Cellular Integration of Information in the Detection and Response to Epithelial Damage

项目摘要 当上皮组织受伤时，细胞的第一反应是胞浆急剧增加。 钙水平，从组织受损后立即开始。这种钙含量的增加不仅见于 细胞位于伤口边缘，但位于伤口周围的大片细胞区域。随着遗传基因的出现 像GCaMP一样编码的钙指示剂，这种创伤诱导的钙反应已经在生活中观察到 整个动物界的生物，然而，已经提出了相互矛盾的机制来解释 钙的诱导，以至于没有一种保守的基本过程的感觉。我们的协作团队 生物物理学家和发育遗传学家最近发表的工作确定了一个主要的潜在障碍： 创伤诱导的钙反应有几种同时发生的机制，我们 已经能够通过我们的高度定量方法和我们的基因组合来梳理 操纵。在不了解诱导钙反应的多种机制的情况下， 不可能在文学作品和受伤模型之间进行比较，也不可能完全 阻断钙反应，分析伤口愈合的下游后果。 我们的分析工具确定了具有不同振荡模式或 在离伤口不同的径向距离处有明显的特征。我们推测这些花纹钙 反应告知细胞与伤口的距离，并决定其下游细胞的行为。喜欢 钙反应、创伤诱导的细胞行为和转录特性是根据 与伤口的距离，边缘有游走细胞和JNK信号，有增殖细胞和 在更远端的环中发送JAK-STAT信号。在第一个目标中，我们研究了钙离子如何 信号在这些模式中的每一种模式中都被启动，既与单独的路径一起工作，也与发展 这些钙模式如何整合的数学模型。在第二个目标中，我们扰乱了特定的 了解钙模式的各个方面，并询问下游细胞的行为和特性是如何改变的。我们能够 实现这些目标是因为我们独特的协作技能，以及我们开发了 无与伦比的创伤模型，允许在一侧进行高时间控制的基因操作 只有伤口。因为它是内部控制的，所以比较两边可以精确量化和 检测到钙信号和创伤诱导的细胞行为的微小变化。 在这个项目完成时，我们希望已经生成了一个高精度的细胞模型 检测远距离的组织损伤，以及他们如何解释这些信息以选择空间上合适的 维修计划。这一基本知识对细胞生物学的许多领域、伤口愈合都很重要。 研究，以及像癌症这样的病理，伤口愈合计划被不适当地激活。

Project Summary When an epithelial tissue is wounded, the first cellular response is a dramatic increase in cytosolic calcium levels, beginning immediately upon tissue damaged. This calcium increase is observed not just in cells at the wound margin but in large domain of cells surrounding the wound. With the advent of genetically encoded calcium indicators like GCaMP, this wound-induced calcium response has been observed in living organisms across the animal kingdom, yet conflicting mechanisms have been proposed to explain the induction of calcium, such that there is no sense of a conserved fundamental process. Our collaborative team of biophysicists and developmental geneticists recently published work identifying a major underlying obstacle: there are several contemporaneous mechanisms underlying the wound-induced calcium response, which we have been able to tease apart with our combination of highly quantitative approaches and our genetic manipulations. Without understanding the multiple mechanisms inducing calcium responses, it has been impossible to draw parallels across the literature and across wounding models, and it has impossible to fully block calcium responses to analyze the downstream consequences for wound healing. Our analysis tools identified stereotyped calcium responses, with different oscillatory patterns or signatures evident at different radial distances from the wound. We hypothesize that these patterned calcium responses inform the cell about its distance from the wound and determine its downstream cell behaviors. Like the calcium responses, wound-induced cell behaviors and transcriptional identities are patterned according to distance from the wound, with migratory cells and JNK signaling near the margin and proliferative cells and JAK-STAT signaling in a more distal ring. In the first Aim, we investigate the mechanisms of how calcium signaling is initiated in each of these patterns, working both with individual pathways and developing mathematical models for how these calcium patterns are integrated. In the second Aim, we perturb specific aspects of calcium patterns and ask how downstream cell behavior and identity are altered. We are able to achieve these Aims because of our unique collaborative skill-set, and because we have developed an unparalleled wounding model that allows genetic manipulation with high temporal control on one side of the wound only. Because it is internally controlled, comparing the two sides allows precise quantification and detection of even small changes, in both calcium signaling and wound-induced cell behaviors. At the completion of this project, we expect to have generated a high-precision model of how cells detect tissue damage at a distance, and how they interpret this information to select a spatially-appropriate repair program. This fundamental knowledge will be important to many areas of cell biology, to wound-healing studies, and to pathologies like cancer where wound-healing programs are inappropriately activated.