Multisite phosphorylation is required for sustained interaction with GRKs and arrestins during rapid μ-opioid receptor desensitization
Multisite phosphorylation is required for sustained interaction with GRKs and arrestins during rapid μ-opioid receptor desensitization
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DOI：
10.1126/scisignal.aas9609
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发表时间：
2018-07-17
影响因子：
7.3
通讯作者：
Canals, Meritxell
中科院分区：
文献类型：
--
作者：
Miess, Elke;Gondin, Arisbel B.;Canals, Meritxell
G蛋白受体激酶（GRKs）和β-抑制蛋白是μ-阿片受体（莫尔）信号传导和运输的关键调节剂。我们以前已经表明，高效阿片类药物，如DAMGO刺激GRK 2/3介导的保守的C-末端尾丝氨酸和苏氨酸残基的多位点磷酸化，这有利于受体的内化。相反，吗啡诱导的莫尔磷酸化仅限于Ser（375），不足以驱动大量受体内化。我们报告了特定的多位点磷酸化是如何控制GRK和β-抑制蛋白与莫尔相互作用的动力学的，并展示了这种磷酸化是如何介导受体脱敏的。我们发现GRK 2/3比β-arrestin更快地被募集到DAMGO激活的莫尔中。β-Arrestin的募集需要GRK 2活性和莫尔磷酸化，但GRK的募集也依赖于C-末端尾部的磷酸化位点，特别是（370）TREHPSTANT（379）基序内的四个丝氨酸和苏氨酸残基。我们的研究结果还表明，该基序以外的其他残基参与了GRK和β-arrestins的初始和瞬时募集。我们确定了莫尔的高效激动剂脱敏的两个组成部分：一个持续的组成部分，这需要GRK 2介导的磷酸化和潜在的可溶性因子，和一个快速的组成部分，这可能是由GRK 2介导的，但独立于受体磷酸化。阐明这些复杂的受体-效应物相互作用代表了对导致耐受性和依赖性发展的莫尔脱敏机制的理解的重要一步。
G protein receptor kinases (GRKs) and beta-arrestins are key regulators of mu-opioid receptor (MOR) signaling and trafficking. We have previously shown that high-efficacy opioids such as DAMGO stimulate a GRK2/3-mediated multisite phosphorylation of conserved C-terminal tail serine and threonine residues, which facilitates internalization of the receptor. In contrast, morphine-induced phosphorylation of MOR is limited to Ser(375) and is not sufficient to drive substantial receptor internalization. We report how specific multisite phosphorylation controlled the dynamics of GRK and beta-arrestin interactions with MOR and show how such phosphorylation mediated receptor desensitization. We showed that GRK2/3 was recruited more quickly than was beta-arrestin to a DAMGO-activated MOR. beta-Arrestin recruitment required GRK2 activity and MOR phosphorylation, but GRK recruitment also depended on the phosphorylation sites in the C-terminal tail, specifically four serine and threonine residues within the (370)TREHPSTANT(379) motif. Our results also suggested that other residues outside this motif participated in the initial and transient recruitment of GRK and beta-arrestins. We identified two components of high-efficacy agonist desensitization of MOR: a sustained component, which required GRK2-mediated phosphorylation and a potential soluble factor, and a rapid component, which was likely mediated by GRK2 but independent of receptor phosphorylation. Elucidating these complex receptor-effector interactions represents an important step toward a mechanistic understanding of MOR desensitization that leads to the development of tolerance and dependence.