INFERRING CHROMOSOME ARCHITECTURE FROM GENOMIC SEQUENCE

从基因组序列推断染色体结构

• 批准号: 6397760
• 负责人: CRAIG J. BENHAM
• 金额: $ 22.14万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-17 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6397760
• 关键词: DNA binding protein; DNA replication origin; binding sites; biotechnology; chemical stability; chromosomes; eukaryote; gene therapy; genome; mathematical model; method development; nuclear matrix; nucleic acid sequence; statistics /biometry; transfection /expression vector

Eukaryotic chromosomal DNA is partitioned into domains by periodic attachment to a protein scaffold. The sites on the DNA where attachment occurs, called scaffold (or matrix) attachment regions (S/MARs), strongly influence many regulatory events including transcriptional augmentation and enhancement, initiation of DNA replication, retroviral integration, and apoptosis. Progress in understanding the mechanisms underlying scaffold attachment and its regulatory effects has been hampered by the fact that S/MARs have no consensus sequence associated with them. Recent work by this research group and our experimentalist collaborators implicates stress-induced DNA duplex destabilization (SIDD) in eukaryotic scaffold binding. When this investigator uses his theoretical method to calculate destabilization profiles of superhelical DNA sequences containing known S/MARs, the S/MAR sites coincide with regions of extensive destabilization. Our experimentalist collaborators have shown that these sites actually undergo base unpairing, both in vitro and in vivo. Moreover, the extent of SIDD is highly correlated with strength of binding. This collaborative research program will extend our investigation of the role of duplex destabilization in scaffold attachment and ancillary regulatory events. We will investigate in detail the associations between strength of SAR binding and SIDD characteristics. We will completely characterize the SIDD properties of SARs, and use this information to design artificial SAR elements based on our understanding of their required attributes. These will be constructed by our collaborators from prokaryotic DNA, and their scaffold binding properties will be assessed. We will develop computational search strategies to detect SAR elements in genomic DNA sequences by their SIDD properties. This will provide the first method for inferring chromosomal architecture and nuclear organization directly from the DNA sequence. It will enable a host of important studies to be performed regarding the organization and regulation of genomic DNA. It will be used to develop a new generation of precisely targeted and regulated transfection vectors for gene therapy.

真核染色体 DNA 通过周期性附着在蛋白质支架上而被划分为多个结构域。 DNA 上发生附着的位点称为支架（或基质）附着区域 (S/MAR)，强烈影响许多调控事件，包括转录增强和增强、DNA 复制起始、逆转录病毒整合和细胞凋亡。 S/MAR 没有与之相关的共有序列，这一事实阻碍了对支架附着机制及其调节作用的理解。该研究小组和我们的实验合作者最近的工作表明真核支架结合中应激诱导的 DNA 双链体不稳定 (SIDD)。当该研究人员使用他的理论方法计算包含已知 S/MAR 的超螺旋 DNA 序列的失稳分布时，S/MAR 位点与广泛失稳区域一致。我们的实验合作者已经证明，这些位点实际上在体外和体内都经历了碱基解配对。此外，SIDD 的程度与结合强度高度相关。该合作研究计划将扩展我们对双链体不稳定在支架附着和辅助调节事件中的作用的研究。我们将详细研究 SAR 结合强度与 SIDD 特征之间的关联。我们将完整地表征 SAR 的 SIDD 属性，并根据我们对其所需属性的理解，使用此信息来设计人工 SAR 元素。这些将由我们的合作者从原核 DNA 构建，并将评估它们的支架结合特性。我们将开发计算搜索策略，通过 SIDD 特性检测基因组 DNA 序列中的 SAR 元素。这将提供第一种直接从 DNA 序列推断染色体结构和核组织的方法。它将能够开展一系列关于基因组 DNA 的组织和调控的重要研究。它将用于开发新一代精确靶向和调控的基因治疗转染载体。