Understanding how cells invade through basement membrane in vivo

项目摘要 基底膜（BM）是包围大多数组织的致密的片状细胞外基质。期间 在发育和免疫细胞运输的过程中，特化细胞获得了突破BM屏障的独特能力， 分散、构建组织并迁移到感染和损伤部位。细胞入侵也是不适当的 在许多疾病中引发，并在哮喘，中风，关节炎，多发性硬化症， 硬化症和转移性癌症。因此，了解细胞如何穿越BM屏障是至关重要的 对改善人类健康的重要性。细胞侵袭事件通常是随机的、快速的，并且涉及动态的 侵袭细胞、BM和邻近组织之间的粘附和通讯。由于这种 由于其复杂性，不可能用体外测定法忠实地概括细胞侵袭，并且一直很困难， 来观察和从遗传学上剖析脊椎动物组织中的入侵。因此，细胞的潜在机制 人们对入侵行为仍然知之甚少。C.锚细胞的侵袭线虫是一种高度定型的体内 细胞侵袭模型，独特地结合了许多强大的实验方法，包括亚细胞 细胞-BM相互作用的可视化分析，分子活性传感器，快速基因组编辑，细胞类型特异性基因 操纵和强大的前向遗传和功能基因组方法。利用这些优势， 这项研究将描述入侵细胞如何获得和使用能量来为BM入侵提供燃料。这项工作将揭示 指导极化葡萄糖输入的机制和专门的电子传递链的构建 富含线粒体，提供局部ATP以驱动BM破坏机制。此外，概述了 实验将确定脂质生物合成是如何整合到一个保守的细胞侵入转录 程序以及在大多数转移性癌症中过度表达的脂质产生酶如何建立一个大的， 短暂的侵入性突出，打开了穿过BM屏障的路径。拟议的研究还将阐明如何 侵袭性细胞通过物理方式使其侵袭程序适应基质金属蛋白酶（MMPs）的缺乏， 取代BM，这将为更有效的方法提供信息，以阻止MMP抑制剂的侵袭， 迄今为止在临床试验中未能有效。最后，这项工作将确定分子机制， 并在BM破坏过程中修复质膜损伤，从而揭示了可以利用的机制 在入侵过程中瞄准细胞。这些综合性研究涵盖了细胞能量学、细胞外基质， 转录调控和膜动力学与NIH的使命相关，因为它们将导致 更深入地了解细胞侵入行为的基本生物学过程，从而允许 开发更好的治疗策略来调节人类疾病的侵袭。

PROJECT SUMMARY Basement membrane (BM) is a dense, sheet-like extracellular matrix that surrounds most tissues. During development and immune cell trafficking, specialized cells acquire the unique ability to breach BM barriers to disperse, construct tissues, and migrate to sites of infection and injury. Cell invasion is also inappropriately initiated during numerous diseases and underlies tissue destruction in asthma, stroke, arthritis, multiple sclerosis, and metastatic cancer. Understanding how cells traverse BM barriers is thus of fundamental importance in improving human health. Cell invasion events are often stochastic, rapid, and involve dynamic adhesions and communication between the invading cell, the BM, and the neighboring tissues. Owing to this complexity, it is not possible to faithfully recapitulate cell invasion with in vitro assays, and it has been difficult to visualize and genetically dissect invasion in vertebrate tissues. As a result, the mechanisms underlying cell invasive behavior remain poorly understood. Anchor cell invasion in C. elegans is a highly stereotyped in vivo model of cell invasion that uniquely combines many powerful experimental approaches including subcellular visual analysis of cell-BM interactions, molecular activity sensors, rapid genome editing, cell-type specific gene manipulation, and powerful forward genetic and functional genomic approaches. Using these strengths, this study will characterize how invading cells acquire and use energy to fuel BM invasion. This work will reveal mechanisms that direct polarized glucose import and the construction of specialized electron transport chain enriched mitochondria that provide localized ATP to power the BM breaching machinery. Further, the outlined experiments will determine how lipid biosynthesis is integrated into a conserved cell invasion transcriptional program and how lipid producing enzymes, which are overexpressed in most metastatic cancers, build a large, transient, invasive protrusion that opens paths through BM barriers. The proposed study will also elucidate how invasive cells adapt their invasion program to the absence of matrix metalloproteinases (MMPs) by physically displacing the BM, which will inform more effective approaches to block invasion with MMP inhibitors that have thus far failed to be effective in clinical trials. Finally, this work will identify molecular mechanisms that prevent and heal plasma membrane damage during BM breaching, thus revealing mechanisms that could be exploited to target cells in the act of invading. These integrative studies spanning cellular energetics, extracellular matrix, transcriptional regulation, and membrane dynamics are relevant to the NIH’s mission as they will lead to a deeper understanding of the fundamental biological process of cell invasive behavior, thus allowing for the development of better therapeutic strategies to modulate invasion in human disease.