Implication of amino acid metabolism and cell surface integrity for the thermotolerance mechanism in the thermally adapted acetic acid bacterium Acetobacter pasteurianus TH-3.

IMPORTANCE: Acetobacter pasteurianus, an industrial vinegar-producing strain, is suffered by fermentation stress such as fermentation heat and/or high concentrations of acetic acid. By an experimental evolution approach, we have obtained a stress-tolerant strain, exhibiting significantly increased growth and acetic acid fermentation ability at higher temperatures. In this study, we report that only the three gene mutations of ones accumulated during the adaptation process, ansP, dctD, and glnD, were sufficient to reproduce the increased thermotolerance of A. pasteurianus. These mutations resulted in cell envelope modification, including increased phospholipid and lipopolysaccharide synthesis, increased respiratory activity, and cell size reduction. The phenotypic changes may cooperatively work to make the adapted cell thermotolerant by enhancing cell surface integrity, nutrient or oxygen availability, and energy generation.

Fpr1, a primary target of rapamycin, functions as a transcription factor for ribosomal protein genes cooperatively with Hmo1 in Saccharomyces cerevisiae.

Fpr1 (FK506-sensitive proline rotamase 1), a protein of the FKBP12 (FK506-binding protein 12 kDa) family in Saccharomyces cerevisiae, is a primary target for the immunosuppressive agents FK506 and rapamycin. Fpr1 inhibits calcineurin and TORC1 (target of rapamycin complex 1) when bound to FK506 and rapamycin, respectively. Although Fpr1 is recognised to play a crucial role in the efficacy of these drugs, its physiological functions remain unclear. In a hmo1Δ (high mobility group family 1-deleted) yeast strain, deletion of FPR1 induced severe growth defects, which could be alleviated by increasing the copy number of RPL25 (ribosome protein of the large subunit 25), suggesting that RPL25 expression was affected in hmo1Δfpr1Δ cells. In the current study, extensive chromatin immunoprecipitation (ChIP) and ChIP-sequencing analyses revealed that Fpr1 associates specifically with the upstream activating sequences of nearly all RPG (ribosomal protein gene) promoters, presumably in a manner dependent on Rap1 (repressor/activator site binding protein 1). Intriguingly, Fpr1 promotes the binding of Fhl1/Ifh1 (forkhead-like 1/interacts with forkhead 1), two key regulators of RPG transcription, to certain RPG promoters independently of and/or cooperatively with Hmo1. Furthermore, mutation analyses of Fpr1 indicated that for transcriptional function on RPG promoters, Fpr1 requires its N-terminal domain and the binding surface for rapamycin, but not peptidyl-prolyl isomerase activity. Notably, Fpr1 orthologues from other species also inhibit TORC1 when bound to rapamycin, but do not regulate transcription in yeast, which suggests that these two functions of Fpr1 are independent of each other.

Possible involvement of extracellular polymeric substrates of Antarctic cyanobacterium Nostoc sp. strain SO-36 in adaptation to harsh environments.

Cyanobacteria are some of the primary producers in extremely cold biospheres such as the Arctic, Antarctic, and vast ice sheets. Many genera of cyanobacteria are identified from these harsh environments, but their specific mechanisms for cold adaptation are not fully understood. Nostoc sp. strain SO-36 is a cyanobacterium isolated in Antarctica more than 30 years ago and regarded as a psychrotolelant species. To determine whether the strain is psychrotolelant or psychrophilic, it was first grown at 30 °C and 10 °C. The cells grew exponentially at 30 °C, but their growth stopped at 10 °C, indicating that the strain is only psychrotolerant. Microscopic analysis revealed that the morphology of the cells grown at 30 °C was filamentous and differentiated heterocysts, which are specialized cells for gaseous nitrogen fixation under nitrogen-deprived conditions, indicating that the strain can grow diazotrophically. The cells grown at 10 °C have a smaller size, shortened filament length and decreased chlorophyll content per cell. At 10 °C, the cells are aggregated with extracellular polymeric substrates (EPSs), which is a common mechanism to protect cells from ultraviolet light. These results imply that segmentation into short filaments was induced by photodamage at low temperatures. To fully understand the adaptation mechanisms of Nostoc sp. strain SO-36 for low-temperature conditions, next-generation sequencing analyses were conducted. Complete genome sequence of the strain revealed that it has one main chromosome of approximately 6.8 Mbp with 4 plasmids, including 6855 coding sequences, 48 tRNA genes, 4 copies of rRNA operons, and 5 CRISPR regions. Putative genes for EPS biosynthesis were found to be conserved in Nostocaceae regardless of their habitat. These results provide basic information to understand the adaptation mechanisms at low temperatures, and the strain can be a model organism to analyze adaptation to extreme environments.

