Understanding the Role of MMPs in Basement Membrane Breaching In vivo

了解 MMP 在体内基底膜破裂中的作用

基本信息

批准号：
8398457
负责人：
Laura Catherine Kelley
金额：
$ 4.92万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-12-01 至 2015-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8398457
关键词：
Animals Basement membrane Behavior Binding Biological Caenorhabditis elegans Cancer cell line Cell Culture Techniques Cell physiology Cells Cessation of life Complex Data Detection Development Developmental Process Disease Disease Progression Disseminated Malignant Neoplasm Drug Delivery Systems Encapsulated Enzymes Event Excision Extracellular Matrix Extracellular Matrix Degradation FOS Protein FOS gene Gene Expression Gene Expression Regulation Genes Genetic Genetic Screening Goals Health Homologous Gene Human Human Development Image Immune System Diseases Immunologic Surveillance In Vitro Infiltration Invaded Knowledge Laboratories Lead Leukocyte Trafficking Life Localized Malignant Neoplasm Malignant Neoplasms Matrix Metalloproteinases Mediating Membrane Proteins Metalloproteases Metalloproteinase Gene Methods Modeling Molecular Molecular Genetics Movement Neoplasm Metastasis Neural Crest Normal Cell Oncogenes Pathway interactions Patients Penetration Process Proteins RNA Interference Research Resolution Role Site Site-Directed Mutagenesis Specificity Stereotyping Time Tissues Transgenes Vertebrates Work Wound Healing Zinc cancer cell cardiogenesis cellular imaging crosslink gene function genetic analysis immune function improved in vivo matrix metalloproteinase 25 migration mutant natural Blastocyst Implantation novel outcome forecast overexpression programs promoter transcription factor tumor progression

项目摘要

DESCRIPTION (provided by applicant): Basement membrane is a dense sheet-like extracellular matrix that encapsulates and separates tissue compartments. The ability of cells to invade through basement membrane barriers is important during normal and disease processes. For example, basement membrane breaching is required for embryo implantation, neural crest migration, heart development, leukocyte trafficking and cancer cell metastasis-a critical step in tumor progression and the leading cause of patient death. Studying how cells breach basement membrane in vertebrates has been hindered by the difficulty of imaging and experimentally examining cell-basement membrane interactions in vivo. As a result, how cells cross basement membrane remains poorly understood. Our laboratory has developed methods that combine high-resolution live-cell imaging with the highly- stereotyped and genetically tractable model of anchor cell invasion in Caenorhabditis elegans to uncover the molecular mechanisms regulating basement membrane breaching in vivo. We have determined that the C. elegans c-fos oncogene homologue, fos-1a functions in the anchor cell to specifically mediate basement membrane penetration. In fos-1a mutant animals, the anchor cell extends cellular processes that flatten at an intact basement membrane. The complete repertoire of FOS-1A protein transcriptional targets mediating basement membrane breaching is not known. Recently, we have found that FOS-1A regulates the expression of three MMPs in the anchor cell during the time of invasion. Matrix metalloproteinases are overexpressed in cells responsible for tissue remodeling, wound healing, and cancer and are hypothesized to enzymatically facilitate BM removal. Matrix metalloproteinases localize to invasive machinery and are required for extracellular matrix degradation in metastatic cancer cell lines in vitro. Due to the high number of matrix metalloproteinases expressed in vertebrates and the difficulty of directly examining cell invasion in vivo, the relevance, and potential function of matrix metalloproteinases in cell invasion through basement membrane is unclear. The goal of the proposed research is to use the strengths of the model of anchor cell invasion in C. elegans - genetic analysis, live cell-imaging, molecular perturbation - to determine the function of matrix metalloproteinases during basement membrane breaching. In addition, I will perform a sensitized genetic screen to identify novel genes and pathways that function downstream of FOS-1A with matrix metalloproteinases to promote invasion. Completion of the aims in this proposal will increase our knowledge of the genetic pathways regulating cell invasion and the functional significance of FOS-1A-directed matrix metalloproteinase-driven basement membrane breaching. This work outlined in this proposal will directly impact human health by identifying specific pathways that could be targeted to limit invasive behavior. PUBLIC HEALTH RELEVANCE: Cell invasion through basement membrane is an important and fundamental process in human development, immune functioning and is a defining step in the lethal progression of most cancers. The proposed research will increase our understanding of the way in which invasive cells penetrate basement membrane barriers and invade into tissues. Completion of this work will lead to better strategies to therapeutically target invasive cellular behavior and directly improve patient prognosis in diseases such as cancer.

描述（由申请人提供）：地下膜是一种薄板状的细胞外基质，可封装并分离组织室。在正常和疾病过程中，细胞通过基底膜屏障侵袭的能力很重要。例如，胚胎植入，神经rest迁移，心脏发育，白细胞运输和癌细胞转移需要基底膜破坏，这是肿瘤进展的关键步骤和患者死亡的主要原因。研究细胞在体内的难度和实验检查细胞膜膜相互作用的困难和实验检查，研究细胞如何违反脊椎动物中的基底膜。结果，细胞如何穿越基底膜仍然了解不足。我们的实验室已经开发了将高分辨率的活细胞成像与秀丽隐杆线虫中锚定细胞侵袭的高分辨率和遗传障碍模型相结合的方法，以发现体内调节基底膜膜破坏的分子机制。我们已经确定秀丽隐杆线虫C-FOS癌基因同源物，FOS-1A在锚固细胞中起作用，以特别介导地下膜穿透。在FOS-1A突变动物中，锚固细胞延伸了在完整的地下膜上扁平的细胞过程。 FOS-1A蛋白转录靶标的完整曲目介导基底膜破坏。最近，我们发现FOS-1A在入侵期间调节了锚固细胞中三个MMP的表达。基质金属蛋白酶在负责组织重塑，伤口愈合和癌症的细胞中过表达，并假设酶促促进BM去除。基质金属蛋白酶本地化为浸润性机械，是在体外转移性癌细胞系中细胞外基质降解所必需的。由于脊椎动物中表达的基质金属蛋白酶的数量大量以及直接检查细胞的难度体内侵袭，基质金属蛋白酶通过基底膜在细胞侵袭中的相关性和潜在功能尚不清楚。拟议的研究的目的是在秀丽隐杆线虫中使用锚固细胞侵袭模型的强度 - 遗传分析，活细胞成像，分子扰动 - 来确定基底膜破坏过程中基质金属蛋白酶的功能。此外，我将执行一个敏化的遗传筛选，以鉴定新的基因和途径，这些基因和途径在FOS-1A下游与基质金属蛋白酶一起促进侵袭。该提案中的目标的完成将增加我们对调节细胞侵袭的遗传途径的了解，以及FOS-1A指导的基质金属蛋白酶驱动的基底膜泄漏的功能意义。该提案中概述的这项工作将直接通过确定可以限制侵入性行为的特定途径来直接影响人类健康。公共卫生相关性：通过基底膜的细胞侵袭是人类发展，免疫功能的重要且基本的过程，是大多数癌症致命进展的决定性一步。拟议的研究将增加我们对侵入性细胞穿透基底膜屏障并侵入组织的方式的理解。这项工作的完成将导致更好地靶向侵入性细胞行为并直接改善癌症等疾病的患者预后。