Receptor Trafficking and the Response to Shh

描述（由申请人提供）：Sonic Hedgehog是一种形态因子。为了完成这一角色，它必须能够从其合成源穿过响应组织行进许多细胞直径。这一要求似乎与Shh的生物学特性相冲突；它同时含有酰基和胆固醇部分，这严重限制了它在亲水性细胞间环境中的溶解度。虽然已经鉴定出一种功能性的、可溶的六聚体形式的Shh，但在Shh表达细胞的条件下，培养基不会积累足够高的Shh水平，从而在神经板外植体中引起反应。然而，当这些应答组织与产生Shh的细胞接触时，可以在外植体中观察到远程Shh信号，这表明Shh在应答组织的平面上被积极运输。据推测，这种远程转运部分是由胞吞作用介导的，在果蝇中观察到，胆固醇相关的Shh同源物Hedgehog （Hh）的输出需要一个专用分子的活性，分派（Disp），而Shh和Hh的摄取是由Disp同源物Patched （Ptc）介导的。此外，ptc1介导的Shh进入晚期核内体是启动Shh应答所必需的。为了在这些应答细胞中完成转胞作用，有必要将Shh从晚期核内体转运到细胞表面，并使其可被邻近细胞使用，并且假设这是由Disp介导的。这些假设将被测试，以确定ptc1介导和dis1介导的Shh运输是否需要协同进行远程信号传递。确定Shh替代性锚定形式的远程信号传导效力。这将决定是否晚期核内体的酸性环境有利于Shh与分派结合，而细胞间空间的中性环境允许Shh与Ptc1结合，从而产生导致胞吞的连续事件链。我们将通过阻断Rabs和Dynamin的功能来测试细胞内运输对产生远程信号的要求。Shh反应与胞裂作用相结合，允许在积极响应Shh的组织中产生分级的Shh反应，而不需要在细胞间空间进行远距离运输，并为Ptc1和Disp1的异常活动提供了解释。更好地了解远距离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，包括很大一部分胰腺肿瘤，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.