The CRP-family transcriptional regulator DevH regulates expression of heterocyst-specific genes at the later stage of differentiation in the cyanobacterium Anabaena sp. strain PCC 7120.

Heterocysts are terminally differentiated cells of filamentous cyanobacteria, which are specialized for nitrogen fixation. Because nitrogenase is easily inactivated by oxygen, the intracellular environment of heterocysts is kept microoxic. In heterocysts, the oxygen-evolving photosystem II is inactivated, a heterocyst-specific envelope with an outer polysaccharide layer and an inner glycolipid layer is formed to limit oxygen entry, and oxygen consumption is activated. Heterocyst differentiation, which is accompanied by drastic morphological and physiological changes, requires strictly controlled gene expression systems. Here, we investigated the functions of a CRP-family transcriptional regulator, DevH, in the process of heterocyst differentiation. A devH-knockdown strain, devH-kd, was created by replacing the original promoter with the gifA promoter, which is repressed during heterocyst differentiation. Although devH-kd formed morphologically distinct cells with the heterocyst envelope polysaccharide layer, it was unable to grow diazotrophically. Genes involved in construction of the microoxic environment, such as cox operons and the hgl island, were not upregulated in devH-kd. Moreover, expression of the nif gene cluster was completely abolished. Although CnfR was expressed in devH-kd, the nif gene cluster was not induced even under microoxic conditions. Thus, DevH is necessary for the establishment of a microoxic environment and induction of nitrogenase in heterocysts.

Novel (p)ppGpp0 suppressor mutations reveal an unexpected link between methionine catabolism and GTP synthesis in Bacillus subtilis.

In bacteria, guanosine (penta)tetra-phosphate ([p]ppGpp) is essential for controlling intracellular metabolism that is needed to adapt to environmental changes, such as amino acid starvation. The (p)ppGpp0 strain of Bacillus subtilis, which lacks (p)ppGpp synthetase, is unable to form colonies on minimal medium. Here, we found suppressor mutations in the (p)ppGpp0 strain, in the purine nucleotide biosynthesis genes, prs, purF and rpoB/C, which encode RNA polymerase core enzymes. In comparing our work with prior studies of ppGpp0 suppressors, we discovered that methionine addition masks the suppression on minimal medium, especially of rpoB/C mutations. Furthermore, methionine addition increases intracellular GTP in rpoB suppressor and this effect is decreased by inhibiting GTP biosynthesis, indicating that methionine addition activated GTP biosynthesis and inhibited growth under amino acid starvation conditions in (p)ppGpp0 backgrounds. Furthermore, we propose that the increase in intracellular GTP levels induced by methionine is due to methionine derivatives that increase the activity of the de novo GTP biosynthesis enzyme, GuaB. Our study sheds light on the potential relationship between GTP homeostasis and methionine metabolism, which may be the key to adapting to environmental changes.

Transcriptome differences between Cupriavidus necator NH9 grown with 3-chlorobenzoate and that grown with benzoate.

