The Role of Centromeres in the Molecular Biology of Prostate Cancer

着丝粒在前列腺癌分子生物学中的作用

• 批准号: 9314219
• 负责人: Anjan Kumar Saha
• 金额: $ 3.91万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9314219
• 关键词: Affect; Aneuploidy; Area; Atlases; Automobile Driving; Base Pairing; Base Sequence; Benign; Binding; Biochemical; Bioinformatics; Biological; Biological Assay; Biology; Bromouridine; Cancer Biology; Cancerous; Cell Culture Techniques; Cell Line; Cell Proliferation; Cell division; Cells; Centromere; ChIP-seq; Characteristics; Chromosome Segregation; Chromosomes; Colon Carcinoma; Complementary DNA; DNA; Data; Data Set; Ensure; Epigenetic Process; Epithelial Cells; Gene Expression; Gene Targeting; Genetic; Genetic Transcription; Genetic study; Genome Stability; Genomics; Goals; Hand; Histone H3; Human; Human Genome; In Vitro; Incubated; Label; Length; Libraries; Light; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of prostate; Measures; Messenger RNA; Methodology; Mitosis; Mitotic; Modernization; Molecular Biology; Molecular Profiling; Nature; Normal tissue morphology; Northern Blotting; Nucleic Acids; Nucleosomes; Nude Mice; Phenotype; Play; Process; Proliferating; Prostate; Prostatic; Prostatic Tissue; Proteins; RNA; RNA Binding; Recruitment Activity; Repetitive Sequence; Reporting; Role; Sampling; Satellite RNA; Structure; Surveys; Technology; Testing; Tissue Microarray; Tissues; Transplantation; VCaP; Variant; Western Blotting; Work; Xenograft procedure; anticancer research; base; cancer genomics; deep sequencing; differential expression; epigenomics; experimental study; frontier; human disease; in vivo; innovative technologies; insight; interest; knock-down; loss of function; malignant breast neoplasm; malignant phenotype; migration; monomer; mouse model; novel; novel therapeutic intervention; overexpression; prostate cancer cell; prostate cancer cell line; tool; transcriptome sequencing

Project Summary/Abstract The centromere is an essential component of the cellular machinery required for the faithful segregation of chromosomes during mitosis, a process that is significantly dysregulated in cancer. Though accurate centromeric assembly is vital for maintaining genomic stability and ensuring accurate cell division, the role centromeres play in cancer biology remains understudied. The highly repetitive underlying sequence that makes up the genomic landscape has made annotation of the centromeric region very difficult, leaving the study of centromere genomics and epigenomics as one of the last frontiers in the quest towards a more complete understanding of the human genome. New methodologies that can navigate the repetitive nature of centromeric sequences are thus required to effectively study centromere genetics and molecular biology in the context of cancer. We have developed novel bioinformatics tools that allow us to survey gene expression across large data sets to ascertain cancer specific expression of target genes. Our group has additionally developed a quantitative PCR assay to detect chromosome-specific centromeric sequences, with a focus on detecting α-satellite repeats, which are the hallmark of centromeres. Further, we have identified significant overexpression of the centromeric H3 histone variant CENPA in prostate cancer. Preliminary studies suggest that CENPA knockdown in prostate cancer cells limits their ability to proliferate. Centromeric α-satellite RNAs (CeASaRs) are known to modulate CENPA function, and we can demonstrate enrichment of specific CeASaRs when comparing prostate cancer to benign prostatic tissue. This enrichment was validated using our chromosome specific qPCR assay in whole cell RNA extracts from prostate cancer and prostatic epithelial cell lines. In view of the above, we hypothesize that centromeres represent a functionally important molecular signature that can drive prostate cancer biology. We will investigate in detail the functions of CENPA and CeASaRs in the context of prostate cancer, through deep sequencing, bioinformatics approaches, cell culture techniques and mouse models. Specifically, modulating CENPA expression in prostate cancer cell lines through overexpression and knockdown studies in cell culture and xenograft transplants will enable us to demonstrate the biological relevance of CENPA to prostate cancer. Additionally, northern blots, RACE, and long read technology available to us through the PacBio RS II platform will assist us in navigating the highly repetitive landscape of centromeric sequences. The importance of these specific CeASaRs in driving prostate cell proliferation, along with CENPA, will then be explored. These studies will shed light on a largely unexplored area in biology and in cancer research, with the potential to inform novel therapeutic approaches.

项目摘要/摘要 着丝粒是细胞机械的重要组成部分，细胞机械需要忠实地分离 染色体在有丝分裂过程中，这一过程在癌症中被显著地失调。尽管着丝粒很精确 组装对于维持基因组的稳定性和确保准确的细胞分裂是至关重要的，着丝粒所起的作用 在癌症方面，生物学仍未得到充分研究。构成基因组的高度重复的潜在序列 景观使着丝粒区域的注释变得非常困难，留下了着丝粒的研究 基因组学和表观基因组学作为寻求更完整地理解人类基因组的最后前沿之一 人类基因组。 因此，需要能够导航着丝粒序列重复性质的新方法来 在癌症的背景下有效地研究着丝粒遗传学和分子生物学。我们已经开发出了小说 生物信息学工具，使我们能够调查大型数据集的基因表达，以确定癌症的特异性 目的基因的表达。我们小组还开发了一种定量聚合酶链式反应检测方法 染色体特定着丝粒序列，重点是检测α卫星重复，这是 着丝粒的标志。此外，我们还发现着丝粒H3组蛋白显著过度表达。 前列腺癌中CENPA的变异。初步研究表明CENPA在前列腺癌细胞中被敲除 限制了它们的扩散能力。着丝粒α卫星RNA(Ceasar)是已知的调节CENPA的分子 功能，当比较前列腺癌和良性前列腺癌时，我们可以证明特异性Ceasar的丰富 前列腺组织。我们在全细胞rna中用我们的染色体特异性qpr分析验证了这种富集物。 前列腺癌和前列腺上皮细胞系的提取物。鉴于上述情况，我们假设 着丝粒是一种重要的功能分子信号，可导致前列腺癌 生物学。我们将详细研究CENPA和Ceasars在前列腺癌背景下的功能， 通过深度测序、生物信息学方法、细胞培养技术和小鼠模型。具体来说， 前列腺癌CENPA基因的过表达和基因敲除研究 细胞培养和异种移植将使我们能够证明CENPA与前列腺的生物学相关性 癌症。此外，我们可以通过PacBio RS II获得Northern blotts、Race和Long Read技术 平台将帮助我们在着丝粒序列的高度重复的景观中导航。重要的是 然后将探索这些特定的Ceasar在推动前列腺细胞增殖方面的作用，以及CENPA。这些 研究将揭示生物学和癌症研究中一个很大程度上未被探索的领域，并有可能为 新的治疗方法。