Mechanisms Regulating Human NK Cell Cytotoxicity

NK细胞是淋巴细胞的亚群，其独特的受体有助于检测感染， 转化的或“应激的”细胞。这种免疫识别随后导致NK细胞的发育- 介导的细胞毒性或细胞因子和趋化因子的产生，所述细胞因子和趋化因子激活细胞的其它组分。 免疫系统高IgE综合征（HIES）患者存在免疫失调， 皮肤疱疹病毒和乳头瘤病毒感染。一个主要的遗传异常发现在这些 患者是编码胞质分裂贡献因子8（DOCK 8）的基因中的缺失或功能丧失突变， 鸟嘌呤核苷酸交换因子（GEF）用于Cdc 42。复发性皮肤病毒感染的高发病率 DOCK 8缺陷患者中的感染提示自然杀伤（NK）细胞功能缺陷。其实别人 我们最近发现，DOCK 8缺陷的NK细胞具有降低的溶解功能， 粘附，F-肌动蛋白在细胞毒性突触处积聚，并且不能将颗粒溶解在细胞内。 靶细胞。然而，DOCK 8如何调节NK细胞杀伤发展中的如此多的关键步骤， 不知道。我们的中心假设是DOCK 8相互作用组协调肌动蛋白的调节， 和微管细胞骨架以促进NK细胞极化并影响NK细胞的细胞毒性。 根据本提案中包含的初步数据，我们假设：（a）DOCK 8激活CDC 42 （B）DOCK 8与WASP的相互作用对其定位是关键的 和在NKIS处的F-肌动蛋白产生;（c）通过DOCK 8的塔林蛋白募集介导NK细胞-靶标粘附;（d） HkRP 3是一种与DOCK 8相互作用的造血表达蛋白，参与MTOC极化， 溶解颗粒聚集，并通过与微管蛋白和动力蛋白的相互作用影响溶解颗粒聚集 马达复合物;（e）septins是调节NK细胞杀伤的DOCK 8相互作用蛋白;（f）博格稳定 Septin丝的活性受到活性Cdc 42的损害，以调节NK细胞的杀伤。为了测试这些 假设我们将：（1）确定DOCK 8调节F-肌动蛋白动力学和细胞增殖的机制。 （2）明确HkRP 3调节溶解颗粒会聚和MTOC的机制 （3）确定septins在NK细胞杀伤活性调节中的作用。的结果 提出的实验将为理解分子事件提供实验基础， 参与调节NK细胞效应功能，并将在更广泛的背景下，推进我们的研究。 理解细胞活化的基本过程，导致F-肌动蛋白调节，MTOC极化 和颗粒胞吐作用。此外，通过这些研究获得的信息可能会指导如何 DOCK 8在其他免疫细胞中发挥作用。

NK cells are a subpopulation of lymphocytes whose unique receptors facilitate the detection of infected, transformed, or `stressed' cells. This immune recognition subsequently leads to the development of NK cell- mediated cytotoxicity or the generation of cytokines and chemokines that activate other components of the immune system. Patients with hyper-IgE syndrome (HIES) have immune dysregulation and can be affected by recalcitrant cutaneous herpes virus and papillomavirus infections. A major genetic abnormality found in these patients is deletion or loss-of-function mutations in the gene encoding Dedicator of Cytokinesis 8 (DOCK8), a guanine nucleotide exchange factor (GEF) for Cdc42. The high incidence of recurrent cutaneous viral infections in DOCK8-deficient patients is suggestive of defects in natural killer (NK) cell function. In fact, others and we have recently shown that NK cells deficient in DOCK8 have reduced lytic function, decreased adhesion, F-actin accumulation at the cytotoxic synapse and an inability to polarize lytic granules toward the target cell. However, how DOCK8 can regulate so many critical steps in the development of NK cell killing is not known. It is our central hypothesis that the DOCK8-interactome coordinates the regulation of the actin and microtubule cytoskeletons to facilitate NK cell polarization and effect NK cellular cytotoxicity. Based on the preliminary data included in this proposal, we hypothesize that: (a) DOCK8 activation of CDC42 is critical to the development of NK cell killing; (b) DOCK8 interaction with WASP is critical for its localization and F-actin generation at the NKIS; (c) talin recruitment by DOCK8 mediates NK cell – target adhesion; (d) HkRP3, a hematopoietically expressed protein that interacts with DOCK8 is involved in MTOC polarization and lytic granule clustering and affects lytic granule clustering through its interactions with tubulin and the dynein motor complex; (e) septins are DOCK8 interacting proteins that regulate NK cell killing; (f) BORG stabilization of septin filaments is impaired by active Cdc42 in order to regulate NK cell killing. In order to test these hypotheses we will: (1) Determine the mechanism by which DOCK8 regulates F-actin dynamics and cell adhesion; (2) Define the mechanism by which HkRP3 regulates lytic granule convergence and MTOC polarization; (3) Determine the role of septins in the regulation of the NK cell cytotoxicity. The outcome of the proposed experiments will provide an experimental basis for understanding the molecular events that are involved in the regulation of NK cell effector functions, and will, in a broader context, advance our understanding of fundamental processes in cellular activation leading to F-actin regulation, MTOC polarization and granule exocytosis. Moreover, the information obtained through these studies will likely instruct how DOCK8 is functioning in other immune cells.