Genetic Dissection of Context-Dependent and Opposing Roles of Androgen Receptor

雄激素受体的背景依赖性和对立作用的基因剖析

• 批准号: 8616350
• 负责人: DIANE M. ROBINS
• 金额: $ 29.51万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2016-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8616350
• 关键词: AR gene; Ablation; Affect; Age; Alleles; Androgen Receptor; Androgens; Bioinformatics; Biological; Biological Assay; Castration; Cell model; Cells; Combined Modality Therapy; Data; Development; Disease; Dissection; Engineering; Epithelial; Epithelium; Event; Future; Gene Expression; Gene Expression Regulation; Gene Targeting; Genetic; Genetic Polymorphism; Genetic Recombination; Genetic Variation; Genomics; Germ Lines; Glutamine; Growth; Health; High-Throughput Nucleotide Sequencing; Hormonal; Hormones; Human; In Vitro; Length; Lentivirus Vector; Ligands; Malignant Neoplasms; Malignant neoplasm of prostate; Maps; Methods; Modeling; Molecular; Molecular Chaperones; Molecular Genetics; Mus; Mutation; N-terminal; Nature; Normal Cell; Nuclear; Oncogenes; Oncogenic; Outcome; PTEN gene; Pathologic; Pathology; Pathway interactions; Patients; Pattern; Phenotype; Physiology; Pre-Clinical Model; Prostate; Prostate Cancer therapy; RNA; RNA Sequences; Receptor Gene; Regulation; Reproduction; Resistance; Resistance development; Role; Sampling; Selection for Treatments; Series; Signal Pathway; Signal Transduction; Somatic Mutation; Staging; Steroids; Testing; Time; Transcript; Transgenes; Transgenic Mice; Transgenic Model; Transgenic Organisms; Tumor Expansion; VCaP; Variant; Xenograft procedure; base; cancer cell; cancer initiation; cancer risk; cancer therapy; cell growth; cell type; chromatin immunoprecipitation; combat; deep sequencing; genetic risk factor; high throughput screening; homologous recombination; human disease; in vivo; male; molecular marker; mouse model; mutant; neoplastic cell; new therapeutic target; next generation; novel; novel strategies; programs; promoter; public health relevance; receptor; receptor expression; receptor function; response; small hairpin RNA; tool; trait; tumor; tumor growth; tumor progression; tumor xenograft; tumorigenesis

DESCRIPTION (provided by applicant): The androgen receptor (AR) regulates male physiology and is central to the ontogeny and progression of prostate cancer. Understanding AR's role strongly impacts human health by identifying genetic risk factors and new therapeutic targets. In normal as well as pathologic cell growth, AR directs divergent actions, such as proliferation or differentiation, through gene targets that vary with promoter, cell type and stage. These context- dependent and opposing functions of AR are evident in prostate cancer where AR initially suppresses tumor development but ultimately promotes oncogenesis. AR's dual nature is highlighted by germ-line and somatic mutations of the AR gene. We hypothesize that mechanisms underlying differential gene regulation by variant ARs may reveal more effective ways to combat pathological actions of the wild type AR. In order to use genetic tools, we converted the mouse AR gene to the human sequence by homologous recombination. We first tested the effect of a polymorphic glutamine (Q) tract associated with male phenotypic variation. Q tract length in humanized AR mice impacts the androgen axis, and affects initiation, progression and therapy response of prostate cancer in a transgenic model. Somatic AR mutations in these tumors evince selection by treatment. Moreover, this was substantiated in a select set of human patients. Mutations chosen for further study affecting the N-terminal domain influence ligand-dependent and -independent activation. The mutant ARs use multiple mechanisms that ultimately alter target gene selectivity to promote disease. We will use a multipronged genetic and genomic analysis to define upstream and downstream mechanisms by which a subset of Q tract variants and mutant ARs ("the variants") confer differential gene regulation. Our aims will define variant AR activity in novel cell and mouse models and identify the downstream effectors and upstream activators of aberrant function by Next Generation deep sequencing. In Aim I, a small set of variant ARs will be stably expressed in normal and cancer cells to view their distinct biological programs in vitro and in xenograft tumors. In Aim II, pathologic activity of variant ARs will be tested in mice with prostate- targeted transgenes, in mice predisposed to cancer by deletion of one PTEN allele, and in mice with varying androgen axis strength. In Aim III, gene expression programs directed by the variant ARs will be defined by high-throughput sequencing of transcripts (RNA-tag-Seq) to identify direct and indirect AR targets. Chromatin immunoprecipitation and high-throughput sequencing (ChIP-Seq) will probe differential promoter recognition and highlight coregulators directing different biological outcomes. Differences shared amongst AR variants, in addition to unique actions, will highlight critical downstream targets and upstream regulators of wild type AR. Combined genetic and genomic study of these AR variants will reveal aspects of wild type AR function so that in future we may selectively block disease-promoting while retaining disease-inhibiting effects of AR in diverse contexts. These results will have immediate applications and broad significance for future studies.

描述(申请人提供)：雄激素受体(AR)调节男性生理，对前列腺癌的个体发生和发展至关重要。通过识别遗传风险因素和新的治疗靶点，了解AR的作用对人类健康有很大影响。在正常和病理细胞生长中，AR通过不同启动子、细胞类型和阶段不同的基因靶点来指导不同的行为，如增殖或分化。AR的这些上下文相关和相反的功能在前列腺癌中很明显，在前列腺癌中，AR最初抑制肿瘤的发展，但最终促进肿瘤的发生。AR基因的胚系和体细胞突变突出了AR的双重性。我们推测，不同ARs差异基因调控的机制可能揭示出更有效的方法来对抗野生型AR的病理行为。为了利用基因工具，我们通过同源重组将小鼠AR基因转化为人的序列。我们首先测试了与男性表型变异相关的多态谷氨酰胺(Q)链的影响。在转基因模型中，人源化AR小鼠的Q道长度影响雄激素轴，并影响前列腺癌的起始、进展和治疗反应。这些肿瘤中的体细胞AR突变表明是通过治疗而选择的。此外，这在一组精选的人类患者中得到了证实。选择用于进一步研究的影响N-末端结构域的突变会影响配体依赖和非配体依赖的激活。突变的ARs使用多种机制，最终改变目标基因的选择性，从而促进疾病。我们将使用多管齐下的遗传和基因组分析来定义上游和下游机制，通过这些机制，Q道变异体和突变ARs(“变异体”)的子集提供差异基因调控。我们的目标是在新的细胞和小鼠模型中定义不同的AR活性，并通过下一代深度测序识别异常功能的下游效应器和上游激活器。在目标I中，一小部分变异的ARs将在正常细胞和癌细胞中稳定表达，以查看它们在体外和异种移植瘤中的不同生物学程序。在AIM II中，将在前列腺靶向转基因小鼠、因缺失一个PTEN等位基因而易患癌症的小鼠以及雄激素轴强度不同的小鼠中测试不同ARs的病理活性。在AIM III中，将通过高通量转录本测序(RNA-Tag-Seq)来确定直接和间接AR靶标，从而定义由变异体ARs指导的基因表达程序。染色质免疫沉淀和高通量测序(CHIP-SEQ)将探测差异启动子识别，并突出共同调节因子指导不同的生物结果。AR变异体之间的差异，除了独特的作用外，还将突出野生型AR的关键下游目标和上游调控因子。对这些AR变异体的遗传学和基因组学联合研究将揭示野生型AR功能的一些方面，以便在未来我们可以选择性地阻止疾病的促进，同时在不同的背景下保留AR的疾病抑制作用。这些结果将对未来的研究具有直接的应用和广泛的意义。