Expression of Chlorite Dismutase and Chlorate Reductase in the Presence of Oxygen and/or Chlorate as the Terminal Electron Acceptor in Ideonella dechloratans

ABSTRACTThe ability of microorganisms to perform dissimilatory (per)chlorate reduction is, for most species, known to be oxygen sensitive. Consequently, bioremediation processes for the removal of oxochlorates will be disturbed if oxygen is present. We measured the expression of chlorite dismutase and chlorate reductase in the presence of different terminal electron acceptors in the chlorate reducerIdeonella dechloratans. Enzyme activity assays and mRNA analyses by real-time quantitative reverse transcription (qRT)-PCR were performed on cell extracts from cells grown aerobically with and without chlorate and on cells grown anaerobically in the presence of chlorate. Our results showed that both chlorite dismutase and chlorate reductase are expressed during aerobic growth. However, transfer to anaerobic conditions with chlorate resulted in significantly enhanced enzyme activities and mRNA levels for both enzymes. Absence of oxygen was necessary for the induction to occur, since chlorate addition under aerobic conditions produced neither increased enzyme activities nor higher relative levels of mRNA. For chlorite dismutase, the observed increase in activity was on the same order of magnitude as the increase in the relative mRNA level, indicating gene regulation at the transcriptional level. However, chlorate reductase showed about 200 times higher enzyme activity in anaerobically induced cells, whereas the increase in mRNA was only about 10-fold, suggesting additional mechanisms influence the enzyme activity.

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Extracellular Enzyme Activity and Microbial Diversity Measured on Seafloor Exposed Basalts from Loihi Seamount Indicate the Importance of Basalts to Global Biogeochemical Cycling

Applied and Environmental Microbiology ◽

10.1128/aem.01038-14 ◽

2014 ◽

Vol 80 (16) ◽

pp. 4854-4864 ◽

Cited By ~ 18

Author(s):

Myrna E. Jacobson Meyers ◽

Jason B. Sylvan ◽

Katrina J. Edwards

Keyword(s):

Enzyme Activity ◽

Microbial Communities ◽

Enzyme Activities ◽

Leucine Aminopeptidase ◽

Extracellular Enzyme ◽

Basaltic Rock ◽

Rrna Gene ◽

Content Type ◽

Shelf Sediments ◽

Potential Enzyme Activity

ABSTRACTSeafloor basalts are widely distributed and host diverse prokaryotic communities, but no data exist concerning the metabolic rates of the resident microbial communities. We present here potential extracellular enzyme activities of leucine aminopeptidase (LAP) and alkaline phosphatase (AP) measured on basalt samples from different locations on Loihi Seamount, HI, coupled with analysis of prokaryotic biomass and pyrosequencing of the bacterial 16S rRNA gene. The community maximum potential enzyme activity (Vmax) of LAP ranged from 0.47 to 0.90 nmol (g rock)−1h−1; theVmaxfor AP was 28 to 60 nmol (g rock)−1h−1. TheKmof LAP ranged from 26 to 33 μM, while theKmfor AP was 2 to 7 μM. Bacterial communities on Loihi basalts were comprised primarily ofAlpha-,Delta-, andGammaproteobacteria,Bacteroidetes, andPlanctomycetes. The putative ability to produce LAP is evenly distributed across the most commonly detected bacterial orders, but the ability to produce AP is likely dominated by bacteria in the ordersXanthomonadales,Flavobacteriales, andPlanctomycetales. The enzyme activities on Loihi basalts were compared to those of other marine environments that have been studied and were found to be similar in magnitude to those from continental shelf sediments and orders of magnitude higher than any measured in the water column, demonstrating that the potential for exposed basalts to transform organic matter is substantial. We propose that microbial communities on basaltic rock play a significant, quantifiable role in benthic biogeochemical processes.

