Transcription factors and genetic circuits orchestrating the complex, multilayered response of Clostridium acetobutylicum to butanol and butyrate stress

Abstract

BackgroundOrganisms of the genus Clostridium are Gram-positive endospore formers of great importance to the carbon cycle, human normo- and pathophysiology, but also in biofuel and biorefinery applications. Exposure of Clostridium organisms to chemical and in particular toxic metabolite stress is ubiquitous in both natural (such as in the human microbiome) and engineered environments, engaging both the general stress response as well as specialized programs. Yet, despite its fundamental and applied significance, it remains largely unexplored at the systems level.ResultsWe generated a total of 96 individual sets of microarray data examining the transcriptional changes in C. acetobutylicum, a model Clostridium organism, in response to three levels of chemical stress from the native metabolites, butanol and butyrate. We identified 164 significantly differentially expressed transcriptional regulators and detailed the cellular programs associated with general and stressor-specific responses, many previously unexplored. Pattern-based, comparative genomic analyses enabled us, for the first time, to construct a detailed picture of the genetic circuitry underlying the stress response. Notably, a list of the regulons and DNA binding motifs of the stress-related transcription factors were identified: two heat-shock response regulators, HrcA and CtsR; the SOS response regulator LexA; the redox sensor Rex; and the peroxide sensor PerR. Moreover, several transcriptional regulators controlling stress-responsive amino acid and purine metabolism and their regulons were also identified, including ArgR (arginine biosynthesis and catabolism regulator), HisR (histidine biosynthesis regulator), CymR (cysteine metabolism repressor) and PurR (purine metabolism repressor).ConclusionsUsing an exceptionally large set of temporal transcriptional data and regulon analyses, we successfully built a STRING-based stress response network model integrating important players for the general and specialized metabolite stress response in C. acetobutylicum. Since the majority of the transcription factors and their target genes are highly conserved in other organisms of the Clostridium genus, this network would be largely applicable to other Clostridium organisms. The network informs the molecular basis of Clostridium responses to toxic metabolites in natural ecosystems and the microbiome, and will facilitate the construction of genome-scale models with added regulatory-network dimensions to guide the development of tolerant strains.

Heterologous Expression of the Clostridium carboxidivorans CO Dehydrogenase Alone or Together with the Acetyl Coenzyme A Synthase Enables both Reduction of CO 2 and Oxidation of CO by Clostridium acetobutylicum

“…The mechanism by which the inability to produce H 2 alters the cell's biosynthetic machinery to produce metabolites requiring electrons is not well understood. It likely involves sensing of the altered redox environment (due to accumulation of reduced ferredoxin, NADH, and other electron carriers) by the redox sensor Rex, which regulates the expression of several product formation genes and pathways (34,35). As stated above, inhibition of the H 2 -producing hydrogenase would increase the Fd red pool and NADH pool derived from Fd red .…”

Rex in Clostridium kluyveri is a global redox-sensing transcriptional regulator

“…Enterococcus faecalis (Vesic and Kristich, 2013) as well as the strict anaerobes Thermoanaerobacter ethanolicus (Pei et al, 2011), C. acetobutylicum (Wang et al, 2013;Wietzke and Bahl, 2012;Zhang et al, 2014), Thermotoga maritima (Ravcheev et al, 2012) and Desulfovibrio vulgaris (Christensen et al, 2015). In these bacteria, Rex was discovered to modulate central carbon metabolism, respiration, solvent and organic acid production, hydrogen formation, tolerance to oxidative stress, biofilm formation, and sulfate and nitrate reduction.…”

Integrated, systems metabolic picture of acetone-butanol-ethanol fermentation by Clostridium acetobutylicum

“…As depicted in Fig. 3C, the model was able to show the time evolution, as well as the composition of cellular toxicity, over the course of fermentation, illustrating the combinatorial feature of toxicity from multiple sources as suggested by experiments (9,24).…”

“…From a system-level perspective, solvent production by C. acetobutylicum is indeed an extraordinarily complex process that consists of genetic regulation, metabolic shift, and cellular signal integration (7,9,10). As illustrated in Fig.…”