Insulin Induces Heterologous Desensitization of G Protein-Coupled Receptor and Insulin-Like Growth Factor I Signaling by Downregulating β-Arrestin-1

β-Arrestin-1 mediates agonist-dependent desensitization and internalization of G protein-coupled receptors (GPCRs) and is also essential for GPCR mitogenic signaling. In addition, insulin-like growth factor I receptor (IGF-IR) endocytosis is facilitated by β-arrestin-1, and internalization is necessary for IGF-I-stimulated mitogen-activated protein (MAP) kinase activation. Here, we report that treatment of cells for 12 h with insulin (100 ng/ml) induces an ∼50% decrease in cellular β-arrestin-1 content due to ubiquitination of β-arrestin-1 and proteosome-mediated degradation. This insulin-induced decrease in β-arrestin-1 content was blocked by inhibition of phosphatidylinositol-3 kinase (PI-3 kinase) and MEK with wortmannin and PD98059, respectively. We also found a marked decrease in the association of β-arrestin-1 with the IGF-IR and a 55% inhibition of IGF-I-stimulated MAP kinase phosphorylation. In insulin-treated, β-arrestin-1-downregulated cells, there was complete inhibition of lysophosphatidic acid (LPA) or isoproterenol (ISO)-stimulated MAP kinase phosphorylation. This was associated with a decrease in β-arrestin-1 association with the β(2)-AR as well as a decrease in β-arrestin-1-Src and Src-β(2)-AR association. Ectopic expression of wild-type β-arrestin-1 in insulin-treated cells in which endogenous β-arrestin-1 had been downregulated rescued IGF-I- and LPA-stimulated MAP kinase phosphorylation. In conclusion, we found the following. (i) Chronic insulin treatment leads to enhanced β-arrestin-1 degradation. (ii) This downregulation of endogenous β-arrestin-1 is associated with decreased IGF-I-, LPA-, and ISO-mediated MAP kinase signaling, which can be rescued by ectopic expression of wild-type β-arrestin-1. (iii) Finally, these results describe a novel mechanism for heterologous desensitization, whereby insulin treatment can impair GPCR signaling, and highlight the importance of β-arrestin-1 as a target molecule for this desensitization mechanism.

β抑制蛋白1（β-Arrestin-1）可介导G蛋白偶联受体（G protein-coupled receptors, GPCRs）的激动剂依赖性脱敏与内化过程，同时对GPCR的促有丝分裂信号转导至关重要。此外，β抑制蛋白1可促进胰岛素样生长因子I受体（insulin-like growth factor I receptor, IGF-IR）的内吞作用，而该内化过程对于胰岛素样生长因子I（IGF-I）刺激下的丝裂原活化蛋白（mitogen-activated protein, MAP）激酶激活不可或缺。本研究报道，以100 ng/ml浓度的胰岛素处理细胞12小时后，细胞内β抑制蛋白1的含量会下降约50%，该现象源于β抑制蛋白1的泛素化及蛋白酶体（proteosome）介导的降解。该胰岛素诱导的β抑制蛋白1含量降低，可分别被磷脂酰肌醇-3激酶（phosphatidylinositol-3 kinase, PI-3 kinase）抑制剂渥曼青霉素（wortmannin）与MEK抑制剂PD98059所阻断。我们同时发现，β抑制蛋白1与IGF-IR的结合显著减少，且IGF-I刺激的MAP激酶磷酸化水平被抑制55%。在经胰岛素处理、β抑制蛋白1表达下调的细胞中，溶血磷脂酸（lysophosphatidic acid, LPA）或异丙肾上腺素（isoproterenol, ISO）诱导的MAP激酶磷酸化过程被完全抑制。该效应与β抑制蛋白1与β₂肾上腺素能受体（β₂-AR）的结合减弱，以及β抑制蛋白1-Src与Src-β₂-AR的结合降低相关。在经胰岛素处理且内源性β抑制蛋白1表达下调的细胞中，异位表达野生型β抑制蛋白1，可挽救IGF-I及LPA刺激的MAP激酶磷酸化反应。综上，本研究得到以下结论：(i) 长期胰岛素处理可促进β抑制蛋白1的降解；(ii) 内源性β抑制蛋白1的该下调过程与IGF-I、LPA及ISO介导的MAP激酶信号转导减弱相关，且该效应可通过异位表达野生型β抑制蛋白1得到挽救；(iii) 本研究揭示了一种全新的异源脱敏机制：胰岛素处理可损害GPCR信号转导，并凸显了β抑制蛋白1作为该脱敏机制靶点分子的重要性。