Exposure to 1-Butanol Exemplifies the Response of the Thermoacidophilic Archaeon Sulfolobus acidocaldarius to Solvent Stress

Abstract

Sulfolobus acidocaldarius is a thermoacidophilic crenarchaeon with optimal growth at 80 °C and pH 2 - 3. Due to its unique physiological properties allowing life at environmental extremes and recent availability of genetic tools, this extremophile receives increasing interest for biotechnological application. In order to elucidate the potential of tolerating process-related stress conditions, we investigated the response of S. acidocaldarius towards the industrially relevant organic solvent 1-butanol. In response to butanol exposure, biofilm formation of S. acidocaldarius was enhanced and occurred up to 1.5% (v/v) 1-butanol, while planktonic growth was only observed up to 1% (v/v) 1-butanol. Confocal laser scanning microscopy revealed that biofilm architecture changed with the formation of denser and higher tower-like structures. Concomitantly, changes in the extracellular polymeric substances with enhanced carbohydrate and protein content were determined in 1-butanol-exposed biofilms. Using scanning electron microscopy three different cell morphotypes were observed in response to 1-butanol. Transcriptome and proteome analyses were performed comparing the response of planktonic and biofilm cells in absence and presence of 1-butanol. In response to 1% (v/v) 1-butanol transcript levels of genes encoding motility and cell envelope structures as well as membrane proteins were reduced. Cell division and/or vesicle formation was upregulated. Furthermore, changes of immune and defence systems, as well as metabolism and general stress response were observed. Our findings show that the extreme lifestyle of S. acidocaldarius coincided with a high tolerance to organic solvents. This study provides first insights into biofilm formation and membrane/cell stress caused by organic solvents in S. acidocaldarius. Importance Archaea are unique in terms of metabolic and cellular processes as well as the adaptation to extreme environments. In the past few years, the development of genetic systems and biochemical, genetic and poly-omics studies have provided deep insights into the physiology of some archaeal model organisms. In this study, we used S. acidocaldarius adapted to two extremes, low pH and high temperature, to study its tolerance and robustness as well as its global cellular response towards organic solvents exemplified by 1-butanol. We were able to identify biofilm formation as primary cellular response to 1-butanol. Furthermore, the triggered cell/membrane stress led to significant changes in culture heterogeneity accompanied by changes in central cellular processes such as cell division and cellular defense systems, thus suggesting a global response for the protection at population level.

Exopolysaccharide composition and size in Sulfolobus acidocaldarius biofilms

“…Compared to other published work on S. acidocaldarius biofilms, using MTPs or dishes for cultivation (Koerdt et al, 2010;Benninghoff et al, 2017), an alternative cultivation method was established to generate high amounts of biomass necessary for subsequent EPS analysis. Different polymeric membranes were tested for their suitability to form homogeneous, thick biofilms.…”

“…Up to now, archaeal biofilms were mostly generated in MTPs, dishes or on slides consisting of different types of polymers (e.g., polystyrene) or glass as substratum for biofilm formation (Benninghoff et al, 2017). While the cultivation of biofilms, e.g., in MTPs, is optimal to analyze biofilm properties on a high-throughput scale (see section "Sulfolobus acidocaldarius biofilm formation in different growth media"), it is suboptimal to receive high amounts of biofilm mass.…”

Perspectives on the microorganism of extreme environments and their applications

“…Biobutanol : Butanol hinders the growth of microorganisms and therefore, most microbes cannot tolerate more than 2% of butanol ( Benninghoff et al., 2021 ). A few species of Bacillus can tolerate 2.5–7% of butanol concentration.…”

Machine Learning Uncovers a Data-Driven Transcriptional Regulatory Network for the Crenarchaeal Thermoacidophile Sulfolobus acidocaldarius

“…There are also six new genes in the FadR iModulon that are not in the currently known regulon (Supplementary Figure 2). All six genes are functionally uncharacterized, with the exception of Saci_1992; this gene is a CRISPR-associated TF (Benninghoff et al, 2021). Five of these genes have negative gene weights, which means that their expression is anticorrelated with the expression of the FadR Sa gene cluster.…”