Elucidating Key Mechanisms Regulating Cell Invasion In Vivo

描述（由申请人提供）：基底膜是一种致密的、高度交联的细胞外基质，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的能力，以分散和运输到感染和损伤部位。在许多疾病中，包括哮喘、关节炎、妊娠期疾病先兆子痫和癌症，细胞入侵程序也被选择或失调。因此，了解细胞如何通过基底膜侵入对人类健康非常重要。细胞侵入涉及侵入细胞、被侵入组织和分隔它们的基底膜之间的动态相互作用。由于无法在体外重现这些复杂的相互作用，以及在体内实验研究入侵的挑战，细胞入侵行为的关键机制仍然知之甚少。C.锚细胞的侵袭elegans是一种实验上可获得的体内细胞入侵模型，其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，调节不同的步骤，在获得侵入性细胞的命运。NHR-67，一种 脊椎动物无尾蛋白，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C。线虫Fos家族转录因子直系同源物FOS-1A启动入侵程序。初步数据表明，FOS-1A调节锚细胞中三种基质金属蛋白酶（MMPs）的表达，暗示了控制基底膜去除的特定途径。最后，我们发现，细胞外基质蛋白的水平升高，在大多数晚期恶性肿瘤中过度表达，降低基底膜中的IV型胶原蛋白水平，并显著增强锚细胞侵袭。本研究的目的是利用活细胞成像技术结合遗传学和分子生物学分析来确定：（1）NHR-67如何维持锚细胞处于有丝分裂后状态并启动细胞侵袭程序;（2）FOS-1A在调节MMP表达中的作用以及MMP在破坏BM中的功能;（3）NHR-67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于入侵的重要性、MMPs在破坏基底膜中的特定作用以及MMP在促进入侵过程中的作用的新见解，从而允许开发更好的治疗策略以限制人类疾病（例如癌症）中的入侵行为。 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关键作用。这种行为也在许多人类疾病中被选择，最明显的是在癌症的传播过程中。我们提出的工作将阐明一个特定的基因程序的功能，关闭细胞增殖，让细胞入侵发生和一个关键的途径，促进入侵细胞穿透基底膜屏障的能力。这些研究将促进我们对细胞入侵基本机制的理解，并产生新的治疗策略来调节人类疾病（如癌症）中的细胞入侵行为。

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.