scholarly journals CztR, a LysR-Type Transcriptional Regulator Involved in Zinc Homeostasis and Oxidative Stress Defense in Caulobacter crescentus

2010 ◽  
Vol 192 (20) ◽  
pp. 5480-5488 ◽  
Author(s):  
Vânia S. Braz ◽  
José F. da Silva Neto ◽  
Valéria C. S. Italiani ◽  
Marilis V. Marques
Keyword(s):  
Stationary Phase ◽  
Mutant Strain ◽  
Caulobacter Crescentus ◽  
Transcriptional Start Site ◽  
Hypothetical Protein ◽  
Zinc Homeostasis ◽  
Gene Encoding ◽  
Transmembrane Segments ◽  
Zinc Depletion

ABSTRACT Caulobacter crescentus is a free-living alphaproteobacterium that has 11 predicted LysR-type transcriptional regulators (LTTRs). Previously, a C. crescentus mutant strain with a mini-Tn5lacZ transposon inserted into a gene encoding an LTTR was isolated; this mutant was sensitive to cadmium. In this work, a mutant strain with a deletion was obtained, and the role of this LTTR (called CztR here) was evaluated. The transcriptional start site of this gene was determined by primer extension analysis, and its promoter was cloned in front of a lacZ reporter gene. β-Galactosidase activity assays, performed with the wild-type and mutant strains, indicated that this gene is 2-fold induced when cells enter stationary phase and that it is negatively autoregulated. Moreover, this regulator is essential for the expression of the divergent cztA gene at stationary phase, in minimal medium, and in response to zinc depletion. This gene encodes a hypothetical protein containing 10 predicted transmembrane segments, and its expression pattern suggests that it encodes a putative zinc transporter. The cztR strain was also shown to be sensitive to superoxide (generated by paraquat) and to hydrogen peroxide but not to tert-butyl hydroperoxide. The expression of katG and ahpC, but not that of the superoxide dismutase genes, was increased in the cztR mutant. A model is proposed to explain how CztR binding to the divergent regulatory regions could activate cztA expression and repress its own transcription.

scholarly journals CAU-1, a Subclass B3 Metallo-β-Lactamase of Low Substrate Affinity Encoded by an Ortholog Present in the Caulobacter crescentus Chromosome

2002 ◽  
Vol 46 (6) ◽  
pp. 1823-1830 ◽  
Author(s):  
Jean-Denis Docquier ◽  
Fabrizio Pantanella ◽  
Francesco Giuliani ◽  
Maria Cristina Thaller ◽  
Gianfranco Amicosante ◽  
...  
Keyword(s):  
Caulobacter Crescentus ◽  
Hypothetical Protein ◽  
Exchange Chromatography ◽  
Substrate Affinity ◽  
Gene Encoding ◽  
Native Form ◽  
Significant Similarity ◽  
Expression Studies ◽  
Genes Encoding ◽  
Isoelectric Ph

ABSTRACT The sequenced chromosome of Caulobacter crescentus CB15 encodes a hypothetical protein that exhibits significant similarity (30 to 35% identical residues) to metallo-β-lactamases of subclass B3. An allelic variant of this gene (divergent by 3% of its nucleotides) was cloned in Escherichia coli from C. crescentus type strain DSM4727. Expression studies confirmed the metallo-β-lactamase activity of its product, CAU-1. The enzyme produced in E. coli was purified by two ion-exchange chromatography steps. CAU-1 contains a 29-kDa polypeptide with an alkaline isoelectric pH (>9), and unlike the L1 enzyme of Stenotrophomonas maltophilia, the native form is monomeric. Kinetic analysis revealed a preferential activity toward penicillins, carbapenems, and narrow-spectrum cephalosporins, while oxyimino cephalosporins were poorly or not hydrolyzed. Affinities for the various β-lactams were poor overall (Km values were always >100 μM and often >400 μM). The interaction with divalent ion chelators appeared to occur by a mechanism similar to that prevailing in other members of subclass B3. In C. crescentus, the CAU-1 enzyme is produced independently of β-lactam exposure and, interestingly, the bla CAU determinant is bracketed by three other genes, including two genes encoding enzymes involved in methionine biosynthesis and a gene encoding a putative transcriptional regulator, in an operon-like structure. The CAU-1 enzyme is the first example of a metallo-β-lactamase in a member of the α subdivision of the class Proteobacteria.

scholarly journals Effect of the Deletion of qmoABC and the Promoter-Distal Gene Encoding a Hypothetical Protein on Sulfate Reduction in Desulfovibrio vulgaris Hildenborough

