The mechanisms underlying the GPCR-mediated chemotaxis in D. discoideum

1：cAR 1介导的Ras信号传导的时空动力学。 为了揭示抑制机制，我们通过观察响应于cAMP梯度的与GFP融合的活性Ras结合结构域（RBD-GFP）的荧光探针的膜易位来监测Ras的时空激活。 我们观察到的Ras激活动力学表明，一个负调节，可能是一个RasGAP，逐渐招募到膜，促进Ras失活。 在哺乳动物细胞中，含有磷脂结合结构域的RasGAP已被证明可以移位到质膜的内表面，使Ras失活，从而抑制PI 3 K。 我们提出，一个类似的磷脂结合的RasGAP可能是一个重要的抑制剂，在cAR 1介导的化学传感在D。盘状突 为了验证这一假设，我们正在研究RasGAP在D.盘状突 图2：一种新型Gbg效应子ElmoE在趋化过程中将GPCR信号转导至肌动蛋白网络。 G蛋白偶联受体（GPCR）的激活导致异源三聚体G蛋白解离成Ga和Gbg亚基，其继续调节参与一系列细胞反应的各种效应物。 在趋化过程中，Gbg亚基调节肌动蛋白的组装和迁移，但将Gbg连接到肌动蛋白细胞骨架的蛋白质仍然未知。 在这里，我们确定了一个新的Gbg效应，ElmoE在Dictyosteopathy，并证明它是GPCR介导的趋化性所需的。值得注意的是，ElmoE与Gbg和Dock样蛋白相互作用，以GPCR依赖的方式激活小GTdR Rac，并且还与Arp 2/3复合物和F-actin缔合。 因此，ElmoE作为化学引诱物GPCR，G蛋白和肌动蛋白细胞骨架之间的第一个直接联系。 由GPCR、Gbg、埃尔莫/Dock、Rac和Arp 2/3组成的途径在趋化性过程中在空间上引导真核细胞伪足中树突状肌动蛋白网络的生长（Yan et al Dev. Cell 2012）。

1: cAR1-mediated spatiotemporal dynamics of Ras signaling. To reveal the inhibitory mechanism, we monitored spatiotemporal activation of Ras by observing the membrane translocation of a fluorescent probe, active Ras Binding Domain fused to GFP (RBD-GFP) in response to a cAMP gradient. The Ras-activation dynamics we observed indicate that a negative regulator, possibly a RasGAP, is gradually recruited to the membrane and promotes Ras inactivation. In mammalian cells, a RasGAP containing a phospholipid-binding domain has been shown to translocate to the plasma membrane's inner face, deactivate Ras, and thereby inhibit PI3K. We proposed that a similar phospholipid-bound RasGAP is likely to be an important inhibitor in cAR1-mediated chemosensing in D. discoideum. To test this hypothesis, we are studying the roles of RasGAP in chemosensing in D. discoideum. 2: A novel Gbg effector, ElmoE, transduces GPCR signaling to the actin network during chemotaxis. Activation of G-protein-coupled receptors (GPCRs) leads to the dissociation of heterotrimeric G-proteins into Ga and Gbg subunits, which go on to regulate various effectors involved in a panoply of cellular responses. During chemotaxis, Gbg subunits regulate actin assembly and migration, but the protein(s) linking Gbg to the actin cytoskeleton remains unknown. Here, we identified a new Gbg effector, ElmoE in Dictyostelium, and demonstrated that it is required for GPCR-mediated chemotaxis. Remarkably, ElmoE interacts with Gbg and Dock-like proteins to activate the small GTPase Rac, in a GPCR-dependent manner, and also associates with Arp2/3 complex and F-actin. Thus, ElmoE serves as the first direct link between chemoattractant GPCRs, G-proteins and the actin cytoskeleton. The pathway, consisting of GPCR, Gbg, Elmo/Dock, Rac, and Arp2/3, spatially guides the growth of dendritic actin networks in pseudopods of eukaryotic cells during chemotaxis (Yan et al Dev. Cell 2012).