INFERRING CHROMOSOME ARCHITECTURE FROM GENOMIC SEQUENCE

从基因组序列推断染色体结构

• 批准号: 6095266
• 负责人: CRAIG J. BENHAM
• 金额: $ 30.43万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 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 2001-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6095266
• 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.

真核细胞的染色体DNA通过周期性地附着在蛋白质支架上而被分割成不同的结构域。发生附着的部位称为支架(或基质)附着区(S/MARS)，它强烈影响许多调控事件，包括转录增强和增强、DNA复制的启动、逆转录病毒整合和细胞凋亡。S/MARS没有与支架附着相关的一致序列，这阻碍了对支架附着及其调控作用机制的理解。该研究小组和我们的实验合作者最近的工作表明，应激诱导的DNA双链失稳(SIDD)在真核细胞支架结合中存在。当这位研究人员使用他的理论方法计算包含已知的S/火星的超螺旋DNA序列的失稳剖面时，S/MAR位点与广泛失稳的区域重合。我们的实验者合作者已经证明，无论是在体外还是在体内，这些位点实际上都经历了碱基去配对。此外，SIDD的程度与结合强度高度相关。这一合作研究计划将扩展我们对支架附着和辅助调控事件中双链失稳作用的研究。我们将详细研究SAR结合强度与SIDD特征之间的关联。我们将完全描述SARS的SIDD属性，并使用这些信息根据我们对其所需属性的理解来设计人造SAR元素。这些将由我们的合作者从原核DNA中构建，并将评估它们的支架结合性能。我们将开发计算搜索策略，根据SIDD属性检测基因组DNA序列中的SAR元件。这将为直接从DNA序列推断染色体结构和核组织提供第一种方法。它将使许多关于基因组DNA的组织和调控的重要研究得以进行。它将被用来开发新一代精确靶向和调控的转基因载体用于基因治疗。