2010 ◽  
Vol 76 (16) ◽  
pp. 5500-5509 ◽  
Author(s):  
Grant M. Zane ◽  
Huei-che Bill Yen ◽  
Judy D. Wall
Keyword(s):  
Sulfate Reduction ◽  
Electron Acceptor ◽  
Transmembrane Protein ◽  
Hypothetical Protein ◽  
Desulfovibrio Vulgaris ◽  
Gene Encoding ◽  
Terminal Electron Acceptor ◽  
Sulfate Reducing ◽  
Reducing Bacteria

ABSTRACTThe pathway of electrons required for the reduction of sulfate in sulfate-reducing bacteria (SRB) is not yet fully characterized. In order to determine the role of a transmembrane protein complex suggested to be involved in this process, a deletion inDesulfovibrio vulgarisHildenborough was created by marker exchange mutagenesis that eliminated four genes putatively encoding the QmoABC complex and a hypothetical protein (DVU0851). The Qmo (quinone-interactingmembrane-boundoxidoreductase) complex is proposed to be responsible for transporting electrons to the dissimilatory adenosine-5′-phosphosulfate reductase in SRB. In support of the predicted role of this complex, the deletion mutant was unable to grow using sulfate as its sole electron acceptor with a range of electron donors. To explore a possible role for the hypothetical protein in sulfate reduction, a second mutant was constructed that had lost only the gene that codes for the DVU0851 protein. The second constructed mutant grew with sulfate as the sole electron acceptor; however, there was a lag that was not present with the wild-type or complemented strain. Neither deletion strain was significantly impaired for growth with sulfite or thiosulfate as the terminal electron acceptor. Complementation of the Δ(qmoABC-DVU0851) mutant with all four genes or only theqmoABCgenes restored its ability to grow by sulfate respiration. These results confirmed the prediction that the Qmo complex is in the electron pathway for sulfate reduction and revealed that no other transmembrane complex could compensate when Qmo was lacking.

scholarly journals Role of the Schizosaccharomyces pombe F-Box DNA Helicase in Processing Recombination Intermediates

2005 ◽  
Vol 25 (18) ◽  
pp. 8074-8083 ◽  
Author(s):  
Takashi Morishita ◽  
Fumiko Furukawa ◽  
Chikako Sakaguchi ◽  
Takashi Toda ◽  
Antony M. Carr ◽  
...  
Keyword(s):  
Stationary Phase ◽  
Schizosaccharomyces Pombe ◽  
Genomic Library ◽  
Dna Helicase ◽  
Gene Encoding ◽  
Focus Formation ◽  
Synthetic Lethal ◽  
Recombination Repair ◽  
Γ Rays

ABSTRACT In an effort to identify novel genes involved in recombination repair, we isolated fission yeast Schizosaccharomyces pombe mutants sensitive to methyl methanesulfonate (MMS) and a synthetic lethal with rad2. A gene that complements such mutations was isolated from the S. pombe genomic library, and subsequent analysis identified it as the fbh1 gene encoding the F-box DNA helicase, which is conserved in mammals but not conserved in Saccharomyces cerevisiae. An fbh1 deletion mutant is moderately sensitive to UV, MMS, and γ rays. The rhp51 (RAD51 ortholog) mutation is epistatic to fbh1. fbh1 is essential for viability in stationary-phase cells and in the absence of either Srs2 or Rqh1 DNA helicase. In each case, lethality is suppressed by deletion of the recombination gene rhp57. These results suggested that fbh1 acts downstream of rhp51 and rhp57. Following UV irradiation or entry into the stationary phase, nuclear chromosomal domains of the fbh1Δ mutant shrank, and accumulation of some recombination intermediates was suggested by pulsed-field gel electrophoresis. Focus formation of Fbh1 protein was induced by treatment that damages DNA. Thus, the F-box DNA helicase appears to process toxic recombination intermediates, the formation of which is dependent on the function of Rhp51.

Zinc and Neurodegenerative Disorders

2019 ◽  
pp. 176-193 ◽  
Author(s):  
Olakunle Bamikole Afolabi ◽  
Bose Damilola Balogun ◽  
Omotade Ibidun Oloyede ◽  
Ayodele Jacob Akinyemi
Keyword(s):  
Neurodegenerative Disorders ◽  
Zinc Homeostasis ◽  
Normal Brain ◽  
Brain Functions ◽  
Zinc Depletion ◽  
Intracellular Storage ◽  
Cellular Zinc ◽  
Excess Zinc ◽  
Insight Into

