Project description:Oxidative stresses commonly exist in natural environments, and microbes have developed a variety of defensive systems to counteract such events. Although increasing evidence has shown that high hydrostatic pressure (HHP) and low temperature (LT) induce antioxidant defense responses in cells, there is no direct evidence to prove the connection between antioxidant defense mechanisms and the adaptation of bacteria to HHP and LT. In this study, using the wild-type (WT) strain of a deep-sea bacterium, Shewanella piezotolerans WP3, as an ancestor, we obtained a mutant, OE100, with an enhanced antioxidant defense capacity by experimental evolution under H2O2 stress. Notably, OE100 exhibited better tolerance not only to H2O2 stress but also to HHP and LT (20 MPa and 4°C, respectively). Whole-genome sequencing identified a deletion mutation in the oxyR gene, which encodes the transcription factor that controls the oxidative stress response. Comparative transcriptome analysis showed that the genes associated with oxidative stress defense, anaerobic respiration, DNA repair, and the synthesis of flagella and bacteriophage were differentially expressed in OE100 compared with the WT at 20 MPa and 4°C. Genetic analysis of oxyR and ccpA2 indicated that the OxyR-regulated cytochrome c peroxidase CcpA2 significantly contributed to the adaptation of WP3 to HHP and LT. Taken together, these results confirmed the inherent relationship between antioxidant defense mechanisms and the adaptation of a benthic microorganism to HHP and LT.IMPORTANCE Oxidative stress exists in various niches, including the deep-sea ecosystem, which is an extreme environment with conditions of HHP and predominantly LT. Although previous studies have shown that HHP and LT induce antioxidant defense responses in cells, direct evidence to prove the connection between antioxidant defense mechanisms and the adaptation of bacteria to HHP and LT is lacking. In this work, using the deep-sea bacterium Shewanella piezotolerans WP3 as a model, we proved that enhancement of the adaptability of WP3 to HHP and LT can benefit from its antioxidant defense mechanism, which provided useful insight into the ecological roles of antioxidant genes in a benthic microorganism and contributed to an improved understanding of microbial adaptation strategies in deep-sea environments.
Project description:Microbial life in deep marine subsurface faces increasing temperatures and hydrostatic pressure with depth. In this study, we have examined growth characteristics and temperature-related adaptation of the Desulfovibrio indonesiensis strain P23 to the in situ pressure of 30 MPa. The strain originates from the deep subsurface of the eastern flank of the Juan de Fuca Ridge (IODP Site U1301). The organism was isolated at 20°C and atmospheric pressure from ~61°C-warm sediments approximately 5 m above the sediment-basement interface. In comparison to standard laboratory conditions (20°C and 0.1 MPa), faster growth was recorded when incubated at in situ pressure and high temperature (45°C), while cell filamentation was induced by further compression. The maximum growth temperature shifted from 48°C at atmospheric pressure to 50°C under high-pressure conditions. Complementary cellular lipid analyses revealed a two-step response of membrane viscosity to increasing temperature with an exchange of unsaturated by saturated fatty acids and subsequent change from branched to unbranched alkyl moieties. While temperature had a stronger effect on the degree of fatty acid saturation and restructuring of main phospholipids, pressure mainly affected branching and length of side chains. The simultaneous decrease of temperature and pressure to ambient laboratory conditions allowed the cultivation of our moderately thermophilic strain. This may in turn be one key to a successful isolation of microorganisms from the deep subsurface adapted to high temperature and pressure.
