siRNA screen identifies the phosphatase acting on the G protein-coupled thyrotropin-releasing hormone receptor.
siRNA screen identifies the phosphatase acting on the G protein-coupled thyrotropin-releasing hormone receptor.
复制标题
siRNA 筛选可鉴定作用于 G 蛋白偶联促甲状腺素释放激素受体的磷酸酶。
DOI：
10.1021/cb3004513
复制
发表时间：
2013
影响因子：
4
通讯作者：
Hinkle,PatriciaM
中科院分区：
文献类型：
--
作者：
Gehret,AustinU;Hinkle,PatriciaM
G蛋白偶联受体（GPCR）是一类广泛表达的跨膜蛋白，参与神经递质、激素和各种其他配体的信号转导。它们的信号输出通过涉及磷酸化、内化和从G蛋白解离的机制而脱敏，并通过涉及去磷酸化的机制而重新敏化，但通常缺乏有关负责的磷酸酶的细节。我们在这里描述了使用基于siRNA的文库敲低特定磷酸酶亚基的表达，以鉴定蛋白磷酸酶1-α（PP 1 α）对促甲状腺激素释放激素（TRH）受体的重要性。在有利于去磷酸化的条件下，抑制PP 1 α合成和过表达显性负性PP 1 α可保护受体磷酸化，而过表达PP 1 α可加速去磷酸化。敲除所有三种PP 1催化亚基比单独敲除PP 1 α更有力地抑制TRH受体磷酸化，这表明不同的PP 1亚型功能冗余。敲除PP 2A的结构亚基（文库筛选中的第二个潜在命中）是无效的。Calyculin A是一种有效的PP 1家族磷酸酶抑制剂，强烈抑制垂体细胞中转染的TRH受体和内源性受体的去磷酸化，但对PP 2A家族磷酸酶具有选择性的fostriecin却没有。我们的结论是PP 1类磷酸酶是必不可少的TRH受体去磷酸化。
G protein-coupled receptors (GPCRs) are an ubiquitously expressed class of transmembrane proteins involved in the signal transduction of neurotransmitters, hormones and various other ligands. Their signaling output is desensitized by mechanisms involving phosphorylation, internalization, and dissociation from G proteins and resensitized by mechanisms involving dephosphorylation, but details about the phosphatases responsible are generally lacking. We describe here the use of an siRNA-based library to knock down expression of specific phosphatase subunits to identify protein phosphatase 1-α (PP1α) as important for the thyrotropin-releasing hormone (TRH) receptor. Inhibition of PP1α synthesis and overexpression of dominant negative PP1α preserved receptor phosphorylation under conditions favoring dephosphorylation, whereas overexpression of PP1α accelerated dephosphorylation. Knockdown of all three PP1 catalytic subunits inhibited TRH receptor phosphorylation much more powerfully than knockdown of PP1α alone, suggesting that different PP1 isoforms function redundantly. Knockdown of a structural subunit of PP2A, a second potential hit in the library screen, was ineffective. Calyculin A, a potent inhibitor of PP1 family phosphatases, strongly inhibited dephosphorylation of transfected TRH receptors and endogenous receptors in pituitary cells, but fostriecin, which is selective for PP2A family phosphatases, did not. We conclude that the PP1 class of phosphatases is essential for TRH receptor dephosphorylation.
登录
查看更多内容
I 组代谢型谷氨酸受体与蛋白磷酸酶 1C 结合
影响因子：
4.8
作者：
C. Croci;H. Sticht;J. Brandstätter;R. Enz
通讯作者：
R. Enz
Ser/Thr 磷酸酶的突变分析。
DOI：
--
发表时间：
1994
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Zhuo,S;Clemens,JC;Stone,RL;Dixon,JE
通讯作者：
Dixon,JE
影响因子：
--
作者：
M. Swingle;Li Ni;R. Honkanen
通讯作者：
M. Swingle;Li Ni;R. Honkanen
影响因子：
4.8
作者：
Poell, Florian;Doll, Christian;Schulz, Stefan
通讯作者：
Schulz, Stefan
影响因子：
13.8
作者：
Bollen, Mathieu;Peti, Wolfgang;Ragusa, Michael J.;Beullens, Monique
通讯作者：
Beullens, Monique