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Transcriptome of Aged and Unaged E. coli populations

ABSTRACT: Escherichia coli can survive for long periods in batch culture in the laboratory, where they experience a stressful and heterogenous environment. During this incubation, E. coli acquires mutations that are selected for in response to this environment, ultimately leading to evolved populations that are better adapted to these complex conditions, which can lead to a better understanding of evolutionary mechanisms. Mutations in regulatory genes often play a role in adapting to heterogenous environments. To identify such mutations, we examined transcriptional differences during log phase growth in unaged cells compared to those that had been aged for 10-days and regrown. We identified expression changes in genes involved in motility and chemotaxis after adaptation to long-term cultures. We hypothesized that aged populations would also have phenotypic changes in motility and that motility may play a role in survival and adaptation to long-term cultures. While aged populations did show an increase in motility, this increase was not essential for survival in long-term cultures. We identified putative mutations in the regulatory gene sspA, as well as other genes, that may contribute to the observed differences in motility. Taken together, these data provide an overall picture of the role of mutations in regulatory genes for adaptation, while underscoring that all changes that occur during evolution in stressful environments are not necessarily adaptive.

ORGANISM(S): Escherichia coli

PROVIDER: GSE188790 | GEO | 2022/05/27

REPOSITORIES: GEO

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Project description:Studies of the RNA polymerase-binding molecule ppGpp in bacteria and plants have shown that changes to the kinetics of the RNA polymerase can have dramatic biological effects in the short-term as a stress response. Here we describe the reprogramming of the kinetic parameters of the RNAP through mutations arising during laboratory adaptive evolution of Escherichia coli in minimal media. The mutations cause a 10- to 30-fold decrease in open complex stability at a ribosomal promoter and approximately a 10-fold decrease in transcriptional pausing in the his operon. The kinetic changes coincide with large scale transcriptional changes, including strong downregulation of motility, acid-resistance, fimbria, and curlin genes which are observed in site-directed mutants containing the RNA polymerase mutations as well as the evolved strains harboring the mutations. Site-directed mutants also grow 60% faster than the parent strain and convert the carbon-source 15% to 35% more efficiently to biomass. The results show that long-term adjustment of the kinetic parameters of RNA polymerase through mutation can be important for adaptation to a condition. Mutations in the RNA polymerase beta prime subunit (rpoC) were discovered in E. coli following adaptation to continual logarithmic growth (OD <= 0.3) in glycerol M9 minimal media at 30 C. We used site-directed mutagenesis to make strains of E. coli isogenic to wild-type except for single adaptive rpoC mutations. We found they increase growth rate in this condition by 60%. In order to understand how the mutations affect gene expression in this condition, we extracted total mRNA from the strains, which had been growing in the adaptive evolution condition, at OD=0.3. There were three RNAP mutants and the wild-type (E. coli K-12 MG1655). Each strain had three flasks from which RNA was extracted (three biological replicates). There were no technical replicates. The mRNA was synthesized into cDNA, labeled, and hybridized to an Affymetrix E. coli 2.0 GeneChip.

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Transcriptome of Aged and Unaged E. coli populations

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