Project description:The effect of pressure and temperature on microbial communities of marine environments contaminated with petroleum hydrocarbons is understudied. This study aims to reveal the responses of marine bacterial communities to low temperature, high pressure, and contamination with petroleum hydrocarbons using seawater samples collected near an offshore Brazilian platform. Microcosms containing only seawater and those containing seawater contaminated with 1% crude oil were subjected to three different treatments of temperature and pressure as follows: (1) 22°C/0.1 MPa; (2) 4°C/0.1 MPa; and (3) 4°C/22 MPa. The effect of depressurization followed by repressurization on bacterial communities was also evaluated (4°C/22 MPaD). The structure and composition of the bacterial communities in the different microcosms were analyzed by PCR-DGGE and DNA sequencing, respectively. Contamination with oil influenced the structure of the bacterial communities in microcosms incubated either at 4°C or 22°C and at low pressure. Incubation at low temperature and high pressure greatly influenced the structure of bacterial communities even in the absence of oil contamination. The 4°C/22 MPa and 4°C/22 MPaD treatments resulted in similar DGGE profiles. DNA sequencing (after 40 days of incubation) revealed that the diversity and relative abundance of bacterial genera were related to the presence or absence of oil contamination in the nonpressurized treatments. In contrast, the variation in the relative abundances of bacterial genera in the 4°C/22 MPa-microcosms either contaminated or not with crude oil was less evident. The highest relative abundance of the phylum Bacteroidetes was observed in the 4°C/22 MPa treatment.
Project description:The chemical reaction of 2,3-naphthyridine, a nitrogen-containing aromatic compound, was investigated at pressures ranging from 0.5 to 1.5 GPa and temperatures from 473 to 573 K. A distinct decrease in the amount of residual 2,3-naphthyridine was observed in the samples recovered after reaction at ˃523 K at 0.5 and 1.0 GPa, and ˃548 K at 1.5 GPa. The formation of o-xylene and o-tolunitrile accompanied a decreasing N/C ratio of the reaction products, indicating decomposition of the aromatic ring and release of nitrogen. Precise analysis of the reaction products indicated the oligomerization of decomposed products with the residual 2,3-naphthyridine to form larger molecules up to 7mers. Nitrogen in the aromatic ring accelerated reactions to decompose the molecule and to oligomerize at lower temperatures than those typically reported for aromatic hydrocarbon oligomerization. The major reaction mechanism was similar between 0.5 and 1.5 GPa, although larger products preferentially formed in the samples at higher pressure.
Project description:Bacterial membranes are typically composed of ester-bonded fatty acid (FA), while archaeal membranes consist of ether-bonded isoprenoids, differentiation referred to as the 'lipid divide'. Some exceptions to this rule are bacteria harboring ether-bonded membrane lipids. Previous research engineered the bacterium Escherichia coli to synthesize archaeal isoprenoid-based ether-bonded lipids together with the bacterial FA ester-linked lipids, showing that heterochiral membranes are stable and more robust to temperature, cold shock, and solvents. However, the impact of ether-bonded lipids, either bacterial or archaeal, on membrane robustness, remains unclear. Here, we investigated the robustness, as survival after shock, of E. coli synthesizing either archaeal or bacterial ether-bonded membrane lipids, under high temperature and/or high hydrostatic pressure (HHP). Our findings reveal E. coli with bacterial ether-bonded lipids is more robust under HHP and high temperature. On the contrary, the presence of archaeal ether-bonded membrane lipids in E. coli does not affect the robustness under HHP nor high temperature under the tested conditions. We observed morphological changes induced by the shock treatments including reduced length under high temperature or HHP, and the presence of elongated cells after a shock of HHP and high temperature combined, suggesting the combined treatments impaired cell division. Our results contribute to a deeper understanding of membrane adaptation to extreme environmental conditions and highlight the significance of HHP as a key parameter to investigate the differentiation of membranes during the lipid divide.
