Elucidation of a eukaryotic chemorepulsion mechanism

有很好的证据表明，一些细胞分泌化学排斥剂，导致特定类型的细胞移动 离他们远点然而，关于化学排斥剂的身份、它们的生物学特性以及它们的生物学特性， 受体，以及它们用于指导细胞运动的机制。我们发现增殖的网骨细胞 分泌一种叫做AprA蛋白质，AprA是一种细胞外信号，起着化学排斥剂的作用。 尽管AprA与哺乳动物蛋白质几乎没有序列相似性，但AprA预测了与哺乳动物蛋白质的结构相似性。 人分泌的二肽基蛋白酶DPPIV，并与DPPIV共享功能特性。我们发现 人DPPIV是人和小鼠嗜中性粒细胞化学排斥剂，当局部应用时，DPPIV可 诱导中性粒细胞离开两个被称为急性呼吸窘迫的肺部疾病小鼠模型的组织 急性呼吸窘迫综合征（ARDS）和类风湿性关节炎的小鼠模型。为了深入了解一个基本的机制， 用于形态发生，诱导中性粒细胞离开组织的方法，以及如何增加或减少 化学排斥剂的作用，我们提出了三个具体的目标，以阐明所使用的分子机制， AprA和DPPIV导致化疗排斥。目的1是鉴定AprA受体，因为它在以下方面起关键作用： Dictyosteoblasts化学排斥机制。我们的初步工作已经确定了一种预测的G蛋白- 偶联受体称为GrlH作为可能的AprA受体。我们会仔细测试，如果GrlH不是 受体，我们将使用几种方法来识别受体。目的2是阐明AprA 化学排斥信号转导途径我们的初步数据表明， 化学排斥机制不同于化学吸引机制所使用的组分， 使网骨藻细胞向cAMP聚集。我们将在多大程度上确定 化学排斥机制使用化学吸引机制的已知组分，以及使用 无偏遗传筛选在网骨藻中鉴定化学排斥的其他成分的能力 机制目的3是检验DPPIV使用称为PAR 2的G蛋白偶联受体来 诱导中性粒细胞化学排斥，并利用我们所了解的网骨藻化学排斥机制， 以确定网骨藻和人类中性粒细胞之间的相似性和差异 化学排斥机制总之，这项工作结合了分子生物学，遗传学，细胞生物学， 生物化学将有助于阐明真核生物的化学排斥机制，并确定潜在的药物靶点 可以增强或抑制化学排斥

There is good evidence that some cells secrete chemorepellents that cause specific cell types to move away from them. However, much remains to be understood about the identity of the chemorepellents, their receptors, and the mechanisms they use to direct cell motility. We found that proliferating Dictyostelium cells secrete a protein called AprA, and that AprA is an extracellular signal that functions as a chemorepellent. Although AprA has little sequence similarity to mammalian proteins, AprA has predicted structural similarity to the human secreted dipeptidyl protease DPPIV, and shares functional properties with DPPIV. We found that human DPPIV is a chemorepellent for human and mouse neutrophils, and when applied locally, DPPIV can induce neutrophils to leave a tissue in two mouse models of a lung disease called acute respiratory distress syndrome (ARDS), and a mouse model of rheumatoid arthritis. To gain insights into a fundamental mechanism used in morphogenesis, ways to induce neutrophils to leave a tissue, and how one could augment or diminish the effect of a chemorepellent, we propose three specific aims to elucidate the molecular mechanisms used by AprA and DPPIV to cause chemorepulsion. Aim 1 is to identify the AprA receptor, since this plays a key role in the Dictyostelium chemorepulsion mechanism. Our preliminary work has identified a predicted G protein- coupled receptor called GrlH as a possible AprA receptor. We will carefully test this, and if GrlH is not the receptor, we will use several approaches to identify the receptor. Aim 2 is to elucidate the AprA chemorepulsion signal transduction pathway. Our preliminary data indicate that some components of the chemorepulsion mechanism are different from components used by the chemoattraction mechanism that allows Dictyostelium cells to aggregate toward cAMP. We will determine the extent to which the chemorepulsion mechanism uses known components of the chemoattraction mechanism, as well as use the power of unbiased genetic screens in Dictyostelium to identify additional components of the chemorepulsion mechanism. Aim 3 is to test the hypothesis that DPPIV uses a G protein-coupled receptor called PAR2 to induce neutrophil chemorepulsion, and use what we learn about the Dictyostelium chemorepulsion mechanism to determine the similarities and differences between the Dictyostelium and the human neutrophil chemorepulsion mechanisms. Together, this work combining molecular biology, genetics, cell biology, and biochemistry will help to elucidate eukaryotic chemorepulsion mechanisms, and identify potential drug targets that could enhance or inhibit chemorepulsion.