Structure and Dynamics of the Bacterial Chromosome in E. Coli Monitored by Gfp-Fis

Abstract

The bacterial cell's ability to control the topology of the 1.5 mm-long DNA in the confined environment of the cell is quite remarkable. Despite a great number of studies on bacteria, and especially E coli, our understanding of the spatio-temporal organization of bacterial chromosomes is minimal, partly because their dynamics have been difficult to observe directly. Using fluorescent-protein techniques we can visualize bacterial chromosome conformation during cell growth and division through fluorescent microscopy. We have developed a bacterial strain containing fluorescent gfp-fusion versions of a chromosome folding protein, Fis, under inducible control. Bacterial chromosomes have been studied in cells and removed from cells, in order to establish their spatial organization and mechanical properties, and to study how those properties are changed by varied external conditions. Space-time studies of the nucleoid in live E coli cells shows how domain structure and overall conformation of chromosomes vary during rapid and slow growth, and it also shows a relation between chromosome segregation and cell division under these different growth conditions. In order to study the bacterial chromosome outside of the cell, we have developed methods for isolation of single bacterial chromosomes and our further objective will be to directly examine nucleoid mechanical properties as a function of protein levels using micromanipulation methods.