Project description:Fungi inhabiting deep subseafloor sediments have been shown to possess anaerobic methane (CH4) production capabilities under atmospheric conditions. However, their ability to produce CH4 under in situ conditions with high hydrostatic pressure (HHP) remains unclear. Here, Schizophyllum commune 20R-7-F01, isolated from ~2 km below the seafloor, was cultured in Seawater Medium (SM) in culture bottles fitted with sterile syringes for pressure equilibration. Subsequently, these culture bottles were transferred into 1 L stainless steel pressure vessels at 30 °C for 5 days to simulate in situ HHP and anaerobic environments. Our comprehensive analysis of bioactivity, biomass, and transcriptomics revealed that the S. commune not only survived but significantly enhanced CH4 production, reaching approximately 2.5 times higher levels under 35 MPa HHP compared to 0.1 MPa standard atmospheric pressure. Pathways associated with carbohydrate metabolism, methylation, hydrolase activity, cysteine and methionine metabolism, and oxidoreductase activity were notably activated under HHP. Specifically, key genes involved in fungal anaerobic CH4 synthesis, including methyltransferase mct1 and dehalogenase dh3, were upregulated 7.9- and 12.5-fold, respectively, under HHP. Enhanced CH4 production under HHP was primarily attributed to oxidative stress induced by pressure, supported by intracellular reactive oxygen species (ROS) levels and comparative treatments with cadmium chloride and hydrogen peroxide. These results may provide a strong theoretical basis and practical guidance for future studies on the contribution of fungi to global CH4 flux.
Project description:Previous studies focused on psychrophilic adaptation generally have demonstrated that multiple mechanisms work together to increase protein flexibility and activity, as well as to decrease the thermostability of proteins. However, the relationship between high and low temperature adaptations remains unclear. To investigate this issue, we collected the available predicted whole proteome sequences of species with different optimal growth temperatures, and analyzed amino acid variations and substitutional asymmetry in pairs of homologous proteins from related species. We found that changes in amino acid composition associated with low temperature adaptation did not exhibit a coherent opposite trend when compared with changes in amino acid composition associated with high temperature adaptation. This result indicates that during their evolutionary histories the proteome-scale evolutionary patterns associated with prokaryotes exposed to low temperature environments were distinct from the proteome-scale evolutionary patterns associated with prokaryotes exposed to high temperature environments in terms of changes in amino acid composition of the proteins.
Project description:Addressing DNA lesion, the SOS response is conserved in bacterial domain and governed by DNA binding protein LexA, which have been well characterized in model microorganism such as Escherichia coli. However, our understanding of the roles of SOS pathway in deep-sea bacteria is limited. To indentify the composition of SOS regulon and function of LexA, we performed whole genome transcriptional profiling using a custom designed microarray which contains 95% open reading frames of Shewanella piezotolerans WP3. Here we describe the experimental procedures and methods in detail to reproduce the results (available at Gene Expression Omnibus database under GSE66790) and provide resource to be employed for comparative analyses of SOS response in microorganisms which inhabited in different environments, and thus broaden our understanding of life strategy of bacteria against various environment stresses.
Project description:Using synchrotron high-pressure X-ray diffraction at cryogenic temperatures, we have established the phase diagram for calcium up to 110 GPa and 5-300 K. We discovered the long-sought for theoretically predicted ?-tin structured calcium with I4(1)/amd symmetry at 35 GPa in a s mall low-temperature range below 10 K, thus resolving the enigma of absence of this lowest enthalpy phase. The stability and relations among various distorted simple-cubic phases in the Ca-III region have also been examined and clarified over a wide range of high pressures and low temperatures.
Project description:Cytochromes P450 (CYP) form one of the largest enzyme superfamilies on Earth, with similar structural fold but biological functions varying from synthesis of physiologically essential compounds to metabolism of myriad xenobiotics. Here we determined the crystal structures of Coryphaenoides armatus and human sterol 14α-demethylases (CYP51s). Both structures reveal elements that imply elevated conformational flexibility, uncovering molecular basis for faster catalytic rates, lower substrate selectivity, and resistance to inhibition. These elements as well as the unique inward/outward location of the FG arm/β4 hairpin in the fish CYP51 structure were not predicted by artificial intelligence molecular modelling. The structural distinction of C. armatus CYP51, which is the first structurally characterized deep-sea P450, suggests stronger involvement of the membrane environment in regulation of this enzyme function. We interpret this as a co-adaptation of membrane protein structure with changes in membrane lipid composition during evolutionary incursion to life in the deep sea.