Persistent cAMP signaling by thyrotropin (TSH) receptors is not dependent on internalization
Persistent cAMP signaling by thyrotropin (TSH) receptors is not dependent on internalization
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DOI：
10.1096/fj.10-161745
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发表时间：
2010-10-01
期刊：
影响因子：
4.8
通讯作者：
Gershengorn, Marvin C.
中科院分区：
文献类型：
--
作者：
Neumann, Susanne;Geras-Raaka, Elizabeth;Gershengorn, Marvin C.
有证据表明，促甲状腺素 [促甲状腺激素 (TSH)] 刺激的持续 cAMP 信号传导依赖于受体（具有 G 蛋白 α 亚基和腺苷酸环化酶）内化。由于尚不清楚 G 蛋白和腺苷酸环化酶是否与受体内化，因此我们测试了 TSH 受体 (TSHR) 持续的 cAMP 信号传导是否依赖于内化。我们测量了永久表达人 TSHR 的 HEK-EM293 细胞中 cAMP 的持续 TSHR 信号积累，在洗涤细胞以去除未结合的 TSH 后，与异丁基甲基黄嘌呤一起孵育 30 分钟，并通过使用 Alexa 标记的 TSH 的荧光显微镜和使用 I-125-TSH 的结合测定来测量 TSHR 内化。 TSHR，而非密切相关的促黄体激素或促卵泡激素受体，表现出持续的 cAMP 信号传导。与未标记的 TSH 一起孵育 30 分钟后，TSHR 并未内化；然而，β-arrestin-2 的表达促进了 TSHR 内化，而 TSHR 内化受到 dynasore（一种动力抑制剂）的抑制。 β-arrestin-2 的表达对 TSHR cAMP 信号传导没有影响，dynasore 在不存在或存在 TSHR 内化的情况下抑制 TSHR cAMP 信号传导，并且抑制内化的显性失活突变体动力蛋白的表达对持续 cAMP 信号传导没有影响。持续的 cAMP 信号传导被小分子 TSHR 拮抗剂特异性抑制。我们的结论是，TSHR 不必内化即可表现出持久的 cAMP 信号传导。-Neumann, S.、Geras-Raaka, E.、Marcus-Samuels, B.、Gershengorn, M. C. 促甲状腺素 (TSH) 受体的持久 cAMP 信号传导并不依赖于内化。 FASEB J. 24, 3992-3999 (2010)。 www.fasebj.org
Evidence was presented that thyrotropin [thyroid-stimulating hormone (TSH)]-stimulated persistent cAMP signaling is dependent on receptor (with G-protein alpha subunits and adenylyl cyclase) internalization. Because it is not clear whether G proteins and adenylyl cyclase internalize with receptors, we tested whether persistent cAMP signaling by TSH receptor (TSHR) is dependent on internalization. We measured persistent TSHR signaling as an accumulation of cAMP in HEK-EM293 cells permanently expressing human TSHRs incubated with isobutylmethylxanthine for 30 min after washing the cells to remove unbound TSH, and TSHR internalization by fluorescence microscopy using Alexa-tagged TSH and binding assays using I-125-TSH. TSHRs, but not the closely related lutropin or follitropin receptors, exhibit persistent cAMP signaling. TSHRs were not internalized by 30 min incubation with unlabeled TSH; however, expression of beta-arrestin-2 promoted TSHR internalization that was inhibited by dynasore, a dynamin inhibitor. Expression of beta-arrestin-2 had no effect on TSHR cAMP signaling, dynasore inhibited TSHR cAMP signaling in the absence or presence of TSHR internalization, and expression of a dominant-negative mutant dynamin, which inhibited internalization, had no effect on persistent cAMP signaling. Persistent cAMP signaling was specifically inhibited by a small molecule TSHR antagonist. We conclude that TSHRs do not have to be internalized to exhibit persistent cAMP signaling.-Neumann, S., Geras-Raaka, E., Marcus-Samuels, B., Gershengorn, M. C. Persistent cAMP signaling by thyrotropin (TSH) receptors is not dependent on internalization. FASEB J. 24, 3992-3999 (2010). www.fasebj.org