Global Transcriptional Regulator Research Articles

Glutathione Synthesis Regulated by CtrA Protects Ehrlichia chaffeensis From Host Cell Oxidative Stress.

Ehrlichia chaffeensis, a small Gram-negative obligatory intracellular bacterium, infects human monocytes or macrophages, and causes human monocytic ehrlichiosis, one of the most prevalent, life-threatening emerging zoonoses. Reactive oxygen species are produced by the host immune cells in response to bacterial infections. The mechanisms exploited by E. chaffeensis to resist oxidative stress have not been comprehensively demonstrated. Here, we found that E. chaffeensis encodes two functional enzymes, GshA and GshB, to synthesize glutathione that confers E. chaffeensis the oxidative stress resistance, and that the expression of gshA and gshB is upregulated by CtrA, a global transcriptional regulator, upon oxidative stress. We found that in E. chaffeensis, the expression of gshA and gshB was upregulated upon oxidative stress using quantitative RT-PCR. Ehrlichia chaffeensis GshA or GshB restored the ability of Pseudomonas aeruginosa GshA or GshB mutant to cope with oxidative stress, respectively. Recombinant E. chaffeensis CtrA directly bound to the promoters of gshA and gshB, determined with electrophoretic mobility shift assay, and activated the expression of gshA and gshB determined with reporter assay. Peptide nucleic acid transfection of E. chaffeensis, which reduced the CtrA protein level, inhibited the oxidative stress-induced upregulation of gshA and gshB. Our findings provide insights into the function and regulation of the two enzymes critical for E. chaffeensis resistance to oxidative stress and may deepen our understanding of E. chaffeensis pathogenesis and adaptation in hosts.

Extracytoplasmic Function Sigma Factors Governing Production of the Primary Siderophores in Pathogenic Burkholderia Species.

Bacteria respond to changing environments by modulating their gene expression programs. One of the mechanisms by which this may be accomplished is by substituting the primary σ factor with an alternative σ factor belonging to the family of extracytoplasmic function (ECF) σ factors. ECF σ factors are activated only in presence of specific signals, and they direct the RNA polymerase (RNAP) to transcribe a defined subset of genes. One condition, which may trigger the activation of an ECF σ factor, is iron limitation. To overcome iron starvation, bacteria produce and secrete siderophores, which chelate iron and facilitate its cellular uptake. In the genus Burkholderia, which includes several serious human pathogens, uptake of iron is critical for virulence, and expression of biosynthetic gene clusters encoding proteins involved in synthesis and transport of the primary siderophores are under control of an ECF σ factor. This review summarizes mechanisms involved in regulation of these gene clusters, including the role of global transcriptional regulators. Since siderophore-mediated iron acquisition is important for virulence, interference with this process constitutes a viable approach to the treatment of bacterial infections.

Drought attenuates plant defence against bacterial pathogens by suppressing the expression of CBP60g/SARD1 during combined stress.

In nature, plants are frequently exposed to drought and bacterial pathogens simultaneously. However, information on how the drought and defence pathways interact and orchestrate global transcriptional regulation is limited. Here, we show that moderate drought stress enhances the susceptibility of Arabidopsis thaliana to Pseudomonas syringae pv. tomato DC3000. Using transcriptome meta-analysis, we found that drought and bacterial stress antagonistically modulate a large set of genes predominantly involved in salicylic acid (SA) and abscisic acid (ABA) signalling networks. We identified that the levels of SA and ABA are dynamically regulated during the course of stress. Importantly, under combined stress, drought through the ABA pathway downregulates the induction of Calmodulin-binding Protein 60 g (CBP60g) and Systemic Acquired Resistance Deficient 1 (SARD1), two transcription factors crucial for SA production upon bacterial infection. We also identified an important role of NPR1-LIKE PROTEIN 3 and 4 (NPR3/4) transcriptional repressors in the drought-mediated negative regulation of CBP60g/SARD1 expression. Using a genetic approach, we show that CBP60g/SARD1 expression is the key determinant of plant defence against bacterial pathogens under combined stress. Thus, these transcription factors act as critical nodes for the crosstalk between drought and bacterial stress signalling under combined stress in plants.

Protein with negative surface charge distribution, Bnr1, shows characteristics of a DNA-mimic protein and may be involved in the adaptation of Burkholderia cenocepacia.

