scholarly journals A Novel Bvg-Repressed Promoter Causesvrg-Like Transcription offim3but Does Not Result in the Production of Serotype 3 Fimbriae in Bvg−ModeBordetella pertussis

2018 ◽  
Vol 200 (20) ◽  
Author(s):  
Qing Chen ◽  
Gloria Lee ◽  
Candice Craig ◽  
Victoria Ng ◽  
Paul E. Carlson ◽  
...  
Keyword(s):  
Bordetella Pertussis ◽  
Transcriptional Profiling ◽  
Phase Variation ◽  
Normal Growth ◽  
Growth Conditions ◽  
Anomalous Behavior ◽  
Content Type ◽  
Fimbrial Subunit ◽  
Stable Production ◽  
High Level

ABSTRACTInBordetella pertussis, two serologically distinct fimbriae, FIM2 and FIM3, undergo on/off phase variation independently of each other via variation in the lengths of C stretches in the promoters for their major subunit genes,fim2andfim3. These two promoters are also part of the BvgAS virulence regulon and therefore, if in an on configuration, are activated by phosporylated BvgA (BvgA~P) under normal growth conditions (Bvg+mode) but not in the Bvg−mode, inducible by growth in medium containing MgSO4or other compounds, termed modulators. In theB. pertussisTohama I strain (FIM2+FIM3−), thefim3promoter is in the off state. However, a high level of transcription of thefim3gene is observed in the Bvg−mode. In this study, we provide an explanation for this anomalous behavior by defining a Bvg-repressed promoter (BRP), located approximately 400 bp upstream of the Pfim3transcriptional start. Although transcription of thefim3gene in the Bvg−mode resulted in Fim3 translation, as measured by LacZ translational fusions, no accumulation of Fim3 protein was detectable. We propose that Fim3 protein resulting from translation of mRNA driven by BRP in the Bvg−mode is unstable due to a lack of the fimbrial assembly apparatus encoded by thefimBCgenes, located within thefhaoperon, and therefore is not expressed in the Bvg−mode.IMPORTANCEInBordetella pertussis, the promoter Pfim3-15C for the major fimbrial subunit genefim3is activated by the two-component system BvgAS in the Bvg+mode but not in the Bvg−mode. However, many transcriptional profiling studies have shown thatfim3is transcribed in the Bvg−mode even when Pfim3is in a nonpermissive state (Pfim3-13C), suggesting the presence of a reciprocally regulated element upstream of Pfim3. Here, we provide evidence that BRP is the cause of this anomalous behavior offim3. Although BRP effectsvrg-like transcription offim3in the Bvg−mode, it does not lead to stable production of FIM3 fimbriae, because expression of the chaperone and usher proteins FimB and FimC occurs only in the Bvg+mode.

scholarly journals Identification and Characterization of Chalcone Isomerase Genes Involved in Flavonoid Production in Dracaena cambodiana

2021 ◽  
Vol 12 ◽  
Author(s):  
Jiahong Zhu ◽  
Wan Zhao ◽  
Rongshuang Li ◽  
Dong Guo ◽  
Huiliang Li ◽  
...  
Keyword(s):  
Transcriptional Profiling ◽  
Normal Growth ◽  
Growth Conditions ◽  
Ultraviolet B ◽  
Chalcone Isomerase ◽  
Enzyme Activity Assay ◽  
Dragon’S Blood ◽  
Dracaena Cambodiana ◽  
Key Enzyme

