Dobkin-Bekman Masha, Naidich Michal, Rahamim Liat, Przedecki Fiorenza, Almog Tal, Lim Stefan, Melamed Philippa, Liu Ping, Wohland Thorsten, Yao Zhong, Seger Rony, Naor Zvi
Department of Biochemistry, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel-Aviv 69978, Israel.
Mol Endocrinol. 2009 Nov;23(11):1850-64. doi: 10.1210/me.2008-0260. Epub 2009 Jul 23.
Most receptor tyrosine kinases and G protein-coupled receptors (GPCRs) operate via a limited number of MAPK cascades but still exert diverse functions, and therefore signal specificity remains an enigma. Also, most GPCR ligands utilize families of receptors for mediation of diverse biological actions; however, the mammalian type I GnRH receptor (GnRHR) seems to be the sole receptor mediating GnRH-induced gonadotropin synthesis and release. Signaling complexes associated with GPCRs may thus provide the means for signal specificity. Here we describe a signaling complex associated with the GnRHR, which is a unique GPCR lacking a C-terminal tail. Unlike other GPCRs, this signaling complex is preformed, and exposure of L beta T2 gonadotropes to GnRH induces its dynamic rearrangement. The signaling complex includes c-Src, protein kinase C delta, -epsilon, and -alpha, Ras, MAPK kinase 1/2, ERK1/2, tubulin, focal adhesion kinase (FAK), paxillin, vinculin, caveolin-1, kinase suppressor of Ras-1, and the GnRHR. Exposure to GnRH (5 min) causes MAPK kinase 1/2, ERK1/2, tubulin, vinculin, and the GnRHR to detach from c-Src, but they reassociate within 30 min. On the other hand, FAK, paxillin, the protein kinase Cs, and caveolin-1 stay bound to c-Src, whereas kinase suppressor of Ras-1 appears in the complex only 30 min after GnRH stimulation. GnRH was found to activate ERK1/2 in the complex in a c-Src-dependent manner, and the activated ERK1/2 subsequently phosphorylates FAK and paxillin. In parallel, caveolin-1, FAK, vinculin, and paxillin are phosphorylated on Tyr residues apparently by GnRH-activated c-Src. Receptor tyrosine kinases and GPCRs translocate ERK1/2 to the nucleus to phosphorylate and activate transcription factors. We therefore propose that the role of the multiprotein signaling complex is to sequester a cytosolic pool of activated ERK1/2 to phosphorylate FAK and paxillin at focal adhesions.
大多数受体酪氨酸激酶和G蛋白偶联受体（GPCR）通过有限数量的丝裂原活化蛋白激酶（MAPK）级联发挥作用，但仍能发挥多种功能，因此信号特异性仍是一个谜。此外，大多数GPCR配体利用受体家族来介导多种生物学作用；然而，哺乳动物I型促性腺激素释放激素受体（GnRHR）似乎是介导GnRH诱导的促性腺激素合成和释放的唯一受体。因此，与GPCR相关的信号复合物可能提供了信号特异性的方式。在这里，我们描述了一种与GnRHR相关的信号复合物，GnRHR是一种独特的缺乏C末端尾巴的GPCR。与其他GPCR不同，这种信号复合物是预先形成的，将LβT2促性腺激素细胞暴露于GnRH会诱导其动态重排。该信号复合物包括c-Src、蛋白激酶Cδ、-ε和-α、Ras、MAPK激酶1/2、细胞外信号调节激酶1/2（ERK1/2）、微管蛋白、粘着斑激酶（FAK）、桩蛋白、纽蛋白、小窝蛋白-1、Ras-1激酶抑制因子和GnRHR。暴露于GnRH（5分钟）会导致MAPK激酶1/2、ERK1/2、微管蛋白、纽蛋白和GnRHR与c-Src分离，但它们会在30分钟内重新结合。另一方面，FAK、桩蛋白、蛋白激酶C和小窝蛋白-1仍与c-Src结合，而Ras-1激酶抑制因子仅在GnRH刺激30分钟后才出现在复合物中。发现GnRH以c-Src依赖性方式激活复合物中的ERK1/2，激活的ERK1/2随后磷酸化FAK和桩蛋白。同时，小窝蛋白-1、FAK、纽蛋白和桩蛋白显然被GnRH激活的c-Src磷酸化酪氨酸残基。受体酪氨酸激酶和GPCR将ERK1/2转运到细胞核中以磷酸化并激活转录因子。因此，我们提出多蛋白信号复合物的作用是隔离细胞质中活化的ERK1/2池，以便在粘着斑处磷酸化FAK和桩蛋白。
