RhoGDI2的磷酸化在SDF-1引起的β1整合素介导的Jurkat细胞迁移过程中的作用研究

The microenvironment in bone marrow is particularly important for the origination and development of leukemia. SDF-1 expressed by bone marrow binds to its receptor CXCR4 which is expressed in various leukemic cells. Their interaction directs the migration of leukemic cells into bone marrow. Additionally, α4β1 mediates binding of cells to marrow stromal elements and cellular migration, which is associated with the poor prognosis. It has been reported that SDF-1 increases the affinity of leukemia cells to the microenvironmental niches in the bone marrow, probably by increasing the affinity of α4β1 to bone matrix. However, this possibility and the specific mechanism has not yet been well studied.The activation of integrin is relevant to Rho which is tightly regulated by RhoGDI2. The role of RhoGDI2 in the migration and metastasis of solid tumor has been widely investigated. However, till now there's no report on its role in leukemia. In our previous experiment, we found obvious phosphorylation of RhoGDI2 in Jurkat cells treated with SDF-1. In this project, we will, via in vivo and in vitro experiments, investigate the effects of specific phosphorylation of RhoGDI2 on the migration of Jurkat cells to bone marrow which is induced by SDF-1, find out the crucial signal pathway in the migration of leukemic cells into bone marrow, and thereby provide theoretical refferences and novel targets for the cure of leukemia.

骨髓微环境对白血病的发生和发展具有重要作用。骨髓内的SDF-1与表达于白血病细胞表面的CXCR4结合，促进白血病细胞向骨髓的侵润。此外，α4β1参与白血病细胞与骨髓基质的结合和细胞迁移，与白血病的愈后较差有关。有报道称，SDF-1很可能通过增加α4β1与骨髓基质的亲和力，增强白血病细胞对骨髓微环境巢亲和力，但具体机制有待研究。整合素的活化与受控于RhoGDI2的Rho的活性有关。RhoGDI2在实体瘤转移中的作用备受关注，但目前尚无RhoGDI2在白血病中的报道。我们的前期研究发现Jurkat细胞受SDF-1刺激后，RhoGDI2发生明显的磷酸化。本项目将通过体内和体外实验，研究RhoGDI2特异位点的磷酸化在SDF-1引起的Jurkat急性白血病细胞向骨髓基质侵润过程中的作用，阐明白血病细胞向骨髓侵润的关键信号通路，为治疗白血病提供理论依据和新的靶点。

一方面，基质细胞衍生因子-1（SDF-1）信号通路通过诱导趋化性迁移在T细胞急性淋巴细胞白血病的维持和进程起重要的作用。然而，SDF-1参与淋巴细胞迁移的分子机制尚需深入研究。另外，有研究表明，受到RhoGDIs负调控的RhoGTP酶参与细胞骨架的调控。为了研究RhoGDIs在SDF-1引起的T-ALL（T细胞免疫表型急性淋巴细胞白血病）迁移过程中的作用和机制，我们使用了Jurkat急性淋巴细胞白血病细胞系。结果显示SDF-1通过调控F-actin的重新分布和聚合诱导Jurkat细胞迁移，此过程依赖于Rho的活化。SDF-1诱导RhoA/C活化，活性氧簇（ROS）和NO的产生。依赖Rho的ROS/NO导致随后的细胞骨架重新分布和聚合。此外，RhoA/C和ROS/NO参与SDF-1诱导的Jurkat细胞迁移。因此，我们发现了一条SDF-1 / CXCR4→RhoA/C→ROS/NO→细胞骨架信号通路，调控SDF-1引起的Jurkat细胞迁移。RhoGDI2 153Y和24Y的磷酸化对于RhoA/C的释放至关重要，可能参与RhoA/C的活化。这一研究有助于更清晰地认识急性淋巴细胞白血病的迁移机制；另一方面，淋巴细胞迁移是免疫反应的必要的生理步骤。L-选择素和P-选择素糖蛋白配体1（PSGL-1）是白细胞表面组成性表达的黏附分子。它们参与白细胞的迁移和活化。尽管他们是不同的分子，却能够引起相似的生理事件。在已发表的论文中，我们研究了在中性粒细胞在E-选择素滚动过程中PI3K对于L-选择素-和PSGL-1-介导的 F-actin 重新分布和聚合的调控作用。在此，我们进一步研究了PI3K连接L-选择素和PSGL-1信号通路的可能性。首先证明了Jurkat细胞中L-选择素和PSGL-1刺激能够上调IL-18的转录水平，然后发现PI3K抑制剂LY294004能够降低L-选择素-和PSGL-1-引起的Jurkat细胞IL-18 mRNA的上调。转染PTEN 的表达质粒能够抑制IL-18的转录水平。因此，我们证明了PI3K是L-选择素和PSGL-1的信号连接体，为迁移引起的相关疾病的干预提供了基础。