The role of ASARs in chromosome dynamics

ASAR 在染色体动力学中的作用

• 批准号: 10396472
• 负责人: MATHEW J THAYER
• 金额: $ 30.8万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10396472
• 关键词: 3-Dimensional; Affect; Alleles; Architecture; Biochemical; Biological Assay; Biology; Biotin; CRISPR/Cas technology; Cell Cycle; Cell Nucleus; Centromere; Characteristics; Chromatin; Chromosome Replication Timing; Chromosome Structures; Chromosome Territory; Chromosomes; Cis-Acting Sequence; Clinical; Control Locus; Cre-LoxP; DNA biosynthesis; Development; Disease; Drug Design; Drug resistance; Elements; Engineering; Ensure; Evolution; Foundations; Gene Expression; Genes; Genetic; Genetic Transcription; Genetic study; Genome; Genomics; Heterogeneity; Hi-C; Human; Human Chromosomes; Image; Immunoprecipitation; Individual; Interphase Chromosome; Lead; Length; Location; Maintenance; Malignant Neoplasms; Mammalian Cell; Mammalian Chromosomes; Maps; Mediating; Mitotic; Modeling; Morphology; Mus; Nature; Nuclear; Nucleotides; Pathogenesis; Phase; Phenotype; Physical condensation; Play; Proteins; Published Comment; RNA; RNA purification; Replication Origin; Role; Site; Small Interfering RNA; Structure; Testing; Time; Transgenes; Untranslated RNA; Work; autosome; cis acting element; design; experimental study; gene function; genome integrity; insight; mammalian genome; model design; neoplastic cell; novel; novel therapeutic intervention; novel therapeutics; prevent; programs; reconstruction; segregation; telomere

Summary/Abstract Mammalian cells initiate DNA replication at multiple sites along each chromosome at different times, following a temporal replication program. We have used a chromosome-engineering strategy to identify cis-acting loci that control this replication-timing program on individual human chromosomes. We found that Cre/loxP- mediated translocations, affecting eight different autosomes, display a delay in replication timing and structural instability of entire chromosomes. Subsequently, we found that Cre/loxP-mediated disruption of the lncRNA genes ASAR6 and ASAR15 result in delayed replication of human chromosomes 6 and 15, respectively. ASAR6 and ASAR15 share numerous characteristics, including: 1) random mono-allelic expression of lncRNAs that can physically “coat” entire chromosomes in cis; 2) asynchronous replication between alleles; 3) genetic disruption results in structural instability of their respective chromosomes; and 4) ectopic integration of transgenes causes delayed replication of entire chromosomes in cis. In earlier studies, we detected the delayed replication and structural instability phenotypes following rearrangement of numerous human and mouse chromosomes, suggesting that all mammalian chromosomes are regulated by similar mechanisms. Our work suggests that all mammalian chromosomes are regulated by similar loci. Therefore, we are proposing that all mammalian chromosomes contain “Inactivation/Stability Centers” (I/SCs), which normally function to promote proper replication timing, monoallelic gene expression and structural stability of individual chromosomes. We believe that I/SCs are as fundamentally important to mammalian chromosome biology as telomeres, centromeres, or origins of replication. Thus, under this scenario every mammalian chromosome contains four essential cis-acting elements, origins, centromeres, telomeres, and I/SCs all functioning to ensure proper replication, segregation and stability of each chromosome. This proposal is designed to elucidate the mechanisms by which ASAR6 and ASAR15 RNAs control chromosome replication timing, to identify chromatin interaction sites of ASAR6 and ASAR15 RNAs, and to determine the mechanisms by which ASAR6 and ASAR15 function to localize chromosomes within the 3D space of the nucleus, and if they control the genome interaction maps of chromosomes 6 and 15. If successful the experiments described in this proposal will help to establish a new paradigm for mammalian chromosome biology, and will serve as the foundation for a fourth type of essential chromosomal element, the “Inactivation/Stability Center”.

总结/摘要 哺乳动物细胞在不同的时间在沿着每条染色体的多个位点启动DNA复制， 一个时间复制程序我们已经使用了染色体工程的策略，以确定顺式作用位点 控制着人类染色体上的复制定时程序。我们发现Cre/loxP- 介导的易位，影响八个不同的常染色体，显示复制时间和结构的延迟， 整个染色体的不稳定性随后，我们发现Cre/loxP介导的lncRNA破坏 基因ASAR 6和ASAR 15分别导致人类染色体6和15的延迟复制。 ASAR 6和ASAR 15共有许多特征，包括：1）ASAR 6和ASAR 15的随机单等位基因表达， lncRNA可以顺式物理地“包覆”整个染色体; 2）等位基因之间的异步复制; 3） 遗传破坏导致其各自染色体的结构不稳定;和4） 转基因引起整个顺式染色体的延迟复制。在早期的研究中，我们检测到 延迟复制和结构不稳定表型后重排的许多人类和 小鼠染色体，这表明所有哺乳动物染色体都受到类似机制的调控。我们 研究表明，所有哺乳动物的染色体都受到类似基因座的调控。因此，我们建议 所有哺乳动物的染色体都含有“失活/稳定中心”（I/SC），其功能通常是 促进适当的复制时机、单等位基因基因表达和个体结构稳定性 染色体我们相信I/SC对哺乳动物染色体生物学的重要性不亚于 端粒、着丝粒或复制起点。因此，在这种情况下， 包含四个基本的顺式作用元件，即起源、着丝粒、端粒和I/SC，所有这些元件都起作用， 确保每个染色体的正确复制、分离和稳定性。这项建议旨在 阐明ASAR 6和ASAR 15 RNA控制染色体复制时间的机制， 确定ASAR 6和ASAR 15 RNA的染色质相互作用位点，并确定其机制， ASAR 6和ASAR 15的功能是将染色体定位在细胞核的3D空间内，并且如果它们控制染色体， 6号和15号染色体的基因组相互作用图谱。如果成功的实验中所描述的， 该提案将有助于为哺乳动物染色体生物学建立一个新的范式，并将作为 第四类基本染色体元件的基础，“失活/稳定中心”。