Adaptation of opportunistic pathogens to their host environment requires reprogramming of a vast array of genes to facilitate survival in the host. Burkholderia cenocepacia, a Gram‐negative bacterium with a large genome of ∼8 Mb that colonizes environmental niches, is exquisitely adaptable to the hypoxic environment of the cystic fibrosis lung and survives in macrophages. We previously identified an immunoreactive acidic protein encoded on replicon 3, BCAS0292. Deletion of the BCAS0292 gene significantly altered the abundance of 979 proteins by 1.5‐fold or more; 19 proteins became undetectable while 545 proteins showed ≥1.5‐fold reduced abundance, suggesting the BCAS0292 protein is a global regulator. Moreover, the ∆BCAS0292 mutant showed a range of pleiotropic effects: virulence and host‐cell attachment were reduced, antibiotic susceptibility was altered, and biofilm formation enhanced. Its growth and survival were impaired in 6% oxygen. In silico prediction of its three‐dimensional structure revealed BCAS0292 presents a dimeric β‐structure with a negative surface charge. The ΔBCAS0292 mutant displayed altered DNA supercoiling, implicated in global regulation of gene expression. Three proteins were identified in pull‐downs with FLAG‐tagged BCAS0292, including the Histone H1‐like protein, HctB, which is recognized as a global transcriptional regulator. We propose that BCAS0292 protein, which we have named Burkholderia negatively surface‐charged regulatory protein 1 (Bnr1), acts as a DNA‐mimic and binds to DNA‐binding proteins, altering DNA topology and regulating the expression of multiple genes, thereby enabling the adaptation of B. cenocepacia to highly diverse environments.

Global pleiotropic effects in adaptively evolved Escherichia coli lacking CRP reveal molecular mechanisms that define the growth physiology.

Evolution facilitates emergence of fitter phenotypes by efficient allocation of cellular resources in conjunction with beneficial mutations. However, system-wide pleiotropic effects that redress the perturbations to the apex node of the transcriptional regulatory networks remain unclear. Here, we elucidate that absence of global transcriptional regulator CRP in Escherichia coli results in alterations in key metabolic pathways under glucose respiratory conditions, favouring stress- or hedging-related functions over growth-enhancing functions. Further, we disentangle the growth-mediated effects from the CRP regulation-specific effects on these metabolic pathways. We quantitatively illustrate that the loss of CRP perturbs proteome efficiency, as evident from metabolic as well as ribosomal proteome fractions, that corroborated with intracellular metabolite profiles. To address how E. coli copes with such systemic defect, we evolved Δcrp mutant in the presence of glucose. Besides acquiring mutations in the promoter of glucose transporter ptsG, the evolved populations recovered the metabolic pathways to their pre-perturbed state coupled with metabolite re-adjustments, which altogether enabled increased growth. By contrast to Δcrp mutant, the evolved strains remodelled their proteome efficiency towards biomass synthesis, albeit at the expense of carbon efficiency. Overall, we comprehensively illustrate the genetic and metabolic basis of pleiotropic effects, fundamental for understanding the growth physiology.

Morphological Phenotypes, Cell Division, and Gene Expression of Escherichia coli under High Concentration of Sodium Sulfate

Sodium and sulfate ions are among the suggested abundant ions on Europa, a moon of Jupiter. In order to investigate the potential habitability of Europa, we study the effects of sodium sulfate (Na2SO4) on a non-halophilic bacterium by subjecting Escherichia coli (E. coli) to a wide range of Na2SO4 concentrations (0–1.0 m). We discover that, as the concentration of sodium sulfate increases, the biomass doubling time increases and the cell growth is completely inhibited at m Na2SO4. Furthermore, we find that E. coli exhibits three distinct morphological phenotypes—(i) shortened, (ii) normal, and (iii) elongated/filamented cells at m and m Na2SO4. We have examined the expression of different genes involved in sodium and sulfate transport (nhaA, nhaB, cysZ, sbp), osmotically driven transport of water (aqpZ), sulfate metabolism (cysN), fatty acid production (fabA), and a global transcriptional regulator (osmZ). Our results suggest that the expression of these genes is not affected significantly at high concentrations of sodium sulfate in the exponential growth phase. Using our experimental data and the existing data in the literature, we show that the osmotic pressure difference may play a major role in determining the growth inhibition of E. coli and B. subtilis at high concentrations of salt.

