The role of HMGN2 in enhancing transcription during prolactin receptor signaling

描述(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。多肽荷尔蒙催乳素(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。这项应用的长期目标是确定可作为新疗法靶点的PRL信号的分子介体。通过与跨膜型PRL受体(PRLr)结合来传递PRL信号。PRLr信号通过两种方式从细胞表面传递到细胞核，1)激活典型的信号通路，2)移位到细胞核。在细胞核中，PRLr促进基因表达，但其机制尚未完全阐明。我们已经证明，核PRLr与染色质修饰蛋白高迁移率组N2(HMGN2)结合，这在促进PRL反应基因的表达方面也是重要的。HMGN2可以介导染色质的解压，这可能通过允许转录机制访问启动子DNA来促进基因的表达。与介导染色质分解一致，HMGN2已被证明促进组蛋白乙酰化和染色质浓缩连接物组蛋白H1的置换。所附的AIMS将确定HMGN2促进PRL反应基因转录的机制。HMGN2介导的PRL反应基因启动子的改变程度，包括组蛋白乙酰化和H1置换，将通过染色质免疫沉淀(ChIP)进行研究。HMGN2促进这种变化的机制，例如通过招募组蛋白乙酰转移酶，也将被研究。此外，所附的AIMS将决定如何监管HMGN2的活动。HMGN2似乎受翻译后乙酰化的调节。模拟或阻止这种修饰的HMGN2突变体将被分析它们介导染色质分解的能力，这将被用芯片来研究，以确定基因启动子上的相关组蛋白修饰。乙酰化的HMGN2与PRLr结合的能力也将通过免疫共沉淀进行评估。这项拟议研究的基本原理是，通过确定HMGN2如何促进转录以及如何调控这一过程，各种蛋白质和过程将被牵连为新的乳腺癌治疗药物的靶点，以减弱PRLr信号。

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.