Integrated analysis of molecular and systems level function of the Crp regulon [ChIP-exo]

A fundamental goal of biological research is to connect molecular events with macroscopic physiological phenomena. By incorporating a multi-scale approach that accounts for mechanistic properties of promoters and maps those findings to a genome-scale metabolic-regulatory network, the Escherichia coli Crp regulon is revealed, enabling systems analytics to be applied to delineate its governing role in regulating cellular carbon flow. Genome-wide promoter functions were characterized using ChIP-exo elucidating stable intermediates formed during transcription initiation and distinctions between transcriptional activators and repressors. Genome-wide analysis of Crp regulated genes showed a coordinate regulation of catabolism, anabolism, and energy-generating chemiosmotic processes. Examination of the functional state of the reconstructed regulon during growth on multiple substrates revealed a coherent quantitative adjustment of these overall metabolic processes based on substrate quality, revealing the detailed mechanistic basis for the phenomenologically observed homoeostatic response. Thus, a multi-scale relationship from individual molecular mechanisms to physiological properties is established. Overall design: A total of 13 different experimental conditions are included yielding a total of 43 individual ChIP-exo samples. Invidividual conditions were produced with a minimum of 2 biological replicates. All samples were collected at mid-exponential phase from shake flasks grown on M9 minimal media supplemented with glucose, fructose, or glycerol. Crp, delta Crp, and the Crp mutants delAr1, delAr2, delAr1delAr2, and RpoD ChIP studies were conducted aerobically. ArcA and Fnr ChIP studies were conducted under anaerobi conditions. Delta Crp was generated as a control for ChIP studies of Crp.