About the Lab
Recent advances in fluorescence technologies have led to the discovery that bacterial chromosomes exist in an extremely precise spatial orientation within the confines of the cell, a trait formerly attributed only to eukaryotic chromosomes. This organization has profound influences on every synthetic event in the cell from DNA replication to gene expression. Moreover, in E. coli, following DNA replication, portions of sister chromosomes align at homologous sequences (cohesion) before separating en masse toward opposite poles. These and other findings illustrate striking similarities between how chromosomes are maintained in bacteria and eukaryotes. With this in mind, we view the E. coli chromosome asa “stripped-down” model of eukaryotic mitosis that will help reveal the underlying molecular mechanisms that drive chromosome behavior in all cells.
My lab is developing a revolutionary fluorescence in situhybridization (FISH) technique to examine the entire chromosome in a single cell by multi-color combinatorial labeling and computer separation of overlapping spectra. This method, inspired by chromosome painting techniques used in human chromosome karyotyping, allows us to visualize the position and orientation of the chromosome in three-dimensional space. Information from the FISH studies is then pooled with other assays including whole-genome microarray analysis of chromatin immunoprecipitates (ChIP-Chip) to map locations of chromosome structure proteins.
DNA Replication and Cell Cycle Regulation
E. coli studies have provided a wealth of information on the biochemistry and genetics of DNA replication, and the basic replication machinery is highly conserved throughout all three domains of life. My lab is interested in the molecular mechanisms of DNA replication initiation and the transition between the replication origin bubble (open-complex) and association of the main replication machinery (replisome). These studies are made possible by a cell synchronization method that we developed, known as a baby cell machine. This technique generates large populations of unperturbed newborn cells that can be grown in unison, removed, and examined at any point in the cell cycle. Cell numbers are sufficient to support even very cell-demanding assays such as microarray analysis, not formerly possible with conventional synchronizing apparatuses.
My lab is also exploring an exciting model of cell cycle control in which replication is coupled to cell division via a transitory physical linkage of the chromosome to the cell membrane at the site of cell division. Breakage of this linkage at the moment of cell birth would then in theory “license” a subsequent round of replication initiation.
Knowledge from the E. coli model will provide critical advances to our understanding of how defects in genome duplication and cell cycle control lead to human diseases including many types of cancers and congenital disorders. The superb manipulability of the bacterial system makes rapid advances possible.