ROLE OF MECHANICAL FORCE IN NOTCH

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 Notch途径是一个进化保守的信号系统，在整个胚胎发育过程中广泛使用，继续在成人的组织和干细胞维持中发挥作用。Noch信号通过对细胞分化、增殖、存活和死亡的影响，指导多种细胞类型的规范，促进组织构型和形态形成。我们关于配体诱导的Notch信号的工作模型是基于Weinmaster在哺乳动物细胞培养中发表的初步发现，并将配体内吞过程中产生的力的关键作用与Notch结构发现结合在一起。具体地说，我们假设配体结合的Notch的内吞作用产生了一个强大的机械力，它物理上将Notch受体拉开，暴露出ADAM裂解位点，促进蛋白分解。我们的模型还提出，通过呋喃裂解产生Notch异二聚体，使Notch易于通过配体内吞过程中产生的机械力进行物理解离。然而，也有人提出通过配体内吞作用来促进循环，以产生高亲和力的Notch配体。为了解决这一争议，我们正在使用光学镊子来确定配体内吞作用在调节结合强度和力产生中的作用。 SMM激光镊子用于测量Notch包被珠和配体表达细胞之间的内吞作用力和亲和力。寻求内吞力量、粘连强度和Notch横裂细胞之间的相关性。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The Notch pathway is an evolutionary conserved signaling system used extensively throughout embryonic development that continues to function in the maintenance of tissues and stem cells in adults. Notch signaling directs the specification of a variety of cell types and contributes to tissue patterning, and morphogenesis through effects on cellular differentiation, proliferation, survival and death. Our working model for ligand-induced Notch signaling is based on Weinmaster's published and preliminary findings obtained with mammalian cell culture, and incorporates the critical role for forces generated during ligand endocytosis with Notch structural findings. Specifically, we hypothesize that endocytosis of ligand-bound Notch generates a robust mechanical force that physically pulls the Notch receptor apart to expose the ADAM cleavage site and facilitate activating proteolysis. Our model also proposes that furin cleavage to produce the Notch heterodimer predisposes Notch to physical dissociation through mechanical force generated during ligand endocytosis. However, ligand endocytosis has also been proposed to facilitate recycling to generate a high affinity Notch ligand. To address this controversy, we are using optical tweezers to determine roles for ligand endocytosis in regulating binding strength and force generation. SMM laser tweezers used to measure endocytic forces and avidity between Notch coated beads and ligand expressing cells. Correlations between endocytic forces, adhesion strength and Notch transedocytoiss are sought.