Agonist stimulation increases the turnover rate of beta(2)AR-bound palmitate and promotes receptor depalmitoylation
Agonist stimulation increases the turnover rate of beta(2)AR-bound palmitate and promotes receptor depalmitoylation
复制标题
DOI：
10.1021/bi9611321
复制
发表时间：
1996-12-10
期刊：
影响因子：
2.9
通讯作者：
Bouvier, M
中科院分区：
文献类型：
--
作者：
Loisel, TP;Adam, L;Bouvier, M
我们已经表征了 Sf9 细胞中 β(2)-肾上腺素受体棕榈酰化的动态性质。在基础条件下，与受体蛋白本身的周转率（半衰期 = 109 +/- 10 分钟）相比，受体结合的棕榈酸酯的周转速度很快（半衰期 = 9.8 +/- 1.8 分钟）。这表明在静息状态下受体的棕榈酰化和非棕榈酰化形式之间存在平衡。激动剂异丙肾上腺素对受体的刺激可将β(2)-肾上腺素能受体结合棕榈酸酯的半衰期缩短1.8倍，而不影响受体本身的周转率。在持续刺激后，这种增加的棕榈酸酯周转率将平衡转向受体的非棕榈酰化形式，这表明长时间的激活要么增加了去棕榈酰化的速率，要么阻止了受体棕榈酰化。与后一种可能性一致，在代谢标记之前用激动剂预处理细胞，将[H-3]棕榈酸酯掺入β(2)-肾上腺素受体的量减少了80%以上。这表明持续激动剂刺激时发生的受体脱敏与受体棕榈酰化的减少之间存在联系。支持这一假设的是，已知参与受体脱敏的 PKA 磷酸化位点的突变消除了激动剂促进的棕榈酸酯掺入的减少。我们之前曾报道过 β(2)-肾上腺素能受体的棕榈酰化对于通过 PKA 控制受体磷酸化非常重要 [Moffett, S., et al. 2017]。 (1993) EMBO J. 12, 349-356;莫菲特，S.，ct al。 (1996) J.Biol。陈省身。 271、21490-21497]。目前的研究表明，受体棕榈酰化状态受到激动剂刺激的调节，并表明在持续的受体刺激下，棕榈酰化和磷酸化之间存在协同的相互调节相互作用。
We have characterized the dynamic nature of beta(2)-adrenergic receptor palmitoylation in Sf9 cells. Under basal conditions, the turnover of receptor-bound palmitate is rapid (half-life = 9.8 +/- 1.8 min) compared to the turnover rate of the receptor protein itself (half-life = 109 +/- 10 min). This suggests that an equilibrium between the palmitoylated and nonpalmitoylated forms of the receptor exists at resting state. Stimulation of the receptor by the agonist isoproterenol reduces the half-life of the beta(2)-adrenergic receptor-bound palmitate by 1.8 fold without affecting the turnover rate of the receptor itself. Upon sustained stimulation, this increased palmitate turnover rate shifted the equilibrium toward the nonpalmitoylated form of the receptor, suggesting that prolonged activation either increases the rate of depalmitoylation or prevents receptor palmitoylation. Consistent with the latter possibility, pretreatment of cells with agonist, prior to metabolic labeling, reduced the incorporation of [H-3]palmitate into the beta(2)-adrenergic receptor by more than 80%. This suggests a link between receptor desensitization occurring upon sustained agonist stimulation and the decrease in receptor palmitoylation. Supporting this hypothesis, mutation of PKA phosphorylation sites known to be involved in receptor desensitization abolished the agonist-promoted reduction in palmitate incorporation. We have previously reported that palmitoylation of the beta(2)-adrenergic receptor is important in controlling receptor phosphorylation by PKA [Moffett, S., et al. (1993) EMBO J. 12, 349-356; Moffett, S., ct al. (1996) J. Biol. Chern. 271, 21490-21497]. The present study now demonstrates that the receptor palmitoylation state is regulated by agonist stimulation and suggests the existence of concerted reciprocal regulatory interactions between palmitoylation and phosphorylation upon sustained receptor stimulation.