Functional Analysis of Histone Demethylase Activity in Hypoxic Cancer Cells

项目总结(见说明)： 肿瘤内不受控制的细胞生长会导致缺氧，因为细胞的新陈代谢需求超过了肿瘤血管提供氧气和营养的能力。作为对低氧微环境的反应，细胞经历了大规模的转录重新编程以促进生存。低氧诱导转录因子(HIF)是低氧反应的主要调节因子，可诱导糖酵解基因、细胞迁移因子和血管生成因子的表达。HIF还诱导几种转录调节因子的表达，包括几种组蛋白去甲基酶，为低氧细胞延长或固定有利于生存的基因的表达提供了一种机制。其中一个组蛋白去甲基酶，JMJD2B，去甲基化组蛋白H3赖氨酸9(H3K9me3)，这是抑制染色质结构的关键标志。低氧诱导JMJD2B可能在激活基因表达促进肿瘤生长中起重要作用。与这一假设一致的是，强制下调JMJD2B的表达会减少肿瘤移植瘤的生长，并在卵巢癌中过度表达。在这项建议中，JMJD2B调节肿瘤生长的程度将通过肿瘤异种移植实验在体内进行测试，并通过分析JMJD2B表达导致的细胞增殖、侵袭和血管生成在体外进行测试(特定目标1)。随后的生化实验将确定低氧中特异性靶基因调控的机制，发现JMJD2B在低氧中调控的新基因，并表征对肿瘤发生表型重要的基因的关键通路的调控(特异性目标2)。本提案中描述的实验将建立JMJD2B用来调控肿瘤发生的机制，而 确定新的途径，以加强肿瘤治疗为目标。

PROJECT SUMMARY (See instructions): Uncontrolled cell growth within a tumor results in hypoxia as the metabolic needs of the cells exceed the ability of the tumor vasculature to provide oxygen and nutrients. In response to the hypoxic microenvironment, cells undergo a massive reprogramming of transcription to promote survival. The Hypoxia Inducible Transcription Factors (HIFs) are primary regulators of the hypoxic response and induce the expression of glycolytic genes, cell migration factors, and angiogenic factors. HIFs also induce expression of several transcriptional regulators, including several histone demethylases, providing a mechanism for the hypoxic cell to extend or fix the expression of pro-survival genes. One of these histone demethylases, JMJD2B, demethylates tri-methylated histone H3 lysine 9 (H3K9me3), a key marker of repressed chromatin structure. Hypoxic induction of JMJD2B may play an important role in activating gene expression to promote tumor growth. Consistent with this hypothesis, forced knock-down of JMJD2B expression reduces growth of tumor xenografts, and is overexpressed in ovarian cancers. In this proposal, the extent to which JMJD2B regulates tumor growth will be tested in vivo using tumor xenograft experiments and in vitro by assaying for cell proliferation, invasion, and angiogenesis as a result of JMJD2B expression (Specific Aim 1). Subsequent biochemical experiments will determine the mechanism of specific target gene regulation in hypoxia, identify new genes regulated by JMJD2B in hypoxia, and characterize the regulation of key pathways of genes important for the tumorigenic phenotype (Specific Aim 2). The experiments described in this proposal will establish the mechanisms utilized by JMJD2B to regulate tumorigenesis, while identifying new pathways to target for enhanced tumor therapies.