Receptor Trafficking and the Response to Shh

描述(申请人提供)：Sonic Hedgehog是一种形态基因。为了完成这一角色，它必须能够远离其合成来源的许多细胞直径通过反应组织。这一要求似乎与Shh的生物学特性相冲突；Shh既含有酰基又含有胆固醇部分，这严重限制了其在亲水性细胞间环境中的溶解性。虽然已经鉴定出一种功能性的、可溶的Shh的六聚体形式，但由Shh表达细胞调节的培养液中Shh的积累水平不足以在神经板外植体中引起反应。然而，当这些反应组织与Shh产生细胞接触时，在外植体中可以观察到长距离的Shh信号，表明Shh在反应组织的平面上被活跃地运输。它假设这种长距离运输部分是通过跨细胞作用介导的，这一点得到了果蝇的观察支持，即输出胆固醇相关的Shh Hedgehog(HH)需要一个专门的分子DISP(DISP)的活性，而Shh和HH的摄取是由DISP同源补丁(PTC)介导的。此外，Ptc1介导的Shh转运到晚期内吞体内是启动Shh反应所必需的。为了在这些反应细胞中完成跨细胞反应，Shh必须从晚期的内体运输到细胞表面，并提供给邻近细胞，并假设这是由DISP介导的。将对这些假设进行测试，以确定Ptc1介导和Disp1介导的Shh运输是否需要协同进行远程信号传递。确定可替代锚定形式的Shh的远程信号有效性。将确定晚内体中的酸性环境是否有利于Shh结合到调度，而细胞间隙中的中性环境允许Shh与Ptc1结合，从而产生一系列导致细胞转运的连续事件。将通过阻断RABS和Dynamin的功能来测试产生远程信号时对细胞内运输的要求。Shh反应与跨细胞反应相结合，允许在积极响应Shh的组织中产生分级Shh反应，而不需要在细胞间隙中进行长距离运输，并为Ptc1和Disp1的异常活动提供了解释。更好地理解长程Shh信号形成的机制是很重要的。Shh反应的不适当激活是多种癌症的原因，包括高比例的胰腺肿瘤，而正常的Shh信号是许多器官系统发育所必需的。

DESCRIPTION (provided by applicant): Sonic Hedgehog is a morphogen. To fulfill this role, it is necessary that it is able to travel many cell diameters away from its source of synthesis through the responding tissue. This requirement appears to be in conflict with the biological properties of Shh; it both contains an Acyl and a cholesterol moiety, which severely curtail its solubility in a hydrophilic intercellular environment. Although a functional, soluble hexameric form of Shh has been identified, medium conditioned by Shh expressing cells do not accumulate Shh to levels high enough to elicit a response in neural plate explants. However, when such responsive tissue is in contact with Shh producing cells, long-range Shh signaling can be observed in the explant, indicating that Shh is actively transported in the plane of the responding tissue. It is hypothesized that this long-range transport is mediated in part by transcytosis, which is supported by observation in Drosophila that export of cholesterol-associated Shh homolog Hedgehog (Hh) requires the activity of a dedicated molecule, Dispatched (Disp), while the uptake of Shh and Hh is mediated by the Disp homolog Patched (Ptc). In addition, the Ptc1-mediated trafficking of Shh into late endosomes is required to initiate the Shh response. To complete transcytosis in these responding cell it is necessary that Shh is transported from the late endosomes to the cell surface and made available to neighboring cells, and it is hypothesized that this is mediated by Disp. The hypotheses will be tested determining if Ptc1-mediated and Disp1-mediated trafficking of Shh are required in concert for long-range signaling. Determine the long-range signaling efficacy of alternatively anchored forms of Shh. It will be determined if the acidic environment in late endosomes favors Shh binding to Dispatched, while the neutral environment in the intercellular space allows Shh to bind to Ptc1, creating a continuous chain of events resulting in transcytosis. The requirement for intracellular transport in generating a long-range signal will be tested by blocking the function of Rabs and Dynamin. The Shh response coupled to transcytosis allows the generation of a graded Shh response, in a tissue actively responding to Shh, without the requirement for long-range transport in the intercellular space, and provides an explanation for the unusual activities of Ptc1 and Disp1. Better understanding of the mechanism by which the long-range Shh signal forms is important. Inappropriate activation of the Shh response is the cause of a wide variety of cancers, including a high fraction of pancreatic tumors, while normal Shh signaling is required for the development of many organ systems.