Dragon’s blood is a traditional medicine in which flavonoids are the main bioactive compounds; however, the underlying formation mechanism of dragon’s blood remains largely poorly understood. Chalcone isomerase (CHI) is the key enzyme in the flavonoid biosynthesis pathway. However, CHI family genes are not well understood in Dracaena cambodiana Pierre ex Gagnep, an important source plant of dragon’s blood. In this study, 11 CHI family genes were identified from D. cambodiana, and they were classified into three types. Evolutionary and transcriptional profiling analysis revealed that DcCHI1 and DcCHI4 might be involved in flavonoid production. Both DcCHI1 and DcCHI4 displayed low expression levels in stem under normal growth conditions and were induced by methyl jasmonate (MeJA), 6-benzyl aminopurine (6-BA, synthetic cytokinin), ultraviolet-B (UV-B), and wounding. The recombinant proteins DcCHI1 and DcCHI4 were expressed in Escherichia coli and purified by His-Bind resin chromatography. Enzyme activity assay indicated that DcCHI1 catalyzed the formation of naringenin from naringenin chalcone, while DcCHI4 lacked this catalytic activity. Overexpression of DcCHI1 or DcCHI4 enhanced the flavonoid production in D. cambodiana and tobacco. These findings implied that DcCHI1 and DcCHI4 play important roles in flavonoid production. Thus, our study will not only contribute to better understand the function and expression regulation of CHI family genes involved in flavonoid production in D. cambodiana but also lay the foundation for developing the effective inducer of dragon’s blood.

scholarly journals Stress-Induced MazF-Mediated Proteins in Escherichia coli

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
Akanksha Nigam ◽  
Tamar Ziv ◽  
Adi Oron-Gottesman ◽  
Hanna Engelberg-Kulka
Keyword(s):  
Escherichia Coli ◽  
Proteomic Analysis ◽  
Normal Growth ◽  
Growth Conditions ◽  
Translation Machinery ◽  
Content Type ◽  
E Coli ◽  
Adverse Conditions ◽  
Almost All ◽  
Induced Proteins

ABSTRACT Escherichia coli mazEF is an extensively studied stress-induced toxin-antitoxin (TA) system. The toxin MazF is an endoribonuclease that cleaves RNAs at ACA sites. By that means, under stress, the induced MazF generates a stress-induced translation machinery (STM) composed of MazF-processed mRNAs and selective ribosomes that specifically translate the processed mRNAs. Here, we performed a proteomic analysis of all the E. coli stress-induced proteins that are mediated through the chromosomally borne mazF gene. We show that the mRNAs of almost all of them are characterized by the presence of an ACA site up to 100 nucleotides upstream of the AUG initiator. Therefore, under stressful conditions, induced MazF processes mRNAs that are translated by STM. Furthermore, the presence of the ACA sites far upstream (up to 100 nucleotides) of the AUG initiator may still permit translation by the canonical translation machinery. Thus, such dual-translation mechanisms enable the bacterium under stress also to prepare proteins for immediate functions while coming back to normal growth conditions. IMPORTANCE The stress response, the strategy that bacteria have developed in order to cope up with all kinds of adverse conditions, is so far understood at the level of transcription. Our previous findings of a uniquely modified stress-induced translation machinery (STM) generated in E. coli under stress by the endoribonucleolytic activity of the toxin MazF opens a new chapter in understanding microbial physiology under stress at the translational level. Here, we performed a proteomic analysis of all the E. coli stress-induced proteins that are mediated by chromosomally borne MazF through STM.

scholarly journals Fks1 and Fks2 Are Functionally Redundant but Differentially Regulated in Candida glabrata: Implications for Echinocandin Resistance

2012 ◽  
Vol 56 (12) ◽  
pp. 6304-6309 ◽  
Author(s):  
Santosh K. Katiyar ◽  
Ana Alastruey-Izquierdo ◽  
Kelley R. Healey ◽  
Michael E. Johnson ◽  
David S. Perlin ◽  
...  
Keyword(s):  
Candida Glabrata ◽  
Transcriptional Control ◽  
Mutational Analysis ◽  
Hot Spot ◽  
Normal Growth ◽  
Integral Membrane Protein ◽  
Content Type ◽  
Echinocandin Resistance ◽  
High Level ◽  
The Impact