RNA-seq analysis of Cupriavidus necator NH9, a 3-chlorobenzoate degradative bacterium, cultured with 3-chlorobenzaote and benzoate, revealed strong induction of genes encoding enzymes in degradation pathways of the respective compound, including the genes to convert 3-chlorobenzaote and benzoate to chlorocatechol and catechol, respectively, and the genes of chlorocatechol ortho-cleavage pathway for conversion to central metabolites. The genes encoding transporters, components of the stress response, flagellar proteins, and chemotaxis proteins showed altered expression patterns between 3-chlorobenzoate and benzoate. Gene Ontology enrichment analysis revealed that chemotaxis-related terms were significantly upregulated by benzoate compared with 3-chlorobenzoate. Consistent with this, in semisolid agar plate assays, NH9 cells showed stronger chemotaxis to benzoate than to 3-chlorobenzoate. These results, combined with the absence of genes related to uptake/chemotaxis for 3-chlorobenzoate located closely to the degradation genes of 3-chlorobenzoate, suggested that NH9 has not fully adapted to the utilization of chlorinated benzoate, unlike benzoate, in nature.

Genes regulated by branched-chain polyamine in the hyperthermophilic archaeon Thermococcus kodakarensis.

Branched-chain polyamine (BCPA) synthase (BpsA), encoded by the bpsA gene, is responsible for the biosynthesis of BCPA in the hyperthermophilic archaeon Thermococcus kodakarensis, which produces N4-bis(aminopropyl)spermidine and spermidine. Here, next-generation DNA sequencing and liquid chromatography-mass spectrometry (LC-MS) were used to perform transcriptomic and proteomic analyses of a T. kodakarensis strain (DBP1) lacking bpsA. Subsequently, the contributions of BCPA to gene transcription (or transcript stabilization) and translation (or protein stabilization) were analyzed. Compared with those in the wild-type strain (KU216) cultivated at 90 °C, the transcript levels of 424 and 21 genes were up- and downregulated in the DBP1 strain, respectively. The expression levels of 12 frequently-used tRNAs were lower in DBP1 cells than KU216 cells, suggesting that BCPA affects translation efficiency in T. kodakarensis. LC-MS analyses of cells grown at 90 °C detected 50 proteins in KU216 cells only, 109 proteins in DBP1 cells only, and 499 proteins in both strains. Notably, the transcript levels of some genes did not correlate with those of the proteins. RNA-seq and RT-qPCR analyses of ten proteins that were detected in KU216 cells only, including three flagellin-related proteins (FlaB2-4) and cytosolic NiFe-hydrogenase subunit alpha (HyhL), revealed that the corresponding transcripts were expressed at higher levels in DBP1 cells than KU216 cells. Electron microscopy analyses showed that flagella formation was disrupted in DBP1 cells at 90 °C, and western blotting confirmed that HyhL expression was eliminated in the DBP1 strain. These results suggest that BCPA plays a regulatory role in gene expression in T. kodakarensis.

Characteristics of physiology of and genomic mutations in aggregation-enhanced mutants of Methanothermobacter sp. CaT2.

The thermophilic hydrogenotrophic methanogen Methanothermobacter sp. CaT2 aggregates by itself. CaT2 is known to have a surface sugar layer and extracellular proteins that may be related to its aggregation. Aggregation-enhanced mutants, CHA001 and CHA002, were isolated after repeated cultivation for more than two years. When treated with proteinase K, CHA001 and CaT2 similarly exhibited a very low degree of aggregation and CHA002 exhibited less aggregation but still retained aggregation, suggesting protein-based aggregation via extracellular proteins in both CHA001 and CHA002, presumably via a putative membrane-bound and extracellularly protruding protein, MTCT_1020, identified previously. Genomic analysis revealed that CHA001 and CHA002 shared a missense mutation of MTCT_1348 and had distinct mutations. These results suggested that the MTCT_1348 mutation provides subsidiary support to the adhesive function of extracellular proteins and that there is an additional mutation(s) in CHA002 for the non-proteinous aggregation capability.

