Laboratorio de Patología y Farmacología Molecular, Instituto de Biología y Medicina Experimental, Buenos Aires, Argentina.
Exp Cell Res. 2010 Feb 1;316(3):401-11. doi: 10.1016/j.yexcr.2009.11.002. Epub 2009 Nov 11.
It is well established that histamine modulates cell proliferation through the activation of the histamine H1 receptor (H1R), a G protein-coupled receptor (GPCR) that is known to couple to phospholipase C (PLC) activation via Gq. In the present study, we aimed to determine whether H1R activation modulates Rho GTPases, well-known effectors of Gq/G(11)-coupled receptors, and whether such modulation influences cell proliferation. Experiments were carried out in CHO cells stably expressing H1R (CHO-H1R). By using pull-down assays, we found that both histamine and a selective H1R agonist activated Rac and RhoA in a time- and dose-dependent manner without significant changes in the activation of Cdc42. Histamine response was abolished by the H1R antagonist mepyramine, RGS2 and the PLC inhibitor U73122, suggesting that Rac and RhoA activation is mediated by H1R via Gq coupling to PLC stimulation. Histamine caused a marked activation of serum response factor activity via the H1R, as determined with a serum-responsive element (SRE) luciferase reporter, and this response was inhibited by RhoA inactivation with C3 toxin. Histamine also caused a significant activation of JNK which was inhibited by expression of the Rac-GAP beta2-chimaerin. On the other hand, H1R-induced ERK1/2 activation was inhibited by U73122 but not affected by C3 or beta2-chimaerin, suggesting that ERK1/2 activation was dependent on PLC and independent of RhoA or Rac. [(3)H]-Thymidine incorporation assays showed that both histamine and the H1R agonist inhibited cell proliferation in a dose-dependent manner and that the effect was independent of RhoA but partially dependent on JNK and Rac. Our results reveal that functional coupling of the H1R to Gq-PLC leads to the activation of RhoA and Rac small GTPases and suggest distinct roles for Rho GTPases in the control of cell proliferation by histamine.
组胺通过激活组胺 H1 受体（H1R）来调节细胞增殖，H1R 是一种 G 蛋白偶联受体（GPCR），已知通过 Gq 与磷脂酶 C（PLC）的激活偶联。在本研究中，我们旨在确定 H1R 激活是否调节 Rho GTPases，即众所周知的 Gq/G(11)偶联受体的效应物，以及这种调节是否影响细胞增殖。实验在稳定表达 H1R（CHO-H1R）的 CHO 细胞中进行。通过使用下拉测定法，我们发现组胺和选择性 H1R 激动剂均以时间和剂量依赖性方式激活 Rac 和 RhoA，而 Cdc42 的激活没有明显变化。组胺反应被 H1R 拮抗剂甲吡拉敏、RGS2 和 PLC 抑制剂 U73122 阻断，表明 Rac 和 RhoA 的激活是通过 H1R 与 Gq 偶联刺激 PLC 来介导的。组胺通过 H1R 引起血清反应因子活性的显著激活，如血清反应元件（SRE）荧光素酶报告所确定的，并且该反应被 C3 毒素抑制 RhoA 失活而抑制。组胺还引起 JNK 的显著激活，该激活被 Rac-GAP beta2-奇美拉表达抑制。另一方面，H1R 诱导的 ERK1/2 激活被 U73122 抑制，但不受 C3 或 beta2-奇美拉影响，表明 ERK1/2 激活依赖于 PLC，而不依赖于 RhoA 或 Rac。[（3）H]-胸苷掺入测定表明，组胺和 H1R 激动剂均以剂量依赖性方式抑制细胞增殖，并且该作用不依赖于 RhoA，但部分依赖于 JNK 和 Rac。我们的结果表明，H1R 与 Gq-PLC 的功能偶联导致 RhoA 和 Rac 小 GTPases 的激活，并表明 Rho GTPases 在组胺控制细胞增殖中具有不同的作用。