ABSTRACTThe echinocandins caspofungin, micafungin, and anidulafungin, inhibitors of cell wall β-1,3-glucan synthesis, were recently elevated to first-line agents for treating infections due to the azole-refractory yeastCandida glabrata. InCandida albicans, echinocandin resistance is strictly associated with mutations in Fks1, a large integral membrane protein and putative β-1,3-glucan synthase, while mutations in both Fks1 and its paralog Fks2 (but not Fks3) have been associated with resistance inC. glabrata. To further explore their function, regulation, and role in resistance,C. glabratafksgenes were disrupted and subjected to mutational analysis, and their differential regulation was explored. Anfks1Δfks2Δ double disruptant was not able to be generated; otherwise, all three single and remaining two double disruptants displayed normal growth and echinocandin susceptibility, indicating Fks1-Fks2 redundancy. Selection on echinocandin-containing medium for resistant mutants was dependent on strain background: onlyfks1Δ andfks1Δfks3Δ strains consistently yielded mutants exhibiting high-level resistance, all with Fks2 hot spot 1 mutations. Thus, Fks1-Fks2 redundancy attenuates the rate of resistance; further analysis showed that it also attenuates the impact of resistance-conferring mutations. Growth of thefks1Δ and, especially,fks1Δfks3Δ strains was specifically susceptible to the calcineurin inhibitor FK506. Relatedly, FK506 addition or calcineurin geneCMP2disruption specifically reversed Fks2-mediated resistance of laboratory mutants and clinical isolates. RNA analysis suggests that transcriptional control is not the sole mechanism by which calcineurin modulates Fks2 activity.

scholarly journals Identification and Characterization of a NaCl-Responsive Genetic Locus Involved in Survival during Desiccation in Sinorhizobium meliloti

2013 ◽  
Vol 79 (18) ◽  
pp. 5693-5700 ◽  
Author(s):  
Jan A. C. Vriezen ◽  
Frans J. de Bruijn ◽  
Klaus Nüsslein
Keyword(s):  
Sinorhizobium Meliloti ◽  
Agricultural Soils ◽  
Sequence Similarity ◽  
Genetic Locus ◽  
Model Organism ◽  
Normal Growth ◽  
Growth Conditions ◽  
Amino Acid Sequence Similarity ◽  
Content Type ◽  
Genetic Mechanisms

ABSTRACTTheRhizobiaceaeare a bacterial family of enormous agricultural importance due to the ability of its members to fix atmospheric nitrogen in an intimate relationship with plants. Their survival as naturally occurring soil bacteria in agricultural soils as well as popular seed inocula is affected directly by drought and salinity. Survival after desiccation in the presence of NaCl is enabled by underlying genetic mechanisms in the model organismSinorhizobium meliloti1021. Since salt stress parallels a loss in water activity, the identification of NaCl-responsive loci may identify loci involved in survival during desiccation. This approach enabled identification of the lociasnOandnggby their reduced ability to grow on increased NaCl concentrations, likely due to their inability to produce the osmoprotectant N-acetylglutaminylglutamine (NAGGN). In addition, the mutant harboringngg::Tn5luxABwas affected in its ability to survive desiccation and responded to osmotic stress. The desiccation sensitivity may have been due to secondary functions of Ngg (N-acetylglutaminylglutamine synthetase)-like cell wall metabolism as suggested by the presence of ad-alanine-d-alanine ligase (dAla-dAla) domain and by sensitivity of the mutant to β-lactam antibiotics.asnO::Tn5luxABis expressed during the stationary phase under normal growth conditions. Amino acid sequence similarity to enzymes producing β-lactam inhibitors and increased resistance to β-lactam antibiotics may indicate thatasnOis involved in the production of a β-lactam inhibitor.

scholarly journals Poaceae Orthologs of Rice OsSGL, DUF1645 Domain-Containing Genes, Positively Regulate Drought Tolerance, Grain Length and Weight in Rice

2020 ◽  
Author(s):  
Kai Liu ◽  
Mingjuan Li ◽  
Bin Zhang ◽  
Yanchun Cui ◽  
Xuming Yin ◽  
...  
Keyword(s):  
Drought Stress ◽  
Stress Tolerance ◽  
Molecular Mechanisms ◽  
Normal Growth ◽  
Growth Conditions ◽  
Growth Stages ◽  
Grain Length ◽  
Drought Stress Tolerance ◽  
High Level ◽  
Seed Setting Percentage

