NRDP1 PROTEIN DEGRADATION PATHWAY IN MAMMARY TUMOR PROGRESSION

描述（由申请方提供）：生长因子受体酪氨酸激酶ErbB家族成员的异常过表达和伴随激活在促进多种实体瘤类型的生长和进展中起关键作用。拟议研究的长期目标是了解一种新的蛋白质降解途径在调节ErbB3诱导的乳腺肿瘤发生和进展中的作用。该途径的中心组成部分是一种称为Nrdp1的环指E3泛素连接酶，它介导ErbB3受体的泛素化，从而促进其向降解细胞区室的运输。在ErbB2诱导的乳腺肿瘤中，Nrdp1蛋白总是在转录后受到抑制，这些肿瘤来自过表达内源性ErbB3的转基因小鼠，并且Nrdp1蛋白在几乎60%的乳腺癌患者的肿瘤中丢失，这与Nrdp1丢失促进ErbB3蛋白在肿瘤中过表达的模型一致。然而，最近的证据表明，Nrdp1及其相关的通路成分在细胞调控中发挥着更广泛的作用。指导拟议研究的假设是Nrdp1通过多种途径发挥作用以维持乳腺上皮细胞的完整性，并且Nrdp1蛋白的转录后丢失消除了乳腺肿瘤起始的障碍。三个关键问题，Nrdp1参与乳腺上皮完整性将得到解决。1)Nrdp1通路相关的去泛素化酶Otub1在肿瘤细胞中介导Nrdp1转录后丢失的作用将通过培养细胞中病毒介导的过表达和敲低来评估。此外，可诱导的转基因过表达的Otub1在乳腺将被用来评估其参与乳腺发育和ErbB2诱导的肿瘤发生。2)Nrdp1通路成分在建立和维持培养的上皮细胞的极性和结构中的作用将使用病毒介导的过表达和敲低方法结合生物化学测定和基于免疫荧光的细胞结构测定来探索。3)Nrdp1基因在抑制ErbB2诱导的肿瘤发生中的作用将使用条件性敲除和移植方法进行检查。总之，拟议的研究可以揭示该途径在乳腺肿瘤发展中的新活性，并强调其在乳腺肿瘤抑制中的作用。

DESCRIPTION (provided by applicant): The aberrant over expression and concomitant activation of members of the ErbB family of growth factor receptor tyrosine kinases plays key roles in promoting the growth and progression of a variety of solid tumor types. The long term objective of the proposed studies is to understand the role of a novel protein degradation pathway in regulating ErbB3-induced mammary tumor initiation and progression. The central component of the pathway is a RING finger E3 ubiquitin ligase called Nrdp1 that mediates the ubiquitination of the ErbB3 receptor, thereby promoting its trafficking to degradative cellular compartments. Nrdp1 protein is invariably post-transcription ally suppressed in ErbB2-induced mammary tumors from transgenic mice that over express endogenous ErbB3, and Nrdp1 protein is lost in tumors from almost 60% of breast cancer patients, consistent with a model whereby Nrdp1 loss facilitates ErbB3 protein over expression in tumors. However, recent evidence points to a broader role for Nrdp1 and its associated pathway components in cellular regulation. The hypothesis guiding the proposed studies is that Nrdp1 acts through multiple pathways to maintain mammary epithelial cell integrity, and that the post-transcriptional loss of Nrdp1 protein removes a barrier to mammary tumor initiation. Three critical issues concerning Nrdp1 involvement in mammary epithelial integrity will be addressed. 1) The role of the Nrdp1 pathway-associated deubiquitinating enzyme Otub1 in mediating Nrdp1 post-transcriptional loss in tumor cells will be assessed by viral-mediated over expression and knockdown in cultured cells. In addition, inducible transgenic over expression of Otub1 in the mammary gland will be used to assess its involvement in mammary gland development and ErbB2-induced tumorigenesis. 2) The role of Nrdp1 pathway components in establishing and maintaining the polarity and architecture of cultured epithelial cells will be explored using viral-mediated over expression and knockdown approaches coupled with biochemical assays and immunofluoresecence-based cell structure assays. 3) The role of the Nrdp1 gene in suppressing the onset of ErbB2-induced tumors will be examined using conditional knockout and transplantation approaches. Together, the proposed studies could uncover novel activities for this pathway in mammary tumor development, and underscore its role in mammary tumor suppression.