RE: Regulation of mammary branching morphogenesis by Slit/Robo 1 signaling

描述(申请人提供)：组织生物学的一个核心目标是了解细胞如何相互作用来产生组织和器官。虽然在定义介导细胞相互作用的信号通路方面取得了很大进展，但关于这些信号系统在哺乳动物器官发生过程中是如何整合的，我们的知识还存在一个根本的缺口。弥合这一知识差距对于确定癌症的可用药靶点非常重要，因为肿瘤的生长和进展以组织结构的紊乱和特殊细胞群体(如干细胞)的放松监管为标志。这项应用的目标是了解细胞外因子如何通过调节干细胞所在的单层基底细胞的增殖来影响组织形态发生。本研究以乳腺分支形态发生为模型系统，在强大的初步数据的指导下，研究了乳腺分支的关键抑制因子--转化生长因子-21通过Sit/Robo1途径上调信号，抑制基底细胞的增殖和黏附。这反过来又通过增加2-连环蛋白在细胞膜上的黏附功能来控制基底细胞的生长和分支形态的形成，而牺牲了它在细胞核中的增殖功能。将使用各种细胞生物学、成像和生化技术，以及强大的移植技术来操纵转基因乳腺组织。三个假说在三个目标中得到验证：i)转化生长因子-21上调Robo1的表达，特别是在基底细胞中，这通过抑制基底细胞的生长来抑制分支；ii)Sit/Robo1信号通过改变2-catenin的亚细胞定位并促进其细胞黏附功能来对抗规范的WNTs的作用，从而以其转录功能为代价；以及iii)Sits是干细胞的非更新因子，通过调节2-catenin来对抗规范的WNT信号的自我更新信号。总之，这些实验将对包含基本干细胞亚群(如前列腺、唾液等)的乳腺和其他腺体器官的癌症生物学产生积极影响，因为它们描绘了一个似乎在干细胞中发挥作用的新的肿瘤抑制信号网络。

DESCRIPTION (provided by applicant): A core objective of tissue biology is to understand how cells interact to generate tissues and organs. While great strides have been made in defining signaling pathways that mediate cellular interactions, there is a fundamental gap in our knowledge concerning how these signaling systems are integrated during mammalian organogenesis. Bridging this knowledge gap is important in identifying druggable targets for cancer, as tumor growth and progression is marked by disorganization of tissue structure, and deregulation of specialized cell populations, such as stem cells. The goal of this application is to understand how extracellular factors, SLITs, influence tissue morphogenesis by regulating the proliferation of a single layer of basal cells in which stem cells reside. Here, mammary branching morphogenesis is used as a model system and, guided by strong preliminary data, the following model is investigated in which a key inhibitor of mammary branching, TGF-21, restrains basal cell proliferation and adhesion by upregulating signaling through the SLIT/ROBO1 pathway. This, in turn, controls basal cell growth and branching morphogenesis by increasing the adhesive functions of 2-catenin at the membrane, at the expense of its proliferative functions in the nucleus. A variety of cell biological, imaging and biochemical techniques will be employed, as well as powerful transplant techniques to manipulate genetically modified mammary tissue. Three hypotheses are tested in three Aims: I) that TGF-21 upregulates Robo1 expression, specifically in basal cells, and that this inhibits branching by restraining basal cell growth; II) that SLIT/ROBO1 signaling opposes the actions of canonical WNTs by altering the subcellular localization of 2-catenin and promoting its cell adhesive functions at the expense of its transcriptional functions; and III), that SLITs are non-renewal factors for stem cells that function to counter the self-renewal signals of canonical WNT signaling by regulating 2-catenin. Together, these experiments will have a positive impact on cancer biology of breast and other glandular organs containing basal stem cell subpopulations (e.g. prostate, salivary etc), because they delineate a novel tumor suppressive signaling network that appears to function in stem cells.