Many cellular functions depend upon interactions between proteins and nucleic acids. These processes can be extremely complicated, often requiring interactions between multiple protein and DNA components to ensure biologically sensible outcomes. Due to this high degree of complexity it has been difficult to obtain a detailed understanding of the molecular nature of many important biological processes. One reason for this is that most methods of biochemical analysis rely on the measurement of population averaged behaviors. Asynchronous or heterogeneous reactions can present significant difficulties when studied with ensemble-based methods. Rare, transient, or unstable intermediates can also be very difficult to detect. Experimental analysis of single molecules allows us to circumvent many of these problems.

Currently we are using these single molecule techniques to study the eukaryotic molecular machinery involved in the recombination and repair of damaged DNA. Specifically, we are examining the roles of Rad51, Dmc1, Rad52, and Rad54 in the repair of double strand DNA breaks via homologous recombination. Our experiments are designed to directly observe these proteins as they interact with one another and their DNA substrates, focusing on understanding how these interactions are coupled to the outcome of the homologous recombination reaction. Our work will contribute to the overall understanding of how a cell recognizes and repairs DNA damage, how defects in this pathway can result in the development of cancer, and how this pathway can be manipulated to treat or prevent the formation of cancerous cells.

Selected Publications

Greene, E.C. and Mizuuchi, K. Direct observation of the assembly and disassembly of the MuB target complex (In preparation).

Greene, E.C. and Mizuuchi, K. Target Immunity during Mu DNA Transposition: Transpososome Assembly and DNA Looping Enhance MuA-Mediated Disassembly of the MuB Target Complex Molecular Cell 2002 10: 1367-1378.

Greene, E.C. and Mizuuchi, K. 2002. Direct observation of single MuB polymers: evidence for a DNA-dependent conformational change for generating an active target complex. Molecular Cell 9(5): 1079-1089.

Greene, E.C. and Mizuuchi, K. 2002. Dynamics of a protein polymer: the assembly and disassembly pathways of the MuB transposition target complex. EMBO J. 21(6):1477-1486.

Greene, E.C., and Shippen D.E. 1998. Developmentally-programmed assembly of higher order telomerase complexes with distinct biochemical and structural properties. Genes & Development. 12:2921-2931.