Overexpression of a novel gene Aokap2 affects the growth and kojic acid production in Aspergillus oryzae.

Aspergillus oryzae is an industrially important filamentous fungus for the production of fermentative food, commercial enzyme and valuable secondary metabolites. Although the whole genome of A. oryzae has been sequenced in 2005, there is currently not enough research on functional genes that affect the growth and secondary metabolites of A. oryzae. This study aimed to identify and characterize functional genes involved in the growth and secondary metabolites of A. oryzae. Our previous work on the developmental transcriptome of A. oryzae found that an uncharacterized gene Aokap2 was repressed during the development of A. oryzae. In this study, the gene expression pattern was verified by qRT-PCR. Phylogenetic analysis revealed that AoKAP2 has the species specificity of Aspergillus. Furthermore, Aokap2 was overexpressed using the A. oryzae amyB promoter and overexpression of Aokap2 caused the inhibition in mycelium growth, conidia formation and biomass. Additionally, overexpression of Aokap2 increased the production of kojic acid. In accordance with the enhanced kojic acid, the overexpression of Aokap2 led to elevated transcription levels of the key kojic acid synthesis gene kojA and the global transcriptional regulator gene of secondary metabolism laeA. Moreover, the expression of Aokap2 was down-regulated significantly in the laeA mutant. Meanwhile, overexpression of Aokap2 in the kojA disrupted strain resulted in a ΔkojA strain-like phenotype with significant inhibition in kojic acid production. Taken together, these data suggest that a novel gene Aokap2 is involved in the growth and overexpression of Aokap2 increased kojic acid production through affecting the expression of laeA and kojA. The identification of Aokap2 provides a new target for genetic modification of the growth and the production of kojic acid in A. oryzae.

Improving phosphate use efficiency in the aquatic crop watercress (Nasturtium officinale).

Watercress is a nutrient-dense leafy green crop, traditionally grown in aquatic outdoor systems and increasingly seen as well-suited for indoor hydroponic systems. However, there is concern that this crop has a detrimental impact on the environment through direct phosphate additions causing environmental pollution. Phosphate-based fertilisers are supplied to enhance crop yield, but their use may contribute to eutrophication of waterways downstream of traditional watercress farms. One option is to develop a more phosphate use efficient (PUE) crop. This review identifies the key traits for this aquatic crop (the ideotype), for future selection, marker development and breeding. Traits identified as important for PUE are (i) increased root surface area through prolific root branching and adventitious root formation, (ii) aerenchyma formation and root hair growth. Functional genomic traits for improved PUE are (iii) efficacious phosphate remobilisation and scavenging strategies and (iv) the use of alternative metabolic pathways. Key genomic targets for this aquatic crop are identified as: PHT phosphate transporter genes, global transcriptional regulators such as those of the SPX family and genes involved in galactolipid and sulfolipid biosynthesis such as MGD2/3, PECP1, PSR2, PLDζ1/2 and SQD2. Breeding for enhanced PUE in watercress will be accelerated by improved molecular genetic resources such as a full reference genome sequence that is currently in development.

Integrative analysis of macrophage ribo-Seq and RNA-Seq data define glucocorticoid receptor regulated inflammatory response genes into distinct regulatory classes

Glucocorticoids such as dexamethasone (Dex) are widely used to treat both acute and chronic inflammatory conditions. They regulate immune responses by dampening cell-mediated immunity in a glucocorticoid receptor (GR)-dependent manner, by suppressing the expression of pro-inflammatory cytokines and chemokines and by stimulating the expression of anti-inflammatory mediators. Despite its evident clinical benefit, the mechanistic underpinnings of the gene regulatory networks transcriptionally controlled by GR in a context-specific manner remain mysterious. Next generation sequencing methods such mRNA sequencing (RNA-seq) and Ribosome profiling (ribo-seq) provide tools to investigate the transcriptional and post-transcriptional mechanisms that govern gene expression. Here, we integrate matched RNA-seq data with ribo-seq data from human acute monocytic leukemia (THP-1) cells treated with the TLR4 ligand lipopolysaccharide (LPS) and with Dex, to investigate the global transcriptional and translational regulation (translational efficiency, ΔTE) of Dex-responsive genes. We find that the expression of most of the Dex-responsive genes are regulated at both the transcriptional and the post-transcriptional level, with the transcriptional changes intensified on the translational level. Overrepresentation pathway analysis combined with STRING protein network analysis and manual functional exploration, identified these genes to encode immune effectors and immunomodulators that contribute to macrophage-mediated immunity and to the maintenance of macrophage-mediated immune homeostasis. Further research into the translational regulatory network underlying the GR anti-inflammatory response could pave the way for the development of novel immunomodulatory therapeutic regimens with fewer undesirable side effects.

