INFERRING CHROMOSOME ARCHITECTURE FROM GENOMIC SEQUENCE

真核细胞染色体DNA通过定期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（S/MAR），强烈影响许多调控事件，包括转录扩增和增强，DNA复制的起始，逆转录病毒整合和凋亡。在理解支架附着及其调节作用的机制方面的进展受到了S/MAR没有与它们相关的共有序列的事实的阻碍。这个研究小组和我们的实验合作者最近的工作涉及真核支架结合中的应力诱导的DNA双链体不稳定化（SIDD）。当这位研究者使用他的理论方法计算含有已知S/MAR的超螺旋DNA序列的不稳定分布时，S/MAR位点与广泛不稳定的区域一致。我们的实验合作者已经表明，这些网站实际上进行碱基解配对，在体外和体内。此外，SIDD的程度与结合强度高度相关。这项合作研究计划将扩大我们的调查的作用，双链体不稳定的支架连接和辅助监管活动。我们将详细研究SAR结合强度和SIDD特征之间的关联。我们将完全描述SAR的SIDD特性，并根据我们对SAR所需属性的理解，使用这些信息来设计人造SAR元件。这些将由我们的合作者从原核DNA构建，并评估其支架结合特性。我们将开发计算搜索策略，以检测基因组DNA序列中的SAR元素，通过它们的SIDD属性。这将提供直接从DNA序列推断染色体结构和核组织的第一种方法。它将使一系列重要的研究能够进行关于基因组DNA的组织和调控。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。

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.