INFERRING CHROMOSOME ARCHITECTURE FROM GENOMIC SEQUENCE

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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.