Structural analyses of the AAA+ ATPase domain of the transcriptional regulator GtrR in the BDSF quorum-sensing system in Burkholderia cenocepacia.

Global transcriptional regulator downstream RpfR (GtrR) is a key downstream regulator for quorum-sensing signaling molecule cis-2-dodecenoic acid (BDSF). As a bacterial enhancer-binding protein (bEBP), GtrR is composed of an N-terminal receiver domain, a central ATPases associated with diverse cellular activities (AAA+) ATPase σ54 -interaction domain, and a C-terminal helix-turn-helix DNA-binding domain. In this work, we solved its AAA+ ATPase domain in both apo and GTP-bound forms. The structure revealed how GtrR specifically recognizes GTP. In addition, we also revealed that GtrR has moderate GTPase activity in vitro in the absence of its activation signal. Finally, we found the residues K170, D236, R311, and R357 in GtrR that are crucial to its biological function, any single mutation leading to completely abolishing GtrR activity.

CRYPTOCHROMES promote daily protein homeostasis.

The daily organisation of most mammalian cellular functions is attributed to circadian regulation of clock‐controlled protein expression, driven by daily cycles of CRYPTOCHROME‐dependent transcriptional feedback repression. To test this, we used quantitative mass spectrometry to compare wild‐type and CRY‐deficient fibroblasts under constant conditions. In CRY‐deficient cells, we found that temporal variation in protein, phosphopeptide, and K+ abundance was at least as great as wild‐type controls. Most strikingly, the extent of temporal variation within either genotype was much smaller than overall differences in proteome composition between WT and CRY‐deficient cells. This proteome imbalance in CRY‐deficient cells and tissues was associated with increased susceptibility to proteotoxic stress, which impairs circadian robustness, and may contribute to the wide‐ranging phenotypes of CRY‐deficient mice. Rather than generating large‐scale daily variation in proteome composition, we suggest it is plausible that the various transcriptional and post‐translational functions of CRY proteins ultimately act to maintain protein and osmotic homeostasis against daily perturbation.

The Type III Effectome of the Symbiotic Bradyrhizobiumvignae Strain ORS3257.

Many Bradyrhizobium strains are able to establish a Nod factor-independent symbiosis with the leguminous plant Aeschynomene indica by the use of a type III secretion system (T3SS). Recently, an important advance in the understanding of the molecular factors supporting this symbiosis has been achieved by the in silico identification and functional characterization of 27 putative T3SS effectors (T3Es) of Bradyrhizobium vignae ORS3257. In the present study, we experimentally extend this catalog of T3Es by using a multi-omics approach. Transcriptome analysis under non-inducing and inducing conditions in the ORS3257 wild-type strain and the ttsI mutant revealed that the expression of 18 out of the 27 putative effectors previously identified, is under the control of TtsI, the global transcriptional regulator of T3SS and T3Es. Quantitative shotgun proteome analysis of culture supernatant in the wild type and T3SS mutant strains confirmed that 15 of the previously determined candidate T3Es are secreted by the T3SS. Moreover, the combined approaches identified nine additional putative T3Es and one of them was experimentally validated as a novel effector. Our study underscores the power of combined proteome and transcriptome analyses to complement in silico predictions and produce nearly complete effector catalogs. The establishment of the ORS3257 effectome will form the basis for a full appraisal of the symbiotic properties of this strain during its interaction with various host legumes via different processes.

