This research program will apply the theoretical methods previously developed by this investigator to the analysis of strand separation in superhelical genomic DNA sequences. The strong associations previously found between sites of stress-induced duplex destabilization (SIDD) and specific types of regulatory regions will be investigated thoroughly. Many additional DNA sequences will be analyzed to determine the SIDD properties associated to transcriptionally active regions, replication origins, centromeres, mutational hotspots, chromosomal breakpoints and other sites of biological activity. The statistical significance of each association will be assessed. The present theoretical methods will be extended to treat the following cases not currently amenable to analysis: 1) long DNAs in which the topological coupling induced by superhelicity may be limited by nucleosomal winding, 2) duplex destabilization induced by the proximal binding of molecules such as transcription factors, 3) effects on transition behavior of local sequence modifications such as methylated or mispaired bases, or abasic sites, and 4) competitions between denaturation and other transitions to which specific local DNA sequences are susceptible, such as cruciform extrusion or transition to Z- or to H-form. Monte Carlo methods will be used to analyze the stabilization of B-form DNA at high temperatures by positive supercoiling. The existing collaboration with Dr. Richard Fye, which focuses on the development of theoretical methods to analyze DNA superhelical equilibria, will be continued. Collaborations with two experimental groups, will investigate the thermodynamics of superhelical DNA conformational transitions, and the roles they play in transcription.