CDHR5 tumor suppressor function in the intestine

钙粘蛋白相关家族成员5(CDHR5)是一种高表达的1型跨膜黏附分子。 在肠道上皮细胞中表达，但在结直肠癌(CRC)中几乎总是缺失。前途无量的CRC 化学预防药物5-ASA(美沙拉嗪)有效上调CDHR5的表达 化学预防活性和CDHR5的直接过表达降低了结直肠癌的成瘤潜能 已经被注射到老鼠体内的细胞。综上所述，这些结果表明CDHR5是一种肿瘤抑制因子 直觉。尽管如此，CDHR5作为肿瘤抑制因子的功能特性还没有得到 已定义。这表明，对CDHR5如何作为对抗儿童权利公约的障碍的认识存在很大差距。这 知识差距阻碍了涉及CDHR5上调的化学预防结直肠癌疗法的发展， 就像药物5-ASA一样。这项研究的长期目标是了解黏附分子的作用。 肠道中的CDHR5。这项提议的当前目标是直接调查CDHR5的性质 这使得它可以在肠道中抑制CRC。初步研究发现CDHR5定位于末端 覆盖在肠上皮细胞顶端表面的微绒毛。在这里，CDHR5形成了一个强大的胞外 与另一种钙粘附素CDHR2相互作用，将相邻的微绒毛物理连接在一起，将它们组织成 一种被称为肠道刷状边界的有序排列。CDHR5的过度表达导致显著的肿瘤效应 结直肠癌细胞，细胞极化形成长度较长的顶端微绒毛密度增加。这 向超极化状态的转变部分依赖于CDHR5的胞浆结构域。 对于这一提议，中心假设是CDHR5有助于肠道上皮细胞的完整性 以依赖于CDHR5细胞质结合伙伴及其受体的方式防止CRC的发生 细胞外黏附活性。这一假设将通过三个具体目标进行检验：1)确定 CDHR5的两种剪接异构体之间的功能相互作用。这一目标将探索发现，这两个 CDHR5的剪接异构体相互作用，形成针对顶端微绒毛的成熟功能复合体。2)识别 与CDHR5胞质结构域相关的因子。这一目标将采用一种创新的蛋白质纯化技术 直接鉴定CDHR5胞浆结构域的体内结合伙伴的方法。的作用 CDHR5结合伙伴的功能将通过肠细胞模型的基因敲除/基因敲除研究来探索。 3)确定降低结直肠癌细胞成瘤潜能所需的CDHR5的功能特性 移植到裸鼠模型中。这一目标将评估粘附力和/或 CDHR5需要胞质结合伙伴才能减少结直肠癌细胞的肿瘤形成。方法是 创新，因为它挑战了CDHR5在肠道中作为肿瘤抑制因子的现有观点 与β-连环蛋白相互作用和隔离。这项拟议的研究具有重要意义，因为CDHR5的丢失 与结直肠癌的发展高度相关，并预测癌症患者的生存不良。

Cadherin related family member 5 (CDHR5) is a type 1 transmembrane adhesion molecule that is highly expressed in the gut epithelium, but is almost always lost in colorectal cancer (CRC). The promising CRC chemopreventive drug 5-ASA (mesalazine) potently upregulates CDHR5 expression as part of its chemopreventive activity, and direct overexpression of CDHR5 reduces the tumor formation potential of CRC cells that have been injected into mice. Together, these results implicate CDHR5 as a tumor suppressor in the gut. Despite this, the functional properties of CDHR5 that allow it to act as a tumor suppressor have not been defined. This represents a significant gap in knowledge of how CDHR5 acts as a barrier against CRC. This knowledge gap prevents the development of chemopreventive CRC therapies involving upregulation of CDHR5, as seen with the drug 5-ASA. The long-term goal of this study is to understand the role of the adhesion molecule CDHR5 in the intestine. The current objective of this proposal is to directly investigate the properties of CDHR5 that allow it to suppress CRC in the gut. Preliminary studies discovered that CDHR5 localizes to the distal tips of microvilli that cover the apical surface of intestinal epithelial cells. Here, CDHR5 forms a strong extracellular interaction with another cadherin, CDHR2, to physically link neighboring microvilli together, organizing them into an ordered array known as the intestinal brush border. Overexpression of CDHR5 results in a striking effect on CRC cells, with cells polarizing to form an increased density of apical microvilli that are longer in length. This shift towards a ‘hyper-polarized’ state is partially dependent upon the intracellular cytoplasmic domain of CDHR5. For this proposal, the central hypothesis is that CDHR5 contributes to the integrity of the intestinal epithelium to guard against CRC development, in a manner dependent on CDHR5 cytoplasmic binding partners and its extracellular adhesion activity. This hypothesis will be tested through three specific aims: 1) Determine the functional interplay between the two splice isoforms of CDHR5. This aim will explore the discovery that the two splice isoforms of CDHR5 interact to form a mature functional complex that targets to apical microvilli. 2) Identify factors associated with the cytoplasmic domain of CDHR5. This aim will employ an innovative protein purification approach to directly identify the in vivo binding partners for the cytoplasmic domain of CDHR5. The role of binding partners in CDHR5 function will be explored using knockdown/knockout studies an enterocyte model. 3) Determine the functional properties of CDHR5 required to reduce the tumor formation potential of CRC cells that have been implanted in a nude mouse model. This aim will assess whether the adhesion capacity and/or cytoplasmic binding partners are required for CDHR5 to reduce tumor formation of CRC cells. The approach is innovative, since it challenges the existing viewpoint that CDHR5 functions as a tumor suppressor in the gut by interacting with and sequestering β-catenin. The proposed research is significant because loss of CDHR5 is highly correlated with development of CRC and predicts poor survival of cancer patients.