Excitable Networks in Directed Cell Migration

定向细胞迁移中的兴奋网络

基本信息

批准号：
10187811
负责人：
Peter N Devreotes
金额：
$ 108.08万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-05-01 至 2026-04-30
项目状态：
未结题

项目摘要

We are investigating molecular mechanisms of directed cell migration, a critical process in health and disease, using Dictyostelium as a discovery tool to inform our studies of neutrophils, macrophages, and epithelial cells. At the core of our working model are coupled Signal Transduction and Cytoskeletal Excitable Networks, referred to as STEN and CEN which drive motility. The STEN integrates inputs from directional sensing and polarity networks to bring about directed migration. In the last grant period, we found that protrusions are governed by waves of coupled STEN-CEN activities and that manipulation of negatively charged lipids on the inner face of the membrane can alter network excitability and control cell behavior. The STEN-CEN concept is conserved in mammalian cells and the networks are hyperactivated in transformed cells, augmenting motility and macropinocytosis. Since these processes require geranyl geranylation, statins cause starvation of cancer cells. Finally, we found that vesicles internalized from retracting protrusions carry “back” components to the rear of the cell contributing to polarity. How do diverse cellular protrusions depend on the setpoint/threshold of STEN-CEN? We are combining imaging, synthetic biological, and computational studies to prove that pseudopods, lamellipods, forming phagosomes, and so on are closely related on a spectrum and interconvertible. We will show that spatiotemporal patterns of activities and responses to acute molecular perturbations are consistent across these protrusions and parallel those established in propagating STEN-CEN waves. What explains the extraordinary coordination of activities in STEN and CEN? Surmising that charge on the inner leaflet of the membrane is an organizer, we are 1) designing methods to directly monitor charge in local regions; 2) determining how anionic lipids transiently decrease; 3) manipulating charge locally with optogenetic systems; 4) examining how the location of key proteins is regulated by surface charge. Is lowered STEN threshold a general property of transformed cells and can it be exploited? To address this question, we are 1) comparing threshold indicators, such as propagating waves, with macropinocytosis and statin sensitivity in a series of increasingly metastatic cell lines and organoids; 2) identifying the essential geranylgeranylated proteins in STEN; 3) genetically engineering cells to increase threshold to normalize cancer cells or further decrease threshold to induce cell death. How does control of STEN and CEN at the cell poles mediate directional sensing and polarity? First, using a novel suppression assay, we are screening kinase and substrate deficient cells to identify global inhibitors. Second, we are studying membrane flow in a variety conditions to pursue our “reverse fountain” model and it reconcile with alternate models that argue membrane flows from front to back. 1

我们正在调查董事迁移董事的分子机制，这是健康的关键过程，疾病，使用dictyostelium作为发现工具，以告知我们对中性粒细胞，巨噬细胞和我们工作模型的核心的上皮细胞是耦合的信号转导和细胞骨架可兴奋的网络，称为sten和cen，驱动运动。方向性的感觉和极性网络在上一个赠款期间引导迁移。发现突出受到挥发的sten-cen活动的浪潮，并操纵膜内表面的负电荷脂质可以改变兴奋性并控制细胞细胞细胞细胞行为。在转化的细胞中，增加运动性和大细胞增多症。汀类药物最终引起癌细胞的饥饿。缩回的突起载有“背部”成分，导致细胞的后部产生极性。各种细胞突起如何取决于我们是Sten-cen的设定点/阈值将成像，合成生物学和计算研究结合给伪足的证明，薄片，形成吞噬体等等，与我们的频谱密切相关将显示激活性急性分子扰动的变形模式是在散布sten-cen波中建立的突出之间保持一致。是什么解释了Sten和CEN活动的内部协调？膜的内部小叶是一个组织者，我们是1）设计直接监控电荷的方法在局部区域；使用光遗传学系统； 4）检查关键蛋白的位置是如何通过表面电荷定制的。降低sten阈值是转化的细胞的一般特性吗？这个问题，我们是1）组成阈值指标，例如传播波，一系列越来越多的转移性细胞系和器官中的大型细胞增多症和他汀类药物的敏感性；识别Sten中必需的黄烷基质基化蛋白）阈值使癌细胞归一化或进一步降低损伤以诱导细胞死亡。细胞杆处的sten和CEN的控制如何介导定向感和极性？使用新型的支持分析，我们是筛选酶和底物缺乏细胞来识别全局抑制剂。第二，我们正在研究各种条件下的膜流程喷泉“模型和它的替代型号进行重新介绍，这些模型会争论膜从前到后面。 1