Autocrine control of neutrophil chemotaxis

趋化性使中性粒细胞(PMN)迅速到达感染和炎症部位。尽管我们对趋化性的理解最近取得了进展，但仍有许多悬而未决的问题。化学诱导剂通过嘌呤能受体控制趋化作用，刺激中性粒细胞释放三磷酸腺苷和自分泌反馈环。在这个项目中，我们修改了拟议工作的范围，以了解趋化信号如何诱导ATP释放，以及腺苷是如何由释放的ATP控制负反馈环的，这些负反馈环在定义细胞极性和尾足回缩方面发挥关键作用。拟议的项目基于以下修订的工作假设：刺激趋化受体诱导快速的细胞内事件，导致ATP释放。通过不同的机制，额外的ATP释放提供了抑制A2a腺苷受体的配体--腺苷。这些受体增加了退缩边缘的cAMP水平，提供了一个抑制反馈环路，引起全局抑制，促进尾足类收缩，定义细胞极性，促进迁移。我们建议通过解决以下修订后的具体目标来测试这一工作假设： 具体目标1.导致ATP释放的上游事件我们将确定在梯度感应过程中连接趋化受体激活和ATP释放的上游信号通路。此外，我们还将研究不同的ATP释放机制及其在梯度感应、细胞极化和迁移过程中对ATP释放的贡献。重点将放在趋化受体下游的信号通路上，例如钙信号和MAPK激活，导致连接蛋白或连接蛋白等ATP释放通道的开放，以及通过囊泡运输释放ATP。 特定目的2.全局抑制和嘌呤能信号我们将研究嘌呤能受体，例如A2a或A2b腺苷受体是否以及如何引起定义极性和促进细胞迁移的全局抑制。重点将放在引起全球抑制的抑制性下游信号通路上。我们将集中于腺苷酸环化酶的激活、PKA和cAMP的积累。此外，我们还将研究这些信号事件如何阻止梯度感知，并诱导Rho激活和肌球蛋白形成，从而在后退边缘诱导尾足收缩。 对这些机制的了解将进一步确定控制趋化性的嘌呤能机制，并可能使我们能够构思新的治疗策略，旨在治疗这些嘌呤能反应，以治疗炎症和感染性疾病。

Chemotaxis allows polymorphonuclear neutrophils (PMN) to rapidly reach infected and inflamed sites. Despite recent progress in our understanding of chemotaxis, many open questions still remain. Chemoattractants stimulate the release of ATP from PMN and autocrine feedback loops via purinergic receptors control chemotaxis. In this project we revised the the scope of the proposed work to understand how the chemotactic signals induce ATP release and how adenosine formed from the released ATP controls negative feedback loops that play a critical role in defining cell polarity and uropod retraction. The proposed project rests on the following revised working hypothesis: Stimulation of chemoattractant receptors induces rapid intracellular events that result in ATP release. Additional ATP release through distinct mechanisms provides the ligand, adenosine, for suppressive A2a adenosine receptors. These receptors increases cAMP levels at the receding edge, providing an inhibitory feedback loop that elicits global inhibition and promotes uropod contraction, defining cell polarity and promoting migration. We propose to test this working hypothesis in by addressing the following Revised Specific Aims: Specific Aim 1. Upstream events leading to ATP release We will determine the upstream signaling pathways that link chemotactic receptor activation and ATP release during gradient sensing. In addition, we will examine different ATP release mechanisms and their contributions to ATP release during gradient sensing, cell polarization, and migration. Emphasis will be placed on the signaling pathway downstream of chemotactic receptors e.g., calcium signaling and MAPK activation that lead to the opening of ATP release channels such as connexins or pannexins and release of ATP via vesicular transport. Specific Aim 2. Global inhibition and purinergic signaling We will study if and how purinergic receptors, e.g., A2a or A2b adenosine receptors induce global inhibition that defines polarity and promotes cell migration. Emphasis will be placed on the suppressive down-stream signaling pathways that elicit global inhibition. We will focus on activation of adenylate cyclases, PKA, and cAMP accumulation. In addition, we will examine how these signaling events block gradient sensing and induce Rho activation and myosin formation that induces uropod contraction at the receding edge. Knowledge of these mechanisms will further define the purinergic mechanisms that control chemotaxis and may allow us to conceive novel therapeutic strategies aimed at treating these purinergic responses to treat inflammatory and infectious diseases.