Abstract BackgroundGrain yield is a polygenic trait influenced by environmental and genetic interactions at all growth stages of the cereal plant. However, the molecular mechanisms responsible for coordinating the trade-off or cross-talk between these traits remain elusive.ResultsWe characterized the hitherto unknown function of four STRESS_tolerance and GRAIN_LENGTH (OsSGL) Poaceae ortholog genes, all encoding DUF1645 domain-containing proteins, in simultaneous regulation of grain length, grain weight, and drought stress-tolerance in rice. In normal growth conditions, the four ortholog genes were mainly expressed in the developing roots and panicles of the corresponding species. Over-expressing or heterologous high-level expressing Poaceae OsSGL ortholog genes conferred remarkably increased grain length, weight, and seed setting percentage, as well as significantly improved drought-stress tolerance in transgenic rice. Microscopical analysis also showed that the transgene expression promoted cell division and development. RNA-seq and qRT-PCR analyses revealed 73.8% (18,711) overlapped DEGs in all transgenic plants. Moreover, GO and KEGG analyses of different comparisons revealed that the key DEGs participating in drought stress-response belonged to hormone (especially auxin and cytokinin) pathways, and signaling processes were apparently affected in the young panicles. ConclusionTogether, these results suggest the four OsSGL orthologs perform a conserved function in regulating stress-tolerance and cell growth by acting via a hormone biosynthesis and signaling pathway. It may also induce a strategy for tailor-made crop yield improvement.

scholarly journals Overproduction of the MtrCDE Efflux Pump in Neisseria gonorrhoeae Produces Unexpected Changes in Cellular Transcription Patterns

2014 ◽  
Vol 59 (1) ◽  
pp. 724-726 ◽  
Author(s):  
Elizabeth A. Ohneck ◽  
Maira Goytia ◽  
Corinne E. Rouquette-Loughlin ◽  
Sandeep J. Joseph ◽  
Timothy D. Read ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Efflux Pump ◽  
Transcriptional Profiling ◽  
Regulatory Proteins ◽  
Drug Efflux ◽  
Content Type ◽  
Genes Encoding ◽  
High Level ◽  
Cellular Transcription ◽  
Global Consequence

ABSTRACTThe global consequence of drug efflux gene overexpression in bacteria has not been specifically analyzed because strains showing high-level expression typically have mutations in genes encoding regulatory proteins that control other genes. Results from a transcriptional profiling study performed with a strain ofNeisseria gonorrhoeaethat is capable of high-level transcription of themtrCDEefflux pump operon independently of control by cognate regulatory proteins revealed that its overexpression has ramifications for systems other than drug efflux.

scholarly journals Salt-Induced Stress Stimulates a Lipoteichoic Acid-Specific Three-Component Glycosylation System inStaphylococcus aureus

2018 ◽  
Vol 200 (12) ◽  
Author(s):  
Kelvin Kho ◽  
Timothy C. Meredith
Keyword(s):  
Staphylococcus Aureus ◽  
Sigma Factor ◽  
Cell Envelope ◽  
Normal Growth ◽  
Lipoteichoic Acid ◽  
Growth Conditions ◽  
Growth Media ◽  
Alternative Sigma Factor ◽  
Content Type