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Gene expression of lipid storage-related enzymes in adipose tissue of the genetically obese Zucker rat. Co-ordinated increase in transcriptional activity and potentiation by hyperinsulinaemia

Biochemical Journal ◽

10.1042/bj2810607 ◽

1992 ◽

Vol 281 (3) ◽

pp. 607-611 ◽

Cited By ~ 23

Author(s):

I Dugail ◽

A Quignard-Boulangé ◽

X Le Liepvre ◽

B Ardouin ◽

M Lavau

Keyword(s):

Adipose Tissue ◽

Mrna Level ◽

Lipid Storage ◽

Zucker Rats ◽

Transcriptional Level ◽

Transcription Rate ◽

Mrna Levels ◽

Zucker Rat ◽

Obese Zucker Rat ◽

Obese Rats

The genetically obese Zucker rat displays excessive fat storage capacity which is due to a tissue-specific increase in the activities of a number of lipid storage-related enzymes in adipose tissue. The aim of this study was to investigate the molecular mechanism responsible for this phenomenon. Lean (Fa/fa) and obese (fa/fa) Zucker rats were studied during the early stages of adipose tissue overdevelopment, both before (at 16 days of age) and after (at 30 days of age) the emergence of hyperinsulinaemia, in order to delineate the effects of the fatty genotype independently of those of hyperinsulinaemia. Lipoprotein lipase (LPL), glycerophosphate dehydrogenase (GPDH), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and malic enzyme (ME) mRNA levels in the adipose tissue of lean and obese rats were assessed by Northern blot analysis, and the relative transcription rates of the corresponding genes were compared in the two genotypes by a nuclear run-on assay. In normoinsulinaemic 16-day-old pre-obese rats, mRNA levels were increased over control values (LPL, 5-fold; ME, 2-fold; GAPDH, 3-fold), in close correlation with genotype-mediated differences in enzyme activities. Stimulation of the transcription rates of the ME and GAPDH genes was observed in obese rats, which could fully account for differences in steady-state mRNA levels. At this age, GPDH activity, mRNA level and transcription rate were similar in the two genotypes. In hyperinsulinaemic 30-day-old obese rats, a 6-7-fold increase in both mRNA and the transcription rate of GPDH emerged, together with an amplification of the genotype-mediated differences observed in younger animals (GAPDH, 6-fold; ME, 7.9-fold; LPL, 10-fold). These results demonstrate that the obese genotype exerts a co-ordinated control on the expression of these genes in adipose tissue, mainly at the transcriptional level. This genotype effect is greatly amplified by the development of hyperinsulinaemia.

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Effect of Soil Salinity on the Expression of Betaine Aldehyde Dehydrogenase in Leaves: Investigation of Hydraulic, Ionic and Biochemical Signals.

Functional Plant Biology ◽

10.1071/pp9920555 ◽

1992 ◽

Vol 19 (5) ◽

pp. 555 ◽

Cited By ~ 12

Author(s):

KF Mccue ◽

AD Hanson

Keyword(s):

Enzyme Activity ◽

Aldehyde Dehydrogenase ◽

Half Life ◽

Mrna Level ◽

State Level ◽

Betaine Aldehyde Dehydrogenase ◽

Mrna Levels ◽

Betaine Aldehyde ◽

Hydraulic Signal ◽

Leaf Disks

Betaine aldehyde dehydrogenase (BADH) catalyses the last step in glycine betaine synthesis. The levels of BADH enzyme and BADH mRNA have previously been shown to be increased several-fold by salt stress. To characterise this induction more thoroughly, BADH mRNA levels and enzyme activities were analysed in leaves of sugar beet plants (Beta vulgaris L.) subjected to different salinisation regimes. Following a salt shock (transfer from 0 to 400 mM NaCI) BADH enzyme activity rose slowly for several days. In contrast, BADH mRNA level first decreased for several hours, and then increased. When salt was leached from the rooting medium of salinised plants, BADH enzyme activity declined, with a half-life of more than 4 days. However, the level of BADH mRNA declined sharply with an apparent half-life of 2 h showing that transcription of the BADH gene or the stability of BADH mRNA in leaves can respond very dynamically to salinity changes around the root. In plants which had been gradually salinised and then held at various NaCl concentrations, the steady state level of enzyme rose continuously between 0 and 500 mM NaCl, whereas that of BADH mRNA reached a plateau at 100 mM NaCl. In general, the observed BADH mRNA fluctuations could not be satisfactorily explained by assuming them to be responses to hydraulic signals. This suggests the participation of a non-hydraulic signal or signals coming from the root. The non-hydraulic signal is unlikely to be NaCl, because leaf disks exposed to salt concentrations typical of the apoplast of salinised leaves did not accumulate BADH mRNA. A biochemical messenger is thus implied. Although abscisic acid application to leaf disks elicited significant increases in BADH mRNA level, these were several-fold smaller than those observed in leaves of intact salinised plants, suggesting the involvement of some other substance.