Zinc (Zn) is an essential trace element that is abundantly present in humans. Despite its importance in normal brain functions, alterations in zinc homeostasis cause various neurological pathologies such as dementia, Parkinson's disease, Prion's disease, etc. A growing body of evidence has shown that zinc might play a dual role: in which both zinc depletion and excess zinc cause severe damage and hence neurotoxicity develops. Homeostatic controls are put in place to avoid the accumulation of excess zinc or its deficiency. This cellular zinc homeostasis results from the actions of a coordinated regulation effected by different proteins involved in the uptake, excretion, and intracellular storage or trafficking of zinc. Further investigation has also shown the role of endogenous carnosine (beta-alanyl-L-histidine) in binding excess zinc. Hence, it has the ability to prevent neurotoxicity. Also, the role of a zinc-rich diet cannot be overemphasized. The authors of the chapter, however, provide an insight into the link between zinc homeostasis and neurodegenerative disorders (NDs).

scholarly journals Disruption of sucA, Which Encodes the E1 Subunit of α-Ketoglutarate Dehydrogenase, Affects the Survival of Nitrosomonas europaea in Stationary Phase

2006 ◽  
Vol 188 (1) ◽  
pp. 343-347 ◽  
Author(s):  
Norman G. Hommes ◽  
Elizabeth G. Kurth ◽  
Luis A. Sayavedra-Soto ◽  
Daniel J. Arp
Keyword(s):  
Stationary Phase ◽  
Mutant Strain ◽  
Nitrosomonas Europaea ◽  
Wild Type ◽  
Late Stationary Phase ◽  
Gene Encoding ◽  
Hydroxylamine Oxidoreductase ◽  
Genes Encoding ◽  
Knockout Mutation ◽  
Ketoglutarate Dehydrogenase

ABSTRACT Although Nitrosomonas europaea lacks measurable α-ketoglutarate dehydrogenase activity, the recent completion of the genome sequence revealed the presence of the genes encoding the enzyme. A knockout mutation was created in the sucA gene encoding the E1 subunit. Compared to wild-type cells, the mutant strain showed an accelerated loss of ammonia monooxygenase and hydroxylamine oxidoreductase activities upon entering stationary phase. In addition, unlike wild-type cells, the mutant strain showed a marked lag in the ability to resume growth in response to pH adjustments in late stationary phase.

scholarly journals Transcriptional Organization and In Vivo Role of theEscherichia coli rsd Gene, Encoding the Regulator of RNA Polymerase Sigma D

1999 ◽  
Vol 181 (12) ◽  
pp. 3768-3776 ◽  
Author(s):  
Miki Jishage ◽  
Akira Ishihama
Keyword(s):  
Rna Polymerase ◽  
Stationary Phase ◽  
Null Mutant ◽  
Gene Encoding ◽  
Transcription In Vitro ◽  
Extension Analysis ◽  
Insight Into

ABSTRACT The regulator of sigma D (Rsd) was identified as an RNA polymerase ς70-associated protein in stationary-phaseEscherichia coli with the inhibitory activity of ς70-dependent transcription in vitro (M. Jishage and A. Ishihama, Proc. Natl. Acad. Sci. USA 95:4953–4958, 1998). Primer extension analysis of rsd mRNA indicated the presence of two promoters, ςS-dependent P1 and ς70-dependent P2 with the gearbox sequence. To get insight into the in vivo role of Rsd, the expression of a reporter gene fused to either the ς70- or ςS-dependent promoter was analyzed in the absence of Rsd or the presence of overexpressed Rsd. In the rsd null mutant, the ς70- and ςS-dependent gene expression was increased or decreased, respectively. On the other hand, the ς70- or ςS-dependent transcription was reduced or enhanced, respectively, after overexpression of Rsd. The repression of the ςS-dependent transcription in the rsd mutant is overcome by increased production of the ςS subunit. Together these observations support the prediction that Rsd is involved in replacement of the RNA polymerase ς subunit from ς70 to ςS during the transition from exponential growth to the stationary phase.

scholarly journals Isolation and Characterization of rpoS from a Pathogenic Bacterium, Vibrio vulnificus: Role of σS in Survival of Exponential-Phase Cells under Oxidative Stress

2004 ◽  
Vol 186 (11) ◽  
pp. 3304-3312 ◽  
Author(s):  
Kyung-Je Park ◽  
Min-Jin Kang ◽  
Songhee H. Kim ◽  
Hyun-Jung Lee ◽  
Jae-Kyu Lim ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Stationary Phase ◽  
Vibrio Vulnificus ◽  
Stationary Phases ◽  
Exponential Phase ◽  
Knockout Mutant ◽  
Gene Encoding ◽  
Isolation And Characterization ◽  
Multiple Copies

