Latent Pathway Activation and Increased Pathway Capacity Enable Escherichia coli Adaptation to Loss of Key Metabolic Enzymes

Abstract

The ability of biological systems to adapt to genetic and environmental perturbations is a fundamental but poorly understood process at the molecular level. By quantifying metabolic fluxes and global mRNA abundance, we investigated the genetic and metabolic mechanisms that underlie adaptive evolution of four metabolic gene deletion mutants of Escherichia coli (⌬pgi, ⌬ppc, ⌬pta, and ⌬tpi) in parallel evolution experiments of each mutant. The initial response to the gene deletions was flux rerouting through local bypass reactions or normally latent pathways. The principal effect of evolution was improved capacity of already active pathways, whereas new flux distributions were not observed. Combinatorial changes in capacity and pathway activation, however, led to different intracellular flux states that enabled evolution in three of the four parallel cases tested. The molecular bases of the evolved phenotypes were then elucidated by global mRNA transcript analyses. Activation of latent pathways and flux changes in the tricarboxylic acid cycle were found to correlate well with molecular changes at the transcriptional level. Flux alterations in other central metabolic pathways, in contrast, were apparently not connected to changes in the transcriptional network. These results give new insight into the dynamics of the evolutionary process by demonstrating the flexibility of the metabolic network of E. coli to compensate for genetic perturbations and the utility of combining multiple high throughput data sets to differentiate between causal and noncausal mechanistic changes.

Computational and Experimental Analysis of Redundancy in the Central Metabolism of Geobacter sulfurreducens

“…Recent genetic and in silico studies have shown that the presence of such redundant metabolic pathways, as well as isozymes, can enable metabolic networks to withstand genetic perturbations [23][24][25][26]. Experimental evidence for alternate optimal pathways have been observed in E. coli, where four metabolic gene deletion mutants had significantly different metabolic flux distributions, but similar overall growth rates [25].…”

Mixotrophic and photoheterotrophic metabolism in Cyanothece sp. ATCC 51142 under continuous light

“…Two nitrogen sources other than N 2 , ammonia and nitrate, were also examined. Precise readouts of metabolic state and activity were based on 13 C-assisted metabolite analysis integrated with biochemical assays and the gene expression patterns obtained by RT-PCR (Fong et al, 2006;Pingitore et al, 2007;Tang et al, 2007cTang et al, , 2009Wu et al, 2010). Superior to the traditional 14 C method (Bottomley & Van Baalen, 1978), the non-radioactive 13 C method can provide rich information about which carbons within a metabolite are labelled, and thus enable an indepth understanding of carbon utilization and metabolic regulation in Cyanothece 51142.…”

Central Role of the Cell in Microbial Ecology

“…It has been demonstrated that even relatively small changes in the genome (single-nucleotide polymorphisms [SNPs] as well as insertions and deletions [indels]) can have substantial effects on the phenotype and fitness of the strains (13,169). Mutations can take place in structural genes influencing membrane fluidity (246); in metabolic enzymes they can reroute carbon and energy fluxes and increase metabolic efficiency (65,101); and in regulatory elements or in the transcription machinery (e.g., transcription factors and the RNA polymerase or promoter region) they can have effects on transcription/translation speed, transcript stability, and strength of induction and repression of genes (40). It has been demonstrated that organisms adapt rapidly on the molecular level to changing environmental conditions by increasing mRNA expression levels (11,41,47,93).…”