Organization of supercoil domains and their reorganization by transcription

Abstract

During a normal cell cycle, chromosomes are exposed to many biochemical reactions that require specific types of DNA movement. Separation forces move replicated chromosomes into separate sister cell compartments during cell division, and the contemporaneous acts of DNA replication, RNA transcription and cotranscriptional translation of membrane proteins cause specific regions of DNA to twist, writhe and expand or contract. Recent experiments indicate that a dynamic and stochastic mechanism creates supercoil DNA domains soon after DNA replication. Domain structure is subsequently reorganized by RNA transcription. Examples of transcription-dependent chromosome remodelling are also emerging from eukaryotic cell systems.

Fig. 1. Assays measuring supercoil domain structure in Salmonella and E. coli

A. A set of Salmonella strains were made with a pair of directly repeated 140 bp Tn3 res sites (red and blue DNA) separated by different segments of bacterial DNA (green). After expressing the γδ site-specific recombinases (Res), a deletion forms if and only if the res sites reside in a common supercoil domain (Stein et al., 2005). B. Escherichia coli cells contain 300 supercoil responsive genes (SSGs) whose expression is rapidly increased or decreased by a loss of negative supercoiling (Peter et al., 2004). Expression of EcoRI causes double-strand breaks and loss of supercoiling from domains with a restriction site. Changes in RNA levels measured with whole genome microarrays measure supercoil diffusion when data are combined with distance from the promoter to restriction sites in the genome (Postow et al., 2004). C. The probability of supercoil loop detection as a function of DNA length in kb is plotted for resolution assays (blue diamonds) SSG transcription data (green triangles) and loop sizes measured from EM images (red squares). (Lisa Postow provided the transcription data and plot.)

Fig. 2. Hysteresis and bistability in single cell transcription profiles

A. Overlayed green fluorescence and inverted phase-contrast images of cells that are initially un-induced for lac expression, then grown for 20 h in 18 μM thio-methylgalactoside (TMG), a non-metabolizable lactose analogue. The cells show a bimodal distribution of lac expression, with induced cells having over one hundred times the green fluorescence of uninduced cells. B. A series of cell populations, initially un-induced (lower panel) or fully induced (upper panel) for lac expression, were grown 20 h in media containing various amounts of TMG. Scatter plots show log (green fluorescence) for about 1000 cells in each population. Each scatter plot is centred at a point indicating the underlying TMG concentration (Ozbudak et al., 2004). Figure reproduced with permission from the Nature Publishing Group.