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Host-Derived Metabolites Modulate Transcription ofSalmonellaGenes Involved in L-Lactate Utilization during Gut Colonization

Infection and Immunity ◽

10.1128/iai.00773-18 ◽

2019 ◽

Vol 87 (4) ◽

Cited By ~ 5

Author(s):

Caroline C. Gillis ◽

Maria G. Winter ◽

Rachael B. Chanin ◽

Wenhan Zhu ◽

Luisella Spiga ◽

...

Keyword(s):

Cell Metabolism ◽

Regulatory Protein ◽

Transcriptional Level ◽

Lactate Oxidation ◽

Terminal Electron Acceptor ◽

Content Type ◽

Gut Colonization ◽

Serovar Typhimurium ◽

Lactate Utilization

ABSTRACTDuringSalmonella entericaserovar Typhimurium infection, host inflammation alters the metabolic environment of the gut lumen to favor the outgrowth of the pathogen at the expense of the microbiota. Inflammation-driven changes in host cell metabolism lead to the release ofl-lactate and molecular oxygen from the tissue into the gut lumen.Salmonellautilizes lactate as an electron donor in conjunction with oxygen as the terminal electron acceptor to support gut colonization. Here, we investigated transcriptional regulation of the respiratoryl-lactate dehydrogenase LldDin vitroand in mouse models ofSalmonellainfection. The two-component system ArcAB repressed transcription ofl-lactate utilization genes under anaerobic conditionsin vitro. The ArcAB-mediated repression oflldDtranscription was relieved under microaerobic conditions. Transcription oflldDwas induced byl-lactate but notd-lactate. A mutant lacking the regulatory protein LldR failed to inducelldDtranscription in response tol-lactate. Furthermore, thelldRmutant exhibited reduced transcription ofl-lactate utilization genes and impaired fitness in murine models of infection. These data provide evidence that the host-derived metabolites oxygen andl-lactate serve as cues forSalmonellato regulate lactate oxidation metabolism on a transcriptional level.

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Syk Plays a Critical Role in the Expression and Activation of IRAK1 in LPS-Treated Macrophages

Mediators of Inflammation ◽

10.1155/2017/1506248 ◽

2017 ◽

Vol 2017 ◽

pp. 1-9 ◽

Cited By ~ 7

Author(s):

Jae Gwang Park ◽

Young-Jin Son ◽

Byong Chul Yoo ◽

Woo Seok Yang ◽

Ji Hye Kim ◽

...

Keyword(s):

Tyrosine Kinase ◽

Protein Tyrosine Kinase ◽

Interleukin 1 ◽

Mrna Level ◽

Critical Role ◽

Primary Response ◽

Transcriptional Level ◽

Mrna Levels ◽

Treatment Conditions ◽

Lps Treatment

To address how interleukin-1 receptor-associated kinase 1 (IRAK1) is controlled by other enzymes activated by toll-like receptor (TLR) 4, we investigated the possibility that spleen tyrosine kinase (Syk), a protein tyrosine kinase that is activated at an earlier stage during TLR4 activation, plays a central role in regulating the functional activation of IRAK1. Indeed, we found that overexpression of myeloid differentiation primary response gene 88 (MyD88), an adaptor molecule that drives TLR signaling, induced IRAK1 expression and that piceatannol, a Syk inhibitor, successfully suppressed the MyD88-dependent upregulation of IRAK1 under LPS treatment conditions. Interestingly, in Syk-knockout RAW