Core--Cytoskeletal Facility

创伤引起大量生物活性介质的释放，如脂质、激素和细胞因子(项目I-A)。这些配体立即引起功能和新陈代谢的变化，但也改变了细胞对后续刺激的反应(无论是破坏性的还是建设性的)(S)。这些适应反应的机制(S)(分别称为启动或预适应)仍然是我们研究的重点(项目II、III、IV、VII和IX)。值得注意的是，配体通过第二信使激活信号，离子通量和蛋白质磷酸化继续得到广泛的研究。但配体结合也会触发受体内化。利用改造后的模型，我们(和其他人)现在接受了几类生物活性介质(如儿茶酚、脂类和肿瘤坏死因子)的激活受体是由笼蛋白包裹的囊泡内化的。包被网状蛋白的内涵体在微管轨道上围绕细胞运输。令人惊讶的是，最近的研究表明，受体内化是启动一个新的信号转导阶段所必需的。内化的G蛋白偶联受体(GPCR)发出的信号影响转录导向的蛋白激酶级联反应(包括PKC和MAPK模块)，导致细胞反应性的持续变化(项目V)。然而，在哺乳动物，尤其是人类细胞中，先前的信号事件如何影响内化受体、内吞机制和微管细胞骨架之间的相互作用尚不清楚。此外，尚不清楚巨噬细胞等促炎细胞对内皮细胞或平滑肌细胞等易受攻击的目标是否有类似的反应(项目六)。我们假设创伤后释放的循环介质(肾上腺素能、溶菌脂和细胞因子)(项目V、VII和IX)增强了膜、受体和随后的信号传递的内在化。我们最初的重点是为I选择的配体。与创伤的相关性，II。通过GPCR进行调解，以及III，激活PKC和MAPK模块。我们将研究这些测试配体如何影响来自人类和啮齿动物的一组髓系和非髓系细胞的A.构成膜/受体运输和B.内化依赖的信号。这些研究将为评估目前可获得的治疗策略是否可以针对笼蛋白包裹的内吞和基于微管的贩运奠定基础。

Trauma provokes the release of a large number of bioactive mediators, such as lipids, hormones and cytokines (Project I-A). These ligands elicit immediate changes in function and metabolism, but also modify the cellular response(s) to subsequent stimuli (both destructively or constructively). The mechanism(s) of these adapted responses (termed priming or preconditioning, respectively) remain the focus of our investigations (with Projects II, III, IV, VII, and IX). Deservedly, ligand activated signaling by second messengers, ion-flux and protein phosphorylation continue to receive extensive study. But ligand binding also triggers receptor internalization. Using transformed models we (and others) now accept that activated receptors for several classes of bioactive mediators (such as catechols, lipids and TNFa) are internalized by clathrin coated vesicles. Clathrin coated endosomes are transported about the cell on microtubular tracks. Surprisingly, recent studies indicate that receptor internalization is necessary to initiate a novel phase of signaling. Signals emanating from internalized G-protein coupled receptors (GPCR) affect transcriptionally directed protein kinase cascades (including PKC and the MAPK modules) causing sustained changes in cell responsiveness (with Project V). However it is unclear how previous signaling events influence the interactions between internalized receptors, the endocytotic machinery and the microtubular cytoskeleton, in mammalian, especially human cells. Moreover it is unclear whether pro-inflammatory cells such as macrophages respond similarly to vulnerable targets such as endothelial or smooth muscle cells (with Project VI). We hypothesize that circulating mediators (adrenergic, lysolipid and cytokines ) (with Projects V, VII and IX) released after trauma augment internalization for membranes, receptors and subsequent signal traffic. We initially focus on ligands selected for i. pertinence to trauma, ii. mediation by GPCR, and iii, activation of PKC and MAPK modules. We will examine how these test ligands influence a. constitutive membrane/receptor trafficking and b. internalization dependent signaling in a set of myeloid and non myeloid cells from humans and rodents. These studies will form the basis for evaluating whether currently accessible therapeutic strategies can be directed against clathrin coated endocytosis and microtubule based trafficking.