Regulation of ADP-ribosylation factor

协调的膜肌动蛋白重塑是许多细胞功能所必需的，包括(I)细胞运动，(Ii)内吞和吐出，以及(Iii)有丝分裂。许多控制膜和肌动蛋白重塑的信号分子包括磷脂酰肌醇、Arf家族GTP结合蛋白和Rho家族GTP结合蛋白。本实验室工作的主要目的是阐明Arf家族蛋白介导的信号调节机制。这项工作导致了一个Arf GTPase激活蛋白家族的鉴定，ASAPS可能整合了至少四个信号通路，提供特定细胞行为所需的膜和肌动蛋白的协调反应。ASAPS由ASAP1/2、ACAP1/2/3、AGAP1/2/3和ARAP1/2/3四个亚族组成。目前正在进行有两个总体目标的研究。 该实验室的重点之一是研究Arf间隙与Arf相互作用并影响Arf活性的特定分子机制。我们发现，一个Arf GAP的PH结构域ASAP1是通过Arf GAP结构域调节催化的变构位点。在这些研究中，我们还发现ASAP1含有独立于PH结构域发挥作用的定位结构域。以外壳适配蛋白GGA为模型，我们还发现Arf与Arf的相互作用要求Arf从其效应蛋白中解离出来。基于这些数据，我们提出了Arf间隙与Arf作用部位结合，并通过脂质环境的变化来感觉，例如，Arf何时应该失活。持续的研究旨在确定ARF和ARF GAP以及ARF和GGA之间的相互作用部位。此外，我们正在研究Arf间隙-Arf相互作用在货物分拣中的潜在作用，这些作用发生在膜运输过程中，如蛋白质分泌和胞吐。 该实验室的第二个重点是检查ASAP家族成员的特定细胞角色。我们从检查每个子科的代表性成员开始。已发现ASAP1可以定位并控制局部黏附的周转，这种黏附是将细胞固定在底物上的结构，必须重塑才能发生细胞运动。已发现ACAP1调节Arf6依赖的富含肌动蛋白的突起的形成，这一事件被认为参与细胞的扩散和运动。ARAP1还被发现同时调节膜运输和肌动蛋白细胞骨架，作为调节细胞运动的信号网络中的一个节点。PIP3与ARAP1结合，激活Arf间隙，从而使Arf失活并控制高尔基体功能。此外，ARAP1还可诱导CDC42的激活和Rho的失活。已经发现，这两种蛋白的相互调节可以诱导细胞迁移。因此，总的来说，ASAP家族的Arf间隙似乎调节着细胞迁移所涉及的事件。继续研究的目的是(I)进一步描述Arf间隙在信号转导中的作用，(Ii)识别依赖于Arf间隙信号的特定生物事件，例如。评估免疫细胞的发育和功能，以及(Iii)评估参与病理过程的可能性，特别是肿瘤细胞的侵袭和转移。 Z01 BC 07365-03 LBC至LCO

Coordinated membrane actin remodeling are integral to a number of cellular functions including (i) cell movement, (ii) endocytosis and exocytosis and (iii) mitosis. A number of signaling molecules that control either or both membrane and actin remodeling include phosphoinositides, Arf family GTP binding proteins and Rho family GTP-binding proteins. The main objective of the work in our laboratory is to elucidate the mechanisms that regulate the signals mediated by Arf family proteins. The work has led to the identification of a family of Arf GTPase-activating proteins, the ASAPs, that may integrate at least four signaling pathways, providing coordinated responses in membranes and actin necessary for particular cellular behaviors. The ASAPs are comprised of four subfamilies: ASAP1/2, ACAP1/2/3, AGAP1/2/3 and ARAP1/2/3. Studies with two general goals are being conducted. One emphasis of the laboratory is to examine specific molecular mechanisms by which Arf GAPs interact with Arf and impact Arf activities. We have found that the PH domain of one Arf GAP, ASAP1, is an allosteric site regulating catalysis through the Arf GAP domain. In those studies, we also found that ASAP1 contains localization domains that function independently of the PH domain. Using the coat adaptor protein, GGA, as a model, we have also found that interaction of Arf GAP with Arf requires that Arf dissociate from its effector proteins. Based on these data, we have proposed that Arf GAPs bind to the site of Arf action, and senses, through changes in the lipid environment for instance, when Arf should be inactivated. Continuing studies are designed to define the site of interaction between Arf and Arf GAP as well as between Arf and GGA. In addition, we are examining the potential role of Arf GAP - Arf interactions in cargo sorting that occurs during membrane trafficking events such as protein secretion and exocytosis. A second emphasis of the laboratory is an examination of the specific cellular roles of ASAP family members. We have started by examining representative members of each subfamily. ASAP1 has been found to localize to and control the turnover of focal adhesion, structures that anchor cells to the substratum and which must be remodeled for cell movement to occur. ACAP1 has been found to regulate Arf6 dependent actin-rich protrusion formation, an event thought to be involved in cell spreading and movement. ARAP1 has been also found to regulate both membrane trafficking and the actin cytoskeleton, functioning as a node in a signaling network that regulates cell movement. PIP3 binds to ARAP1, activating the Arf GAP, thereby inactivating Arf and controlling Golgi function. In addition, ARAP1 both induces the activation of Cdc42 and the inactivation of Rho. Reciprocal regulation of these two proteins has been found to induce cell migration. Therefore, in general the ASAP family of Arf GAPs appear to regulate events involved in cell migration. Continuing studies are designed to (i) further delineate the role of Arf GAPs in signaling, (ii) identify specific biological events dependent on signaling through Arf GAPs, eg. Development and function of immunological cells and (iii) evaluate a possible involvement in pathological processes, particularly tumor cell invasion and metastasis. Z01 BC 07365-03 LBC to LCO