Acidophilic green algal genome provides insights into adaptation to an acidic environment.

Some microalgae are adapted to extremely acidic environments in which toxic metals are present at high levels. However, little is known about how acidophilic algae evolved from their respective neutrophilic ancestors by adapting to particular acidic environments. To gain insights into this issue, we determined the draft genome sequence of the acidophilic green alga Chlamydomonas eustigma and performed comparative genome and transcriptome analyses between Ceustigma and its neutrophilic relative Chlamydomonas reinhardtii The results revealed the following features in Ceustigma that probably contributed to the adaptation to an acidic environment. Genes encoding heat-shock proteins and plasma membrane H+-ATPase are highly expressed in Ceustigma This species has also lost fermentation pathways that acidify the cytosol and has acquired an energy shuttle and buffering system and arsenic detoxification genes through horizontal gene transfer. Moreover, the arsenic detoxification genes have been multiplied in the genome. These features have also been found in other acidophilic green and red algae, suggesting the existence of common mechanisms in the adaptation to acidic environments.

Algal Protein Kinase, Triacylglycerol Accumulation Regulator 1, Modulates Cell Viability and Gametogenesis in Carbon/Nitrogen-Imbalanced Conditions.

Nutrient-deprived microalgae accumulate triacylglycerol (TAG) in lipid droplets. A dual-specificity tyrosine phosphorylation-regulated kinase, TAG accumulation regulator 1 (TAR1) has been shown to be required for acetate-dependent TAG accumulation and the degradation of chlorophyll and photosynthesis-related proteins in photomixotrophic nitrogen (N)-deficient conditions (Kajikawa et�al. 2015). However, this previous report only examined particular condition. Here, we report that in photoautotrophic N-deficient conditions, tar1-1 cells, with a mutation in the TAR1 gene, maintained higher levels of cell viability and lower levels of hydrogen peroxide generation and accumulated higher levels of TAG and starch compared with those of wild type (WT) cells with bubbling of air containing 5% carbon dioxide. Transcriptomic analyses suggested that genes involved in the scavenging of reactive oxygen species are not repressed in tar1-1 cells. In contrast, the mating efficiency and mRNA levels of key regulatory genes for gametogenesis, MID, MTD and FUS, were suppressed in tar1-1 cells. Among the TAR1-dependent phosphopeptides deduced by phosphoproteomic analysis, protein kinases and enzymes related to N assimilation and carbon (C) metabolism are of particular interest. Characterization of these putative downstream factors may elucidate the molecular pathway whereby TAR1 mediates cellular propagation and C and N metabolism in C/N-imbalanced stress conditions.

Pseudofructophilic Leuconostoc citreum Strain F192-5, Isolated from Satsuma Mandarin Peel.

Fructophilic lactic acid bacteria (FLAB), composed of Fructobacillus spp., Lactobacillus kunkeei, and Lactobacillus apinorum, are unique in that they prefer d-fructose over d-glucose as a carbon source. Strain F192-5, isolated from the peel of a satsuma mandarin and identified as Leuconostoc citreum, grows well on d-fructose but poorly on d-glucose and produces mainly lactate and acetate, with trace amounts of ethanol, from the metabolism of d-glucose. These characteristics are identical to those of obligate FLAB. However, strain F192-5 ferments a greater variety of carbohydrates than known FLAB. Comparative analyses of the genomes of strain F192-5 and reference strains of L. citreum revealed no signs of specific gene reductions, especially genes involved in carbohydrate transport and metabolism, in the genome of F192-5. The bifunctional alcohol/acetaldehyde dehydrogenase gene (adhE) is conserved in strain F192-5 but is not transcribed. This is most likely due to a deletion in the promoter region upstream of the adhE gene. Strain F192-5 did, however, ferment d-glucose when transformed with a plasmid containing the allochthonous adhE gene. L. citreum F192-5 is an example of a pseudo-FLAB strain with a deficiency in d-glucose metabolism. This unique phenotypic characteristic appears to be strain specific within the species L. citreum This might be one of the strategies lactic acid bacteria use to adapt to diverse environmental conditions.IMPORTANCE Obligate fructophilic lactic acid bacteria (FLAB) lack the metabolic pathways used in the metabolism of most carbohydrates and differ from other lactic acid bacteria in that they prefer to ferment d-fructose instead of d-glucose. These characteristics are well conserved at the genus or species level. Leuconostoc citreum F192-5 shows similar growth characteristics. However, the strain is metabolically and genomically different from obligate FLAB. This is an example of a strain that evolved a pseudo-FLAB phenotype to adapt to a fructose-rich environment.

