The role of HMGN2 in enhancing transcription during prolactin receptor signaling

描述（由申请人提供）：乳腺癌是女性最常见的癌症，也是女性癌症死亡的第二大原因。肽激素催乳素（PRL）促进正常乳腺组织的生长和成熟，但PRL也有助于乳腺癌的发展。本申请的长期目标是确定PRL信号传导的分子介质，其可以作为新疗法的靶点。PRL通过与跨膜PRL受体（PRLr）结合来发出信号。PRLr以两种方式从细胞表面向细胞核发出信号，1）通过激活经典信号传导途径和2）通过易位到细胞核。在细胞核中，PRLr促进基因表达，但其机制尚未完全阐明。我们已经表明，核PRL r结合染色质修饰蛋白高迁移率族N2（HMGN 2），这也是重要的促进PRL反应基因的表达。HMGN 2可以介导染色质解压缩，其可以通过允许转录机器接近启动子DNA来促进基因表达。与介导染色质解压缩一致，HMGN 2已显示促进组蛋白乙酰化和染色质凝聚接头组蛋白H1的置换。所附的目的将确定HMGN 2促进PRL应答基因转录的机制。将通过染色质免疫沉淀（ChIP）研究HMGN 2介导的PRL应答基因启动子改变的程度，包括组蛋白乙酰化和H1置换。HMGN 2促进这种变化的机制，如通过招募组蛋白乙酰转移酶，也将进行研究。此外，所附的目标将确定如何调节HMGN 2活性。HMGN 2似乎受翻译后乙酰化调控。将分析模拟或阻止此类修饰的HMGN 2突变体介导染色质解压缩的能力，将使用ChIP研究其以鉴定基因启动子处的相关组蛋白修饰。还将通过免疫共沉淀评估乙酰化HMGN 2结合PRLr的能力。这项研究的基本原理是，通过确定HMGN 2如何促进转录以及如何调节该过程，各种蛋白质和过程将被用作新型乳腺癌治疗剂的靶点，以减弱PRLr信号传导。

DESCRIPTION (provided by applicant): Breast cancer is the most common cancer in women and the second leading cause of cancer death in women. The peptide hormone prolactin (PRL) promotes normal breast tissue growth and maturation, but PRL also contributes to breast cancer development. The long-term goal of this application is to determine molecular mediators of PRL signaling which can serve as targets for novel therapeutics. PRL signals by binding to the transmembrane PRL receptor (PRLr). The PRLr signals from the cell surface to the nucleus in two ways, 1) by activating canonical signaling pathways and 2) by translocating to the nucleus. In the nucleus, the PRLr promotes gene expression, but the mechanism by which it does so has not been fully elucidated. We have shown that nuclear PRLr binds to the chromatin-modifying protein high-mobility group N2 (HMGN2), which is also important in promoting the expression of PRL-responsive genes. HMGN2 can mediate chromatin decompaction, which may promote gene expression by allowing the transcriptional machinery to access the promoter DNA. Consistent with mediating chromatin decompaction, HMGN2 has been shown to promote histone acetylation and displacement of the chromatin-condensing linker histone H1. The enclosed aims will determine the mechanism by which HMGN2 promotes the transcription of PRL- responsive genes. The extent of HMGN2-mediated alterations at the promoters of PRL-responsive genes, including histone acetylation and H1 displacement, will be investigated by chromatin immunoprecipitation (ChIP). The mechanism by which HMGN2 promotes such changes, such as by recruiting histone acetyltransferases, will also be investigated. In addition, the enclosed aims will determine how HMGN2 activity is regulated. HMGN2 appears to be regulated by post-translational acetylation. HMGN2 mutants mimicking or preventing such modification will be analyzed for their ability to mediate chromatin decompaction, which will be investigated using ChIP to identify relevant histone modifications at gene promoters. The ability of acetylated HMGN2 to bind to the PRLr will also be assessed by co-immunoprecipitation. The rationale for the proposed research is that by determining how HMGN2 promotes transcription and how that process is regulated, various proteins and processes will be implicated as targets for novel breast cancer therapeutics to attenuate PRLr signaling.