OsnR is an autoregulatory negative transcription factor controlling redox-dependent stress responses in Corynebacterium glutamicum

BackgroundCorynebacterium glutamicum is used in the industrial production of amino acids and nucleotides. During the course of fermentation, C. glutamicum cells face various stresses and employ multiple regulatory genes to cope with the oxidative stress. The osnR gene plays a negative regulatory role in redox-dependent oxidative-stress responses, but the underlying mechanism is not known yet.ResultsOverexpression of the osnR gene in C. glutamicum affected the expression of genes involved in the mycothiol metabolism. ChIP-seq analysis revealed that OsnR binds to the promoter region of multiple genes, including osnR and cg0026, which seems to function in the membrane-associated redox metabolism. Studies on the role of the osnR gene involving in vitro assays employing purified OsnR proteins and in vivo physiological analyses have identified that OsnR inhibits the transcription of its own gene. Further, oxidant diamide stimulates OsnR-binding to the promoter region of the osnR gene. The genes affected by the overexpression of osnR have been found to be under the control of σH. In the osnR-overexpressing strain, the transcription of sigH is significantly decreased and the stimulation of sigH transcription by external stress is lost, suggesting that osnR and sigH form an intimate regulatory network.ConclusionsOur study suggests that OsnR not only functions as a transcriptional repressor of its own gene and of those involved in redox-dependent stress responses but also participates in the global transcriptional regulation by controlling the transcription of other master regulators, such as sigH.

Transcriptional Response of Mycobacterium tuberculosis to Cigarette Smoke Condensate.

Smoking is known to be an added risk factor for tuberculosis (TB), with nearly a quarter of the TB cases attributed to cigarette smokers in the 22 countries with the highest TB burden. Many studies have indicated a link between risk of active TB and cigarette smoke. Smoking is also known to significantly decrease TB cure and treatment completion rate and increase mortality rates. Cigarette smoke contains thousands of volatile compounds including carcinogens, toxins, reactive solids, and oxidants in both particulate and gaseous phase. Yet, to date, limited studies have analyzed the impact of cigarette smoke components on Mycobacterium tuberculosis (Mtb), the causative agent of TB. Here we report the impact of cigarette smoke condensate (CSC) on survival, mutation frequency, and gene expression of Mtb in vitro. We show that exposure of virulent Mtb to cigarette smoke increases the mutation frequency of the pathogen and strongly induces the expression of the regulon controlled by SigH—a global transcriptional regulator of oxidative stress. SigH has previously been shown to be required for Mtb to respond to oxidative stress, survival, and granuloma formation in vivo. A high-SigH expression phenotype is known to be associated with greater virulence of Mtb. In patients with pulmonary TB who smoke, these changes may therefore play an important, yet unexplored, role in the treatment efficacy by potentially enhancing the virulence of tubercle bacilli.

Exploring the Meta-regulon of the CRP/FNR Family of Global Transcriptional Regulators in a Partial-Nitritation Anammox Microbiome.

ABSTRACTMicroorganisms must respond to environmental changes to survive, often by controlling transcription initiation. Intermittent aeration during wastewater treatment presents a cyclically changing environment to which microorganisms must react. We used an intermittently aerated bioreactor performing partial nitritation and anammox (PNA) to investigate how the microbiome responds to recurring change. Meta-transcriptomic analysis revealed a dramatic disconnect between the relative DNA abundance and gene expression within the metagenome-assembled genomes (MAGs) of community members, suggesting the importance of transcriptional regulation in this microbiome. To explore how community members responded to cyclic aeration via transcriptional regulation, we searched for homologs of the catabolite repressor protein/fumarate and nitrate reductase regulatory protein (CRP/FNR) family of transcription factors (TFs) within the MAGs. Using phylogenetic analyses, evaluation of sequence conservation in important amino acid residues, and prediction of genes regulated by TFs in the MAGs, we identified homologs of the oxygen-sensing FNR in Nitrosomonas and Rhodocyclaceae, nitrogen-sensing dissimilative nitrate respiration regulator that responds to nitrogen species (DNR) in Rhodocyclaceae, and nitrogen-sensing nitrite and nitric oxide reductase regulator that responds to nitrogen species (NnrR) in Nitrospira MAGs. Our data also predict that CRP/FNR homologs in Ignavibacteria, Flavobacteriales, and Saprospiraceae MAGs sense carbon availability. In addition, a CRP/FNR homolog in a Brocadia MAG was most closely related to CRP TFs known to sense carbon sources in well-studied organisms. However, we predict that in autotrophic Brocadia, this TF most likely regulates a diverse set of functions, including a response to stress during the cyclic aerobic/anoxic conditions. Overall, this analysis allowed us to define a meta-regulon of the PNA microbiome that explains functions and interactions of the most active community members.IMPORTANCE Microbiomes are important contributors to many ecosystems, including ones where nutrient cycling is stimulated by aeration control. Optimizing cyclic aeration helps reduce energy needs and maximize microbiome performance during wastewater treatment; however, little is known about how most microbial community members respond to these alternating conditions. We defined the meta-regulon of a PNA microbiome by combining existing knowledge of how the CRP/FNR family of bacterial TFs respond to stimuli, with metatranscriptomic analyses to characterize gene expression changes during aeration cycles. Our results indicated that, for some members of the community, prior knowledge is sufficient for high-confidence assignments of TF function, whereas other community members have CRP/FNR TFs for which inferences of function are limited by lack of prior knowledge. This study provides a framework to begin elucidating meta-regulons in microbiomes, where pure cultures are not available for traditional transcriptional regulation studies. Defining the meta-regulon can help in optimizing microbiome performance.

