DNA minor groove binding drugs, or MGBDs, are novel antitumor agents that have been the subject of intense study because their ability to interact with DNA in a sequence-specific fashion suggests they may be developed as targets of specific gene regulation. Several MGBDs have been shown to have potent antitumor activity in preclinical trials and are now in clinical studies. Studies aimed at understanding the sequence- and structure-dependent ways that MGBDs interact with DNA are needed to understand their biological effects and to guide the development of agents that can target the expression of specific genes. The proposed project combines in vitro and in vivo approaches to the study of the interaction of MGBDs with DNA and their effects on gene expression. Review of the literature and consideration of the results of preliminary investigations have led to the following hypotheses. First, it is proposed that the optimal binding sites for MGBDs are A/T rich, contain a high degree of homopolymeric character, and conform to the sequence requirements for DNA bending. Second, the optimal binding sites for MGBDs have structural features that make them attractive substrates for minor groove binding proteins. Third, the expression of individual genes in yeast with upstream promoter elements that contain optimal MGBD binding sites are affected at the level of transcription by MGBDs. Fourth, global patterns of gene expression in yeast are affected by MGBDs. The hypotheses will be addressed by using a novel in vitro selection protocol to determine the optimal DNA sequence and structural requirements for MGBDs, using capillary electrophoretic mobility shift assays, fluorescence enhancement, and fluorescence resonance energy transfer to measure the interaction of MGBDs and minor groove binding proteins with the optimal binding sequences, and employing both single gene and genome-wide evaluations of the effects of MGBDs on gene expression in yeast cells.
| Eckdahl, Todd T; Brown, Adam D; Hart, Steven N et al. (2008) Microarray analysis of the in vivo sequence preferences of a minor groove binding drug. BMC Genomics 9:32 |
