Department of Biochemistry and Immunology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Genes Immun. 2010 Sep;11(6):447-57. doi: 10.1038/gene.2009.107. Epub 2010 Jan 21.
Guanine-nucleotide exchange factors (GEFs) stimulate the intrinsic GDP/GTP exchange activity of Ras and promote the formation of active Ras-GTP, which in turn controls diverse signalling networks important for the regulation of cell proliferation, survival, differentiation, vesicular trafficking, and gene expression. RasGEF1b is a GEF, whose expression is induced in macrophages on stimulation with toll-like receptor (TLR) agonists. Here, we showed that in vitro RasGEF1b expression by macrophages is mostly induced by TLR3 (poly I:C) and TLR4 (lipopolysaccharyde) through the MyD88-independent pathway. In vivo infection with the protozoan parasites Trypanosoma cruzi and Plasmodium chabaudi induced RasGEF1b in an MyD88-, TRIF-, and IFN-gamma-dependent manner. Ectopically expressed RasGEF1b was found, mostly, in the heavy membrane fraction of HEK 293T, and by confocal microscopy, it was found to be located at early endosomes. Computational modelling of the RasGEF1b-Ras interaction revealed that RasGEF1b interacts with the binding domain site of Ras, a critical region for interacting with GEFs involved in the activation of Ras-Raf-MEK-ERK pathway. More important, RasGEF1b was found to be closely associated with Ras in live cells and to trigger Ras activity. Altogether, these results indicate that on TLR activation, RasGEF1b may trigger Ras-like proteins and regulate specific biological activities described for this subtype of GTPases.
鸟嘌呤核苷酸交换因子（GEFs）刺激 Ras 的固有 GDP/GTP 交换活性，并促进活性 Ras-GTP 的形成，从而控制对细胞增殖、存活、分化、囊泡运输和基因表达的调节至关重要的各种信号转导网络。RasGEF1b 是一种 GEF，其表达在巨噬细胞受到 Toll 样受体（TLR）激动剂刺激时被诱导。在这里，我们表明，巨噬细胞中的 RasGEF1b 表达主要是由 TLR3（poly I:C）和 TLR4（脂多糖）通过 MyD88 非依赖性途径诱导的。原虫寄生虫 Trypanosoma cruzi 和 Plasmodium chabaudi 的体内感染以 MyD88、TRIF 和 IFN-γ依赖性方式诱导 RasGEF1b。异位表达的 RasGEF1b 主要存在于 HEK 293T 的重膜部分，通过共聚焦显微镜观察，发现它位于早期内体中。RasGEF1b-Ras 相互作用的计算模型表明，RasGEF1b 与 Ras 的结合域位点相互作用，该位点对于与 Ras-Raf-MEK-ERK 途径激活相关的 GEFs 相互作用至关重要。更重要的是，发现 RasGEF1b 在活细胞中与 Ras 密切相关，并触发 Ras 活性。总之，这些结果表明，在 TLR 激活时，RasGEF1b 可能触发 Ras 样蛋白并调节该亚型 GTPase 描述的特定生物学活性。