Analysis of transcriptional regulators HpaR1 and Clp regulating the expression of glycoside hydrolase-encoding gene in the Xanthomonas campestris pv. campestris.

Xanthomonas campestris pv. campestris (Xcc) is a vascular pathogen that causes black rot in host. It is an important model strain for studying the interaction between the phytopathogen and plants. In Xcc, global transcription regulator HpaR1 that belongs to the GntR family regulates many cellular processes such as the movement and synthesis of extracellular polysaccharides and extracellular enzymes, and is associated with hypersensitive response (HR) and pathogenicity. On the other hand, the global transcriptional regulator Clp regulates the secretion and synthesis of extracellular enzymes and extracellular polysaccharides, and is associated with the pathogenicity of Xanthomonas. Previous studies have shown that both HpaR1 and Clp bind to the promoter region of the glycoside hydrolase encoding gene (named ghy gene). This study investigates the molecular mechanism of the co-regulation of HpaR1 and Clp on the expression of ghy gene. Through electrophoresis mobility shift assay (EMSA), we found that both HpaR1 and Clp bind to the promoter regions of gene ghy in vitro. Both HpaR1 and Clp also bind to the promoter regions of gene ghy in vivo by chromatin immunoprecipitation (ChIP) assays. DNase I footprinting and 5'-RACE assays showed that both HpaR1 and Clp bind to the -35 region upstream of the ghy promoter. The HpaR1 binding site was located upstream of the Clp binding site. RT-qPCR and in vitro transcription assays showed that HpaR1 negatively while Clp positively regulates the transcription of gene ghy. Furthermore, HpaR1 inhibits the activation of Clp on the transcription of gene ghy in vitro. Our findings indicate that HpaR1 and Clp exhibit opposite effect on the transcription of gene ghy. It is speculated that HpaR1 may regulate the expression of gene ghy by inhibiting the activity of RNA polymerase.

Structural basis of transcription activation by the global regulator Spx

Spx is a global transcriptional regulator in Gram-positive bacteria and has been inferred to efficiently activate transcription upon oxidative stress by engaging RNA polymerase (RNAP) and promoter DNA. However, the precise mechanism by which it interacts with RNAP and promoter DNA to initiate transcription remains obscure. Here, we report the cryo-EM structure of an intact Spx-dependent transcription activation complex (Spx–TAC) from Bacillus subtilis at 4.2 Å resolution. The structure traps Spx in an active conformation and defines key interactions accounting for Spx-dependent transcription activation. Strikingly, an oxidized Spx monomer engages RNAP by simultaneously interacting with the C-terminal domain of RNAP alpha subunit (αCTD) and σA. The interface between Spx and αCTD is distinct from those previously reported activators, indicating αCTD as a multiple target for the interaction between RNAP and various transcription activators. Notably, Spx specifically wraps the conserved –44 element of promoter DNA, thereby stabilizing Spx–TAC. Besides, Spx interacts extensively with σA through three different interfaces and promotes Spx-dependent transcription activation. Together, our structural and biochemical results provide a novel mechanistic framework for the regulation of bacterial transcription activation and shed new light on the physiological roles of the global Spx-family transcription factors.

