Elucidating Key Mechanisms Regulating Cell Invasion In Vivo

描述(申请人提供)：基底膜是一种致密的、高度交联的细胞外基质形式，包围着大多数组织。在发育和免疫监视过程中，特化细胞获得了突破基底膜扩散并运输到感染和损伤部位的能力。在许多疾病中，包括哮喘、关节炎、妊娠障碍、先兆子痫和癌症，细胞侵袭计划也被增选或调节不当。因此，了解细胞如何通过基底膜侵袭对人类健康具有重要意义。细胞侵袭涉及侵袭细胞、被侵袭组织和分隔它们的基底膜之间的动态相互作用。由于无法在体外概括这些复杂的相互作用，以及在体内实验检测侵袭的挑战，细胞侵袭行为的关键机制仍然知之甚少。线虫锚定细胞入侵是一种实验上可获得的体内细胞入侵模型，它独特地将单细胞视觉分析与强大的遗传和基因组方法相结合。利用这些优势，我们已经确定了两个保守的转录因子，它们调控着获得侵袭性细胞命运的不同步骤。NHR-67，一个直系同源的 脊椎动物的无尾蛋白，维持锚细胞在有丝分裂后的状态。退出细胞周期似乎是必要的，然后允许线虫Fos家族转录因子同源FOS-1A启动入侵程序。初步数据表明，FOS-1A调节锚定细胞中三种基质金属蛋白酶(MMPs)的表达，暗示了一条控制基底膜去除的特定途径。最后，我们发现，在大多数晚期癌症中过度表达的细胞外基质蛋白SPARC的水平升高，降低了基底膜中的IV型胶原水平，并显著增强了锚定细胞的侵袭。这项建议的目标是使用活细胞成像结合遗传学和分子分析来确定：(1)NHR-67如何维持有丝分裂后状态的锚定细胞并允许细胞侵袭程序启动，(2)FOS-1A在调节MMP表达中的作用以及MMPs在破坏BM中的功能，(3)SPARC在增强细胞侵袭中的作用。这些研究与NIH的使命相关，因为它们将导致对细胞周期退出对侵袭的重要性、MMPs在Breachin基底膜中的特定作用以及SPARC在促进侵袭过程中的作用的新见解，从而使开发出更好的治疗策略来限制癌症等人类疾病的侵袭行为。 公共卫生相关性：通过基底膜的细胞侵袭在发育和免疫细胞运输中的许多细胞迁移中起着关键作用。这一行为也出现在许多人类疾病中，最明显的是在癌症扩散期间。我们拟议的工作将阐明特定的遗传程序的功能，该程序关闭细胞增殖并允许细胞入侵发生，以及促进侵袭细胞穿透基底膜屏障的能力的关键途径。这些研究将促进我们对细胞侵袭的基本机制的理解，并产生新的治疗策略来调节癌症等人类疾病的细胞侵袭行为。

DESCRIPTION (provided by applicant): Basement membrane is a dense, highly cross-linked form of extracellular matrix that surrounds most tissues. During development and immune surveillance, specialized cells acquire the ability to breach basement membrane to disperse and traffic to sites of infection and injury. The cell invasion program is also co-opted or misregulate during many diseases, including asthma, arthritis, the pregnancy disorder pre-eclampsia, and cancer. Understanding how cells invade through basement membrane is thus of great importance to human health. Cell invasion involves dynamic interactions between the invading cell, the tissue being invaded, and the basement membrane separating them. Owing to an inability to recapitulate these complex interactions in vitro, and the challenge of experimentally examining invasion in vivo, the key mechanisms underlying cell invasive behavior remain poorly understood. Anchor cell invasion in C. elegans is an experimentally accessible in vivo model of cell invasion that uniquely combines single cell visual analysis with powerful genetic and genomic approaches. Using these strengths, we have identified two conserved transcription factors that regulate distinct steps in acquiring an invasive cell fate. NHR-67, an ortholog of the vertebrate Tailless protein, maintains the anchor cell in a post-mitotic state. Exit from the cell cycle appears necessary to then permit the C. elegans Fos family transcription factor ortholog FOS-1A to initiate the invasion program. Preliminary data indicate that FOS-1A regulates the expression of three matrix metalloproteinases (MMPs) in the anchor cell, implicating a specific pathway that controls basement membrane removal. Finally, we have found that elevated levels of the extracellular matrix protein SPARC, which is overexpressed in most advanced cancer malignancies, decreases type IV collagen levels in basement membrane, and dramatically enhances anchor cell invasion. The goal of this proposal is to use live-cell imaging with genetic and molecular analysis to determine: (1) How NHR-67 maintains the anchor cell in a post-mitotic state and allows the cell invasion program to initiate, (2) the role of FOS-1A in regulatin MMP expression and the function of MMPs in breaching the BM, (3) the role of SPARC in enhancing cell invasion. These studies are relevant to NIH's mission as they will lead to new insights into the importance of cell cycle exit for invasion, the specific role of MMPs in breachin basement membrane and the role of SPARC in facilitating the invasive process, thus allowing the development of better therapeutic strategies to limit invasive behavior in human diseases such as cancer. PUBLIC HEALTH RELEVANCE: Cell invasion through basement membrane plays pivotal roles in numerous cell migrations in development and immune cell trafficking. This behavior is also co-opted in many human diseases, most notably during the spread of cancer. Our proposed work will elucidate the function of a specific genetic program that shuts down cell proliferation t allow cell invasion to occur and a key pathway that promotes the ability of invasive cells to penetrate through basement membrane barriers. These studies will advance our understanding of the fundamental mechanisms underlying cell invasion and generate new therapeutic strategies to regulate cell invasive behavior in human diseases such as cancer.