ABSTRACTLipoteichoic acid (LTA) inStaphylococcus aureusis a poly-glycerophosphate polymer anchored to the outer surface of the cell membrane. LTA has numerous roles in cell envelope physiology, including regulating cell autolysis, coordinating cell division, and adapting to environmental growth conditions. LTA is often further modified with substituents, includingd-alanine and glycosyl groups, to alter cellular function. While the genetic determinants ofd-alanylation have been largely defined, the route of LTA glycosylation and its role in cell envelope physiology have remained unknown, in part due to the low levels of basal LTA glycosylation inS. aureus. We demonstrate here thatS. aureusutilizes a membrane-associated three-component glycosylation system composed of an undecaprenol (Und)N-acetylglucosamine (GlcNAc) charging enzyme (CsbB; SAOUHSC_00713), a putative flippase to transport loaded substrate to the outside surface of the cell (GtcA; SAOUHSC_02722), and finally an LTA-specific glycosyltransferase that adds α-GlcNAc moieties to LTA (YfhO; SAOUHSC_01213). We demonstrate that this system is specific for LTA with no cross recognition of the structurally similar polyribitol phosphate containing wall teichoic acids. We show that while wild-typeS. aureusLTA has only a trace of GlcNAcylated LTA under normal growth conditions, amounts are raised upon either overexpressing CsbB, reducing endogenousd-alanylation activity, expressing the cell envelope stress responsive alternative sigma factor SigB, or by exposure to environmental stress-inducing culture conditions, including growth media containing high levels of sodium chloride.IMPORTANCEThe role of glycosylation in the structure and function ofStaphylococcus aureuslipoteichoic acid (LTA) is largely unknown. By defining key components of the LTA three-component glycosylation pathway and uncovering stress-induced regulation by the alternative sigma factor SigB, the role ofN-acetylglucosamine tailoring during adaptation to environmental stresses can now be elucidated. As thedltand glycosylation pathways compete for the same sites on LTA and induction of glycosylation results in decreasedd-alanylation, the interplay between the two modification systems holds implications for resistance to antibiotics and antimicrobial peptides.

scholarly journals Laboratory Adaptation of Bordetella pertussis Is Associated with the Loss of Type Three Secretion System Functionality

2011 ◽  
Vol 79 (9) ◽  
pp. 3677-3682 ◽  
Author(s):  
M. E. Gaillard ◽  
D. Bottero ◽  
C. E. Castuma ◽  
L. A. Basile ◽  
D. Hozbor
Keyword(s):  
Bordetella Pertussis ◽  
Growth Conditions ◽  
Secretion System ◽  
Clinical Isolates ◽  
Content Type ◽  
Type Three Secretion System ◽  
Associated Proteins ◽  
Type Three Secretion

ABSTRACTAlthoughBordetella pertussiscontains and transcribes loci encoding type III secretion system (TTSS) homologues, expression of TTSS-associated proteins has been reported only for non-laboratory-adapted Irish clinical isolates. Here we confirm such a result for clinical isolates obtained from patients treated in Argentinean hospitals. Moreover, we demonstrate that the expression of TTSS-associated proteins is independent both of the year in which the isolate was obtained and of the types of polymorphic alleles for other virulence factors but is dependent on environmental growth conditions. Interestingly, we observed that TTSS-associated protein expression is lost after successivein vitropassages but becomes operative again when bacteria come into contact with the host. Thisin vivoactivation of TTSS expression was observed not only for clinical isolates previously adapted to the laboratory after successivein vitropassages but also for vaccine strains that did not express the systemin vitro. The reversibility of TTSS expression, demonstrated by its switching off-on when the bacterium comes into contact with the host, appears to be an adaptive response of this pathogen.

scholarly journals The Bacillus subtilis tyrZ Gene Encodes a Highly Selective Tyrosyl-tRNA Synthetase and Is Regulated by a MarR Regulator and T Box Riboswitch

2015 ◽  
Vol 197 (9) ◽  
pp. 1624-1631 ◽  
Author(s):  
Rebecca N. Williams-Wagner ◽  
Frank J. Grundy ◽  
Medha Raina ◽  
Michael Ibba ◽  
Tina M. Henkin
Keyword(s):  
Bacillus Subtilis ◽  
Biofilm Formation ◽  
Normal Growth ◽  
Growth Conditions ◽  
Trna Synthetase ◽  
Cellular Proteins ◽  
Gene Encoding ◽  
Content Type ◽  
T Box ◽  
Related Compounds