MgaSpn is a negative regulator of capsule and phosphorylcholine biosynthesis and influences the virulence of Streptococcus pneumoniae D39

ABSTRACT Global transcriptional regulators are prevalent in gram-positive pathogens. The transcriptional regulators of the Mga/AtxA family regulate target gene expression by directly binding to the promoter regions, that results in the coordinated expression of virulence factors. The spd_1587 gene of Streptococcus pneumoniae strain D39 encodes MgaSpn, which shares sequence similarity with global transcriptional regulators of the Mga/AtxA family. In this study, we demonstrated that MgaSpn regulates the biosynthesis of the capsule and phosphorylcholine, which play key roles in disease severity in S. pneumoniae infections. MgaSpn directly binds to the cps and lic1 promoters and affects the biosynthesis of the capsule and phosphorylcholine. MgaSpn binds to two specific sites on the promoter of cps, one of which contains the −35 box of the promoter, with high affinity. Consistently, low-molecular-weight capsule components were observed in the mgaSpn-null mutant strain. Moreover, we found that phosphorylcholine content was notably increased in the unencapsulated mgaSpn mutant strain. The mgaSpn null mutant caused more severe systemic disease than the parental strain D39. These findings indicate that the pneumococcal MgaSpn protein can inhibit capsule and phosphorylcholine production, thereby affecting the virulence of S. pneumoniae.

A HU-like protein is required for full virulence in Xanthomonas campestris pv. campestris.

Bacteria harbour several abundant small DNA‐binding proteins known as nucleoid‐associated proteins (NAPs) that contribute to the structure of the bacterial nucleoid as well as to gene regulation. Although the function of NAPs as global transcriptional regulators has been comprehensively studied in the model organism Escherichia coli, their regulatory functions in other bacteria remain relatively poorly understood. Xanthomonas campestris pv. campestris (Xcc) is a gram‐negative bacterium that causes black rot disease in almost all members of the crucifer family. In previous work, we demonstrated that a Fis homologue protein, which we named Fis‐like protein (Flp), contributes to the regulation of virulence, type III secretion, and a series of other phenotypes in Xcc. Here we have examined the role of XC_1355, which is predicted to encode a DNA‐binding protein belonging to the HU family herein named HU‐like protein (Hlp). We show that mutation of XC_1355 in Xcc reduces the virulence, extracellular polysaccharide production, and cell motility, but has no effect on the production of extracellular enzymes and induction of the hypersensitive response. These data together with transcriptome analysis indicate that hlp is a previously uncharacterized gene involved in virulence that has partially overlapping and complementary functions with flp in Xcc, although the two regulators have opposite effects on the expression of genes involved in type III secretion. The findings add to our understanding of the complex regulatory pathways that act to regulate virulence in Xcc.

IGluK-Deep: computational identification of lysine glutarylation sites using deep neural networks with general pseudo amino acid compositions

Lysine glutarylation is a post-translation modification which plays an important regulatory role in a variety of physiological and enzymatic processes including mitochondrial functions and metabolic processes both in eukaryotic and prokaryotic cells. This post-translational modification influences chromatin structure and thereby results in global regulation of transcription, defects in cell-cycle progression, DNA damage repair, and telomere silencing. To better understand the mechanism of lysine glutarylation, its identification in a protein is necessary, however, experimental methods are time-consuming and labor-intensive. Herein, we propose a new computational prediction approach to supplement experimental methods for identification of lysine glutarylation site prediction by deep neural networks and Chou’s Pseudo Amino Acid Composition (PseAAC). We employed well-known deep neural networks for feature representation learning and classification of peptide sequences. Our approach opts raw pseudo amino acid compositions and obsoletes the need to separately perform costly and cumbersome feature extraction and selection. Among the developed deep learning-based predictors, the standard neural network-based predictor demonstrated highest scores in terms of accuracy and all other performance evaluation measures and outperforms majority of previously reported predictors without requiring expensive feature extraction process. iGluK-Deep:Computational Identification of lysine glutarylationsites using deep neural networks with general Pseudo Amino Acid Compositions Sheraz Naseer, Rao Faizan Ali, Yaser Daanial Khan, P.D.D Dominic Communicated by Ramaswamy H. Sarma