Interaction of Protein Occupancy, Chromosome Structure, and Gene Regulation in E. coli
Authors: Kroner Grace Mahony , Scholz Scott , Goss Thomas J. , Lin Xiaoxia , Freddolino Peter L. 
Published: November 2023

Chromosome structure facilitates coordinated regulation of related genes as well as controlling the access of transcription machinery to DNA. In human cells, the control of a chromosome's location within the nucleus, its contacts with other sections of DNA, and its individual collection of DNA binding proteins is a well‐established method of regulation. In the simpler bacterial cell, with no nuclear membrane and with genetic information contained in a circular chromosome, initial hypotheses were that the DNA does not have a controlling architecture. However, studies by Valens et al. (EMBO 2004) and Cagliero et al. (Nucleic Acids Research 2013) in Escherichia coli and Umbarger et al. (Mol. Cell 2011) in Caulobacter crescentus illustrated that the genetic material of bacteria also appears to follow some structural rules. From these studies, researchers hypothesized that the E. coli chromosome follows a macrodomain structure, with nearby segments forming clusters distinct from neighboring DNA. In contrast, the Caulobacter chromosome appeared to twist on itself, producing interactions between points halfway across the genome. Independently, Vora et al. (Mol. Cell 2009) observed that the E. coli chromosome contains regions of very high protein occupancy, termed EPODs, many of which are transcriptionally silent and which are enriched for nucleoid protein binding sites.To unify these observations, we have performed experiments to characterize the conformation of E. coli genomes at different stages of growth and to analyze the interaction of protein occupancy with genome structure and gene transcription. Our genome conformation capture studies revealed no signs of higher order structure in either logarithmic or stationary E. coli. However, we do see correlations between methylation, recombination hot spots and regions of high DNA‐DNA interaction. In addition, studies of genome wide protein occupancy reveal an anti‐correlation between extended protein occupancy domains (EPODs) and DNA accessibility; i.e., EPODs are usually found in regions of DNA with low interaction frequencies. EPODs appear to result from the cooperative activity of several nucleoid associated proteins including Fis and Hfq, so it is unsurprising that they appear to have a connection with chromosome structure. The presence of such EPODs would be presumed to affect the likelihood of gene expression just as heterochromatic regions of eukaryotic DNA are known to repress transcription. To assess the effect of EPOD occupancy on gene expression, we have inserted a uniform construct into various locations on the E. coli chromosome and measured expression, identifying a crucial role for EPODs in modulating transcription rates. These studies provide a global picture of the fundamental role of chromosome organization in E. coli.Support or Funding InformationSupport from the Cellular Biotechnology Training Program (GMK), R00 GM097033 (PLF) and University of Michigan startup funding (PLF)