Ubr1p-Cup9p-Ptr2p pathway involves in the sensitivity to readthrough compounds negamycin derivatives in budding yeast.

In this study, we found that dipeptide transporter Ptr2p is the putative transporter of read-through compounds (+)-negamycin derivatives TCP-126 and TCP-112, in budding yeast. Ptr2p expression and activity were correlated with the TCP-112 sensitivity, and dipeptide with high affinity to Ptr2p suppressed the TCP-112 activity. These results suggest that dipeptide transporter is one of the determinants of negamycin analogs sensitivity. Abbreviation: PTC: premature termination codon.

CyanoBase: a large-scale update on its 20th anniversary.

The first ever cyanobacterial genome sequence was determined two decades ago and CyanoBase (http://genome.microbedb.jp/cyanobase), the first database for cyanobacteria was simultaneously developed to allow this genomic information to be used more efficiently. Since then, CyanoBase has constantly been extended and has received several updates. Here, we describe a new large-scale update of the database, which coincides with its 20th anniversary. We have expanded the number of cyanobacterial genomic sequences from 39 to 376 species, which consists of 86 complete and 290 draft genomes. We have also optimized the user interface for large genomic data to include the use of semantic web technologies and JBrowse and have extended community-based reannotation resources through the re-annotation of Synechocystis sp. PCC 6803 by the cyanobacterial research community. These updates have markedly improved CyanoBase, providing cyanobacterial genome annotations as references for cyanobacterial research.

Sulfur-Containing Carotenoids from A Marine Coral Symbiont Erythrobacter flavus Strain KJ5.

Erythrobacter flavus strain KJ5 (formerly called Erythrobacter sp. strain KJ5) is a yellowish marine bacterium that was isolated from a hard coral Acropora nasuta in the Karimunjawa Islands, Indonesia. The complete genome sequence of the bacterium has been reported recently. In this study, we examined the carotenoid composition of this bacterium using high-performance liquid chromatography coupled with ESI-MS/MS. We found that the bacterium produced sulfur-containing carotenoids, i.e., caloxanthin sulfate and nostoxanthin sulfate, as the most abundant carotenoids. A new carotenoid zeaxanthin sulfate was detected based on its ESI-MS/MS spectrum. The unique presence of sulfated carotenoids found among the currently known species of the Erythrobacter genus were discussed.

Genetic Analysis of Collective Motility of Paenibacillus sp. NAIST15-1.

Bacteria have developed various motility mechanisms to adapt to a variety of solid surfaces. A rhizosphere isolate, Paenibacillus sp. NAIST15-1, exhibited unusual motility behavior. When spotted onto 1.5% agar media, Paenibacillus sp. formed many colonies, each of which moved around actively at a speed of 3.6 µm/sec. As their density increased, each moving colony began to spiral, finally forming a static round colony. Despite its unusual motility behavior, draft genome sequencing revealed that both the composition and organization of flagellar genes in Paenibacillus sp. were very similar to those in Bacillus subtilis. Disruption of flagellar genes and flagellar stator operons resulted in loss of motility. Paenibacillus sp. showed increased transcription of flagellar genes and hyperflagellation on hard agar media. Thus, increased flagella and their rotation drive Paenibacillus sp. motility. We also identified a large extracellular protein, CmoA, which is conserved only in several Paenibacillus and related species. A cmoA mutant could neither form moving colonies nor move on hard agar media; however, motility was restored by exogenous CmoA. CmoA was located around cells and enveloped cell clusters. Comparison of cellular behavior between the wild type and cmoA mutant indicated that extracellular CmoA is involved in drawing water out of agar media and/or smoothing the cell surface interface. This function of CmoA probably enables Paenibacillus sp. to move on hard agar media.