ABSTRACTMisincorporation ofd-tyrosine (d-Tyr) into cellular proteins due to mischarging of tRNATyrwithd-Tyr by tyrosyl-tRNA synthetase inhibits growth and biofilm formation ofBacillus subtilis. Furthermore, manyB. subtilisstrains lack a functional gene encodingd-aminoacyl-tRNA deacylase, which prevents misincorporation ofd-Tyr in most organisms.B. subtilishas two genes that encode tyrosyl-tRNA synthetase:tyrSis expressed under normal growth conditions, andtyrZis known to be expressed only whentyrSis inactivated by mutation. We hypothesized thattyrZencodes an alternate tyrosyl-tRNA synthetase, expression of which allows the cell to grow whend-Tyr is present. We show that TyrZ is more selective forl-Tyr overd-Tyr than is TyrS; however, TyrZ is less efficient overall. We also show that expression oftyrZis required for growth and biofilm formation in the presence ofd-Tyr. BothtyrSandtyrZare preceded by a T box riboswitch, buttyrZis found in an operon withywaE, which is predicted to encode a MarR family transcriptional regulator. Expression oftyrZis repressed by YwaE and also is regulated at the level of transcription attenuation by the T box riboswitch. We conclude that expression oftyrZmay allow growth when excessd-Tyr is present.IMPORTANCEAccurate protein synthesis requires correct aminoacylation of each tRNA with the cognate amino acid and discrimination against related compounds.Bacillus subtilisproducesd-Tyr, an analog ofl-Tyr that is toxic when incorporated into protein, during stationary phase. Most organisms utilize ad-aminoacyl-tRNA deacylase to prevent misincorporation ofd-Tyr. This work demonstrates that the increased selectivity of the TyrZ form of tyrosyl-tRNA synthetase may provide a mechanism by whichB. subtilisprevents misincorporation ofd-Tyr in the absence of a functionald-aminoacyl-tRNA deacylase gene.

scholarly journals Hansenula polymorpha Hac1p Is Critical to ProteinN-Glycosylation Activity Modulation, as Revealed by Functional and Transcriptomic Analyses

2015 ◽  
Vol 81 (20) ◽  
pp. 6982-6993 ◽  
Author(s):  
Hye-Yun Moon ◽  
Seon Ah Cheon ◽  
Hyunah Kim ◽  
M. O. Agaphonov ◽  
Ohsuk Kwon ◽  
...  
Keyword(s):  
Mutant Strain ◽  
Leucine Zipper ◽  
Hansenula Polymorpha ◽  
Normal Growth ◽  
Growth Conditions ◽  
Protein Glycosylation ◽  
Bzip Transcription Factor ◽  
Carboxypeptidase Y ◽  
Basic Leucine Zipper ◽  
Content Type

ABSTRACTAggregation of misfolded protein in the endoplasmic reticulum (ER) induces a cellular protective response to ER stress, the unfolded protein response (UPR), which is mediated by a basic leucine zipper (bZIP) transcription factor, Hac1p/Xbp1. In this study, we identified and studied the molecular functions of aHAC1homolog from the thermotolerant yeastHansenula polymorpha(HpHAC1). We found that the HpHAC1mRNA contains a nonconventional intron of 177 bp whose interaction with the 5′ untranslated region is responsible for the translational inhibition of the HpHAC1mRNA. TheH. polymorphahac1-null (Hphac1Δ) mutant strain grew slowly, even under normal growth conditions, and was less thermotolerant than the wild-type (WT) strain. The mutant strain was also more sensitive to cell wall-perturbing agents and to the UPR-inducing agents dithiothreitol (DTT) and tunicamycin (TM). Using comparative transcriptome analysis of the WT and Hphac1Δ strains treated with DTT and TM, we identified HpHAC1-dependent core UPR targets, which included genes involved in protein secretion and processing, particularly those required forN-linked protein glycosylation. Notably, different glycosylation and processing patterns of the vacuolar glycoprotein carboxypeptidase Y were observed in the WT and Hphac1Δ strains. Moreover, overexpression of active HpHac1p significantly increased theN-linked glycosylation efficiency and TM resistance. Collectively, our results suggest that the function of HpHac1p is important not only for UPR induction but also for efficient glycosylation inH. polymorpha.

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