New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D(2) receptor regulation and signaling.
New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D(2) receptor regulation and signaling.
复制标题
新的磷酸位点特异性抗体，以阐明GRK磷酸化在多巴胺D（2）受体调节和信号传导中的作用。
DOI：
10.1038/s41598-021-87417-2
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发表时间：
2021-04-15
影响因子：
4.6
通讯作者：
Robert Lane J
中科院分区：
文献类型：
--
作者：
Mann A;Keen AC;Mark H;Dasgupta P;Javitch JA;Canals M;Schulz S;Robert Lane J
多巴胺D2受体（D2 R）是用于治疗帕金森病和精神分裂症症状的药物的靶点。D2 R通过其与G蛋白受体激酶（GRKs）的相互作用和被其磷酸化以及与抑制蛋白的相互作用来调节。最近，D2 R抑制蛋白介导的信号传导已被证明具有与G蛋白信号传导不同的生理功能。关于可能控制这些过程的D2 R磷酸化模式，我们知之甚少。我们的目的是产生细胞内D2 R磷酸化位点的特异性抗体，以促进这些机制的调查。我们合成了对应于hD2 R的细胞内环3内区域的双磷酸化肽，并使用它们来产生磷酸位点特异性抗体以捕获GRK介导的磷酸化的广泛筛选。我们鉴定了对D2 R细胞内环3中GRK 2/3磷酸化位点特异的抗体。我们将D2 R磷酸化的测量与D2 R信号传导的其他测量进行比较，以分析所选的D2 R激动剂，包括先前描述的偏向性激动剂。这些研究证明了新型磷酸位点特异性抗体在研究D2 R调节和信号传导方面的实用性。
The dopamine D2 receptor (D2R) is the target of drugs used to treat the symptoms of Parkinson’s disease and schizophrenia. The D2R is regulated through its interaction with and phosphorylation by G protein receptor kinases (GRKs) and interaction with arrestins. More recently, D2R arrestin-mediated signaling has been shown to have distinct physiological functions to those of G protein signalling. Relatively little is known regarding the patterns of D2R phosphorylation that might control these processes. We aimed to generate antibodies specific for intracellular D2R phosphorylation sites to facilitate the investigation of these mechanisms. We synthesised double phosphorylated peptides corresponding to regions within intracellular loop 3 of the hD2R and used them to raise phosphosite-specific antibodies to capture a broad screen of GRK-mediated phosphorylation. We identify an antibody specific to a GRK2/3 phosphorylation site in intracellular loop 3 of the D2R. We compared measurements of D2R phosphorylation with other measurements of D2R signalling to profile selected D2R agonists including previously described biased agonists. These studies demonstrate the utility of novel phosphosite-specific antibodies to investigate D2R regulation and signalling.
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影响因子：
3.6
作者：
Free, R. Benjamin;Chun, Lani S.;Sibley, David R.
通讯作者：
Sibley, David R.
DOI：
10.1073/pnas.83.9.2797
发表时间：
1986-05-01
影响因子：
11.1
作者：
BENOVIC, JL;STRASSER, RH;LEFKOWITZ, RJ
通讯作者：
LEFKOWITZ, RJ
影响因子：
4.8
作者：
Hollins B;Kuravi S;Digby GJ;Lambert NA
通讯作者：
Lambert NA
影响因子：
3.7
作者：
Chun, Lani S.;Vekariya, Rakesh H.;Sibley, David R.
通讯作者：
Sibley, David R.
影响因子：
16.6
作者：
Klein Herenbrink C;Sykes DA;Donthamsetti P;Canals M;Coudrat T;Shonberg J;Scammells PJ;Capuano B;Sexton PM;Charlton SJ;Javitch JA;Christopoulos A;Lane JR
通讯作者：
Lane JR