ABSTRACT A gene homologous to rpoS was cloned from a fatal human pathogen, Vibrio vulnificus. The functional role of rpoS in V. vulnificus was accessed by using an rpoS knockout mutant strain. This mutant was impaired in terms of the ability to survive under oxidative stress, nutrient starvation, UV irradiation, or acidic conditions. The increased susceptibility of the V. vulnificus mutant in the exponential phase to H2O2 was attributed to the reduced activity of hydroperoxidase I (HPI). Although σS synthesis was induced and HPI activity reached the maximal level in the stationary phase, the mutant in the stationary phase showed the same susceptibility to H2O2 as the wild-type strain in the stationary phase. In addition, HPII activity, which is known to be controlled by σS in Escherichia coli, was not detectable in V. vulnificus strains under the conditions tested. The mutant in the exponential phase complemented with multiple copies of either the rpoS or katG gene of V. vulnificus recovered both resistance to H2O2 and HPI activity compared with the control strain. Expression of the katG gene encoding HPI in V. vulnificus was monitored by using a katG::luxAB transcriptional fusion. The expression of this gene was significantly reduced by deletion of σS in both the early exponential and late stationary phases. Thus, σS is necessary for increased synthesis and activity of HPI, and σS is required for exponentially growing V. vulnificus to develop the ability to survive in the presence of H2O2.

scholarly journals The MAL Protein, an Integral Component of Specialized Membranes, in Normal Cells and Cancer

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1065
Author(s):  
Armando Rubio-Ramos ◽  
Leticia Labat-de-Hoz ◽  
Isabel Correas ◽  
Miguel A. Alonso
Keyword(s):  
Epithelial Cells ◽  
Large Body ◽  
Original Description ◽  
Cell Types ◽  
Future Research ◽  
Transmembrane Segments ◽  
Epsilon Toxin ◽  
Proteolipid Proteins ◽  
Polarized Epithelial Cells

The MAL gene encodes a 17-kDa protein containing four putative transmembrane segments whose expression is restricted to human T cells, polarized epithelial cells and myelin-forming cells. The MAL protein has two unusual biochemical features. First, it has lipid-like properties that qualify it as a member of the group of proteolipid proteins. Second, it partitions selectively into detergent-insoluble membranes, which are known to be enriched in condensed cell membranes, consistent with MAL being distributed in highly ordered membranes in the cell. Since its original description more than thirty years ago, a large body of evidence has accumulated supporting a role of MAL in specialized membranes in all the cell types in which it is expressed. Here, we review the structure, expression and biochemical characteristics of MAL, and discuss the association of MAL with raft membranes and the function of MAL in polarized epithelial cells, T lymphocytes, and myelin-forming cells. The evidence that MAL is a putative receptor of the epsilon toxin of Clostridium perfringens, the expression of MAL in lymphomas, the hypermethylation of the MAL gene and subsequent loss of MAL expression in carcinomas are also presented. We propose a model of MAL as the organizer of specialized condensed membranes to make them functional, discuss the role of MAL as a tumor suppressor in carcinomas, consider its potential use as a cancer biomarker, and summarize the directions for future research.

scholarly journals Effects of A6E Mutation on Protein Expression and Supramolecular Assembly of Yeast Asparagine Synthetase

Biology ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 294
Author(s):  
Thunyarat Surasiang ◽  
Chalongrat Noree
Keyword(s):  
Protein Expression ◽  
Stationary Phase ◽  
Lymphoblastic Leukemia ◽  
Energy Levels ◽  
Supramolecular Assembly ◽  
Asparagine Synthetase ◽  
Gene Encoding ◽  
Synthetase Deficiency ◽  
Asparagine Synthetase Deficiency ◽  
Intracellular Energy

Asparagine synthetase deficiency (ASD) has been found to be caused by certain mutations in the gene encoding human asparagine synthetase (ASNS). Among reported mutations, A6E mutation showed the greatest reduction in ASNS abundance. However, the effect of A6E mutation has not yet been tested with yeast asparagine synthetase (Asn1/2p). Here, we constructed a yeast strain by deleting ASN2 from its genome, introducing the A6E mutation codon to ASN1, along with GFP downstream of ASN1. Our mutant yeast construct showed a noticeable decrease of Asn1p(A6E)-GFP levels as compared to the control yeast expressing Asn1p(WT)-GFP. At the stationary phase, the A6E mutation also markedly lowered the assembly frequency of the enzyme. In contrast to Asn1p(WT)-GFP, Asn1p(A6E)-GFP was insensitive to changes in the intracellular energy levels upon treatment with sodium azide during the log phase or fresh glucose at the stationary phase. Our study has confirmed that the effect of A6E mutation on protein expression levels of asparagine synthetase is common in both unicellular and multicellular eukaryotes, suggesting that yeast could be a model of ASD. Furthermore, A6E mutation could be introduced to the ASNS gene of acute lymphoblastic leukemia patients to inhibit the upregulation of ASNS by cancer cells, reducing the risk of developing resistance to the asparaginase treatment.

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