Hypoxia and long noncoding RNA

缺氧和长非编码RNA

• 批准号: 8883446
• 负责人: Nobuaki Kikyo
• 金额: $ 16.53万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8883446
• 关键词: 6-Phosphofructo-2-kinase; Antibodies; Automobile Driving; Binding; Binding Sites; Biological Markers; Breast Cancer Cell; Breast Cancer cell line; Cancer Patient; Cell Culture Techniques; Cell Proliferation; Cells; Chromatin; Chromatin Structure; Complex; DNA Binding; Deposition; Disadvantaged; Energy Metabolism; Enzymes; Family; Gene Expression Regulation; Gene Targeting; Genes; Genetic Transcription; Glucose; Glycolysis; Growth Factor; Health; Hexokinase 2; Histones; Hypoxia; Hypoxia Inducible Factor; Link; Maps; Messenger RNA; Metabolic; Metabolic Pathway; Molecular Chaperones; Names; Neoplasm Metastasis; Oxidative Phosphorylation; Pharmaceutical Preparations; Proteins; RNA Binding; RNA Sequences; RNA Splicing; Reactive Oxygen Species; Recruitment Activity; Regenerative Medicine; Regulation; Reporting; Resistance; Ribosomal RNA; Role; Signal Transduction; Testing; Transfer RNA; Translations; Undifferentiated; Untranslated RNA; Up-Regulation; Variant; angiogenesis; base; cancer cell; cancer initiation; cancer therapy; cancer type; chemotherapy; chromatin immunoprecipitation; chromatin protein; design; dimer; embryonic stem cell; genome-wide; glucose metabolism; human embryonic stem cell; hypoxia inducible factor 1; improved; innovation; knock-down; lactate dehydrogenase A; new therapeutic target; novel; outcome forecast; pluripotency; prevent; promoter; response; screening; targeted cancer therapy; therapeutic target; therapy resistant; transcription factor; transcriptome sequencing; tumor; tumor progression; vpr Genes

DESCRIPTION (provided by applicant): Cancer cells express hypoxia-inducible factors (HIFs) in hypovascularized regions inside a tumor mass in response to hypoxia. HIFs activate more than 800 genes including those involved in angiogenesis, glycolysis, and growth factor signaling, which collectively facilitate cancer progression and metastasis. Indeed, increased expression of HIFs is an indicator for poor prognosis of cancer patients. In addition, embryonic stem cells (ESCs) maintain an undifferentiated state more efficiently in hypoxia than in normoxia. Therefore, it is important to understand the functions and regulation of HIFs for cancer therapy and regenerative medicine. The current project will identify and characterize long noncoding RNAs (lncRNAs) that are bound to HIF-1 in human breast cancer cells and embryonic stem cells (ESCs) cultured in hypoxia. LncRNAs are defined as RNAs that are longer than 200 bases and do not encode mRNA, rRNA, or tRNA. Chromatin immunoprecipitation and RNA sequencing were used to identify novel HIF-1-bound lncRNAs (collectively called R-HIFs) that have been mapped closely to genes encoding glycolysis enzymes. Cancer cells and ESCs use glycolysis as a primary metabolic pathway of glucose rather than oxidative phosphorylation in normoxia and this tendency is enhanced by HIFs in hypoxia. Enhanced glycolysis has an advantage of producing more metabolic intermediates necessary for rapid cell proliferation. Thus, the regulation of glycolytic enzymes by lncRNAs can serve as a novel target for cancer therapy. Currently virtually nothing is known about the involvement of lncRNAs in hypoxia or glycolysis. It was hypothesized that HIF-1 uses lncRNAs as novel coactivators to regulate its target genes, including glycolysis genes, in hypoxia and thereby promotes proliferation of breast cancer cells and ESCs. The following 3 aims have been proposed to test this hypothesis. Aim 1 is designed to identify direct target genes of an R-HIF by screening of its DNA binding sites and by identification of genes whose expression level is altered by the knockdown of the R-HIF. In Aim 2 the roles of other R-HIFs in hypoxic gene regulation will be determined by the combination of knockdown, analysis of their genome-wide binding sites, and identification of associated chromatin proteins. In Aim 3 the roles of the R-HIFs in increased glycolysis and decreased oxidative phosphorylation in hypoxic cancer cells and ESCs will be determined through the analysis of energy and glucose metabolism. These studies are expected to establish lncRNAs as a novel entity essential for the regulation of glycolytic genes by HIF-1 in hypoxia.

描述（由申请人提供）：癌细胞响应缺氧而在肿瘤块内血管不足的区域表达缺氧诱导因子（HIF）。 HIF 可激活 800 多个基因，包括参与血管生成、糖酵解和生长因子信号传导的基因，这些基因共同促进癌症进展和转移。事实上，HIF 表达增加是癌症患者预后不良的一个指标。此外，胚胎干细胞（ESC）在缺氧条件下比在常氧条件下更有效地维持未分化状态。因此，了解 HIF 的功能和调节对于癌症治疗和再生医学非常重要。当前的项目将鉴定并表征在缺氧条件下培养的人类乳腺癌细胞和胚胎干细胞 (ESC) 中与 HIF-1 结合的长非编码 RNA (lncRNA)。 LncRNA 被定义为长度超过 200 个碱基且不编码 mRNA、rRNA 或 tRNA 的 RNA。使用染色质免疫沉淀和 RNA 测序来鉴定新型 HIF-1 结合 lncRNA（统称为 R-HIF），这些 lncRNA 与编码糖酵解酶的基因紧密定位。在常氧条件下，癌细胞和 ESC 使用糖酵解而不是氧化磷酸化作为葡萄糖的主要代谢途径，并且缺氧条件下的 HIF 会增强这种趋势。增强的糖酵解具有产生更多细胞快速增殖所需的代谢中间体的优点。因此，lncRNA对糖酵解酶的调节可以作为癌症治疗的新靶点。目前，对于 lncRNA 参与缺氧或糖酵解的情况几乎一无所知。据推测，HIF-1在缺氧条件下利用lncRNA作为新型共激活剂来调节其靶基因，包括糖酵解基因，从而促进乳腺癌细胞和ESC的增殖。为了检验这一假设，提出了以下 3 个目标。目标 1 旨在通过筛选 DNA 结合位点并鉴定表达水平因 R-HIF 敲低而改变的基因来鉴定 R-HIF 的直接靶基因。在目标 2 中，其他 R-HIF 在缺氧基因调控中的作用将通过组合敲低、分析其全基因组结合位点以及识别相关染色质蛋白来确定。在目标 3 中，将通过分析能量和葡萄糖代谢来确定 R-HIF 在缺氧癌细胞和 ESC 中糖酵解增加和氧化磷酸化减少中的作用。这些研究有望将 lncRNA 确立为缺氧条件下 HIF-1 调节糖酵解基因所必需的新实体。