Identifying the Growth Modulon of Corynebacterium glutamicum

Abstract

The growth rate (μ) of industrially relevant microbes, such as Corynebacterium glutamicum , is a fundamental property that indicates its production capacity. Therefore, understanding the mechanism underlying the growth rate is imperative for improving productivity and performance through metabolic engineering. Despite recent progress in the understanding of global regulatory interactions, knowledge of mechanisms directing cell growth remains fragmented and incomplete. The current study investigated RNA-Seq data of three growth rate transitions, induced by different pre-culture conditions, in order to identify transcriptomic changes corresponding to increasing growth rates. These transitions took place in minimal medium and ranged from 0.02 to 0.4 h -1 μ. This study enabled the identification of 447 genes as components of the growth modulon. Enrichment of genes within the growth modulon revealed 10 regulons exhibiting a significant effect over growth rate transition. In summary, central metabolism was observed to be regulated by a combination of metabolic and transcriptional activities orchestrating control over glycolysis, pentose phosphate pathway, and the tricarboxylic acid cycle. Additionally, major responses to changes in the growth rate were linked to iron uptake and carbon metabolism. In particular, genes encoding glycolytic enzymes and the glucose uptake system showed a positive correlation with the growth rate.

Revisiting the Growth Modulon of Corynebacterium glutamicum Under Glucose Limited Chemostat Conditions

“…Further transcript analysis focused on genes encoding glucose uptake and regulation in C. glutamicum ( Figure 6 ). Noteworthy, published data of Haas et al (2019) serve as reference. The expression profiles mirror transcript dynamics of the same strain in the same synthetic medium after its exposure to glucose limitation (highlighted in red in Figure 6 ).…”

“…Transcriptome sampling was performed by withdrawing 1 mL biosuspension from the bioreactor, centrifuging at 20,817 × g and 4°C for 30 s (5430 R, Eppendorf, Hamburg, Germany), discarding the supernatant and immediately quenching the biomass pellet in liquid nitrogen. Extraction and isolation of RNA from the samples, whole transcriptome sequencing, read mapping, and raw read count calculation was performed as described in Haas et al (2019) . The raw sequencing data have been deposited in the ArrayExpress database at EMBL-EBI under accession number E-MTAB-9371 .…”

“…However, a putative drawback lies in the somewhat reduced growth rates compared to other hosts such as Escherichia coli . With about 0.4 h –1 wild-type (WT) C. glutamicum shows only moderate growth in synthetic media in classical batch cultivations ( Blombach et al, 2013 ; Grünberger et al, 2013 ; Graf et al, 2018 ; Haas et al, 2019 ) compared to about 0.7–0.8 h –1 maximum growth rate of other prokaryotes such as E. coli , 1.7 h –1 for the putative future competitors Vibrio natriegens ( Hoffart et al, 2017 ) and Geobacillus LC300 with 2.15 h –1 ( Long et al, 2017 ). Relatively low space-time yields compared to other hosts may be the consequence.…”

“…Under the tested glucose-limited growth conditions, Noack et al (2017) concluded that metabolic fluxes within C. glutamicum ’s central carbon metabolism are likely more affected by concentrations of local substrates and allosteric metabolite effectors than by the concentration of enzymes. Recently, another systems biology study was performed in traditional non-substrate limited batch cultivations to investigate determinants of global regulation orchestrating the growth of C. glutamicum ( Haas et al, 2019 ). The so-called growth modulon was discovered by investigating transcriptional dynamics of strains in batch cultivation.…”

“…Based on these recent systems biology insights, the current study tries to broaden knowledge about the strain’s metabolic, transcriptional, and fluxomic response to different growth rates. Accordingly, we continuously cultivated C. glutamicum under glucose-limited growth conditions setting dilution rates of 0.2, 0.3, and 0.4 h –1 , applying the same strain as Haas et al (2019) . We analyzed whether the transcriptional regulatory response identified in glucose-rich batch cultures is similar to the one operational under chemostat conditions at similar growth rates.…”