Chloroplast-mediated regulation of CO2-concentrating mechanism by Ca2+-binding protein CAS in the green alga Chlamydomonas reinhardtii.

Aquatic photosynthetic organisms, including the green alga Chlamydomonas reinhardtii, induce a CO2-concentrating mechanism (CCM) to maintain photosynthetic activity in CO2-limiting conditions by sensing environmental CO2 and light availability. Previously, a novel high-CO2-requiring mutant, H82, defective in the induction of the CCM, was isolated. A homolog of calcium (Ca2+)-binding protein CAS, originally found in Arabidopsis thaliana, was disrupted in H82 cells. Although Arabidopsis CAS is reported to be associated with stomatal closure or immune responses via a chloroplast-mediated retrograde signal, the relationship between a Ca2+ signal and the CCM associated with the function of CAS in an aquatic environment is still unclear. In this study, the introduction of an intact CAS gene into H82 cells restored photosynthetic affinity for inorganic carbon, and RNA-seq analyses revealed that CAS could function in maintaining the expression levels of nuclear-encoded CO2-limiting-inducible genes, including the HCO3- transporters high-light activated 3 (HLA3) and low-CO2-inducible gene A (LCIA). CAS changed its localization from dispersed across the thylakoid membrane in high-CO2 conditions or in the dark to being associated with tubule-like structures in the pyrenoid in CO2-limiting conditions, along with a significant increase of the fluorescent signals of the Ca2+ indicator in the pyrenoid. Chlamydomonas CAS had Ca2+-binding activity, and the perturbation of intracellular Ca2+ homeostasis by a Ca2+-chelator or calmodulin antagonist impaired the accumulation of HLA3 and LCIA. These results suggest that Chlamydomonas CAS is a Ca2+-mediated regulator of CCM-related genes via a retrograde signal from the pyrenoid in the chloroplast to the nucleus.

Conserved two-component Hik34-Rre1 module directly activates heat-stress inducible transcription of major chaperone and other genes in Synechococcus elongatus PCC 7942.

Bacteria and other organisms, including cyanobacteria, employ two-component signal transducing modules comprising histidine kinases and response regulators to acclimate to changing environments. While the number and composition of these modules differ among cyanobacteria, two response regulators that contain DNA binding domains, RpaB and Rre1, are conserved in all sequenced cyanobacterial genomes and are essential for viability. Although RpaB negatively or positively regulates high light and other stress-responsive gene expression, little is known about the function of Rre1. Here, they investigated the direct regulatory targets of Rre1 in the cyanobacterium Synechococcus elongatus PCC 7942. Chromatin immunoprecipitation and high-density tiling array analysis were used to map Rre1 binding sites. The sites included promoter regions for chaperone genes such as dnaK2, groESL-1, groEL-2, hspA and htpG, as well as the group 2 sigma factor gene rpoD2. In vivo and in vitro analyses revealed that Rre1 phosphorylation level, DNA binding activity and adjacent gene transcription increased in response to heat stress. These responses were much diminished in a knock-out mutant of Hik34, a previously identified heat shock regulator. Based on our results, we propose Hik34-Rre1 is the heat shock-responsive signaling module that positively regulates major chaperone and other genes in cyanobacteria.

Transcriptional Activation of Glycogen Catabolism and the Oxidative Pentose Phosphate Pathway by NrrA Facilitates Cell Survival Under Nitrogen Starvation in the Cyanobacterium Synechococcus sp. Strain PCC 7002.

Cyanobacteria respond to nitrogen deprivation by changing cellular metabolism. Glycogen is accumulated within cells to assimilate excess carbon and energy during nitrogen starvation, and inhibition of glycogen synthesis results in impaired nitrogen response and decreased ability to survive. In spite of glycogen accumulation, genes related to glycogen catabolism are up-regulated by nitrogen deprivation. In this study, we found that glycogen catabolism was also involved in acclimation to nitrogen deprivation in the cyanobacterium Synechococcus sp. PCC 7002. The glgP2 gene, encoding glycogen phosphorylase, was induced by nitrogen deprivation, and its expression was regulated by the nitrogen-regulated response regulator A (NrrA), which is a highly conserved transcriptional regulator in cyanobacteria. Activation of glycogen phosphorylase under nitrogen-deprived conditions was abolished by disruption of the nrrA gene, and survival of the nrrA mutant declined. In addition, a glgP2 mutant was highly susceptible to nitrogen starvation. NrrA also regulated expression of the tal-zwf-opcA operon, encoding enzymes of the oxidative pentose phosphate (OPP) pathway, and inactivation of glucose-6-phosphate dehydrogenase, the first enzyme of the OPP pathway, decreased the ability to survive under nitrogen starvation. It was concluded that NrrA facilitates cell survival by activating glycogen degradation and the OPP pathway under nitrogen-deprived conditions.

ParA-like protein influences the distribution of multi-copy chromosomes in cyanobacterium Synechococcus elongatus PCC 7942.

While many bacteria, such as Escherichia coli and Bacillus subtilis, harbour a single-copy chromosome, freshwater cyanobacteria have multiple copies of each chromosome per cell. Although it has been reported that multi-copy chromosomes are evenly distributed along the major axis of the cell in cyanobacterium Synechococcus elongatus PCC 7942, the distribution mechanism of these chromosomes remains unclear. In S. elongatus, the carboxysome, a metabolic microcompartment for carbon fixation that is distributed in a similar manner to the multi-copy chromosomes, is regulated by ParA-like protein (hereafter ParA). To elucidate the role of ParA in the distribution of multi-copy chromosomes, we constructed and analysed ParA disruptant and overexpressing strains of S. elongatus. Our fluorescence in situ hybridization assay revealed that the parA disruptants displayed an aberrant distribution of their multi-copy chromosomes. In the parA disruptant the multiple origin and terminus foci, corresponding to the intracellular position of each chromosomal region, were aggregated, which was compensated by the expression of exogenous ParA from other genomic loci. The parA disruptant is sensitive to UV-C compared to the WT strain. Additionally, giant cells appeared under ParA overexpression at the late stage of growth indicating that excess ParA indirectly inhibits cell division. Screening of the ParA-interacting proteins by yeast two-hybrid analysis revealed four candidates that are involved in DNA repair and cell membrane biogenesis. These results suggest that ParA is involved in the pleiotropic cellular functions with these proteins, while parA is dispensable for cell viability in S. elongatus.

In vitro and in silico characterisation of Lactobacillus paraplantarum D2-1, a starter culture for soymilk fermentation.

Soymilk contains several functional nutrients and is thus a promising ingredient for production of functional foods. The present research aimed to study starter properties, functional characteristics and safety of Lactobacillus paraplantarum D2-1, a promising starter culture for soymilk fermentation. Strain D2-1 actively fermented soymilk within 24 h but had weak activity of additional acid production after 7 d. Succinate and acetoin, which could be linked to flavour and taste, were accumulated in fermented soymilk. In vitro study revealed that the organism has several beneficial properties, including high survival ability in artificial gastric juice, high abilities of mucus adhesion and biofilm formation and production of γ-aminobutyric acid and conjugated linoleic acid, without any significant risks for consumption. Genome sequencing supported the desirable metabolic properties of the strain. These results indicate that L. paraplantarum D2-1 is a suitable starter for soymilk fermentation and is a promising probiotic candidate that can be used safely.