Área de Endocrinología Molecular y Celular, Instituto de Investigación Sanitaria de Santiago, Hospital Clinico Universitario de Santiago, Servicio Gallego de Salud, 15706 Santiago de Compostela, Spain.
Mol Biol Cell. 2011 Nov;22(21):4182-91. doi: 10.1091/mbc.E11-04-0373. Epub 2011 Sep 7.
The aim of the present study was to identify the signaling mechanism(s) responsible for the modulation of growth hormone secretagogue receptor type 1a (GHSR1a)-associated Akt activity. Ghrelin leads to the activation of Akt through the interplay of distinct signaling mechanisms: an early G(i/o) protein-dependent pathway and a late pathway mediated by β-arrestins. We found that the Src homology 2-containing protein tyrosine phosphatase (SHP-1) was an essential molecule in both G(i/o) protein-dependent and β-arrestin-mediated pathways. More specifically, the role of SHP-1 in the G(i/o) protein-dependent pathway was demonstrated by the fact that the overexpression of a catalytically defective SHP-1 augments tyrosine phosphorylation of the PI3K regulatory subunit p85, leading to an increase in the phosphorylation of cSrc and phosphoinositide-dependent protein kinase 1, and finally activating Akt. The presence of SHP-1 in the β-arrestin-scaffolded complex and its attenuating effect on the cSrc and Akt activities verified that SHP-1 regulates not only the G(i/o) protein-dependent pathway but also the β-arrestin-mediated pathway. Assays performed in preadipocyte and adipocyte 3T3-L1 cells showed SHP-1 expression. According to our results in HEK-GHSR1a cells, ghrelin stimulated SHP-1 phosphorylation in 3T3-L1 cells. The increase in ghrelin-induced Akt activity was enhanced by small interfering RNA of SHP-1 in preadipocyte 3T3-L1 cells. These results were reproduced in white adipose tissue obtained from mice, in which SHP-1 exhibited higher expression in omental than in subcutaneous tissue. Furthermore, this pattern of expression was inverted in mice fed a high-fat diet, suggesting a role for SHP-1 in controlling ghrelin sensitivity in adipose tissue. Indeed, SHP-1 deficiency was associated with augmented ghrelin-evoked Akt phosphorylation in omental tissue, as well as decreased phosphorylation under overexpression of SHP-1 in subcutaneous tissue. These findings showed a novel role for SHP-1 in the regulation of Akt activity through the modulation of the ghrelin/GHSR1a system signaling.
本研究旨在确定负责生长激素促分泌素受体 1a (GHSR1a)相关 Akt 活性调节的信号转导机制。Ghrelin 通过独特的信号转导机制激活 Akt：早期 G(i/o) 蛋白依赖性途径和晚期途径由β-arrestin 介导。我们发现含Src 同源 2 结构域的蛋白酪氨酸磷酸酶（SHP-1）是 G(i/o) 蛋白依赖性途径和β-arrestin 介导途径中必不可少的分子。更具体地说，过表达催化失活的 SHP-1 会增强 PI3K 调节亚基 p85 的酪氨酸磷酸化，导致 cSrc 和磷酸肌醇依赖性蛋白激酶 1 的磷酸化增加，最终激活 Akt，这表明 SHP-1 在 G(i/o) 蛋白依赖性途径中的作用。SHP-1 存在于β-arrestin 支架复合物中，并减弱其对 cSrc 和 Akt 活性的影响，证实 SHP-1 不仅调节 G(i/o) 蛋白依赖性途径，还调节β-arrestin 介导的途径。在脂肪前体细胞和脂肪细胞 3T3-L1 细胞中进行的测定显示 SHP-1 的表达。根据我们在 HEK-GHSR1a 细胞中的结果，ghrelin 刺激 3T3-L1 细胞中的 SHP-1 磷酸化。在脂肪前体细胞 3T3-L1 细胞中，用 SHP-1 的小干扰 RNA 增强了 ghrelin 诱导的 Akt 活性的增加。在从小鼠获得的白色脂肪组织中重现了这些结果，其中 SHP-1 在网膜中表达高于皮下组织。此外，这种表达模式在喂食高脂肪饮食的小鼠中发生逆转，表明 SHP-1 在控制脂肪组织中 ghrelin 的敏感性中起作用。事实上，SHP-1 缺陷与网膜组织中 ghrelin 诱导的 Akt 磷酸化增加以及皮下组织中 SHP-1 过表达时的磷酸化减少有关。这些发现表明 SHP-1 在通过调节 ghrelin/GHSR1a 系统信号转导来调节 Akt 活性方面发挥了新的作用。
