions, MyRIP does not link this motor protein to SGs. Glucose-stimulated insulin secretion is augmented by incretin hormones, which increase cAMP levels and leads to MyRIP phosphorylation, its interaction with BR-MyoVa, and phosphorylation of the BR-MyoVa receptor rabphilin-3A (Rph-3A). Rph-3A phosphorylation on Ser-234 was inhibited by small interfering RNA knockdown of MyRIP, which also reduced cAMP-mediated hormone secretion. Demonstrating the importance of this phosphorylation, nonphosphorylatable and phosphomimic Rph-3A mutants significantly altered hormone release when PKA was activated. These data suggest that MyRIP only forms a functional protein complex with BR-MyoVa on SGs when cAMP is elevated and under this condition facilitates phosphorylation of SG-associated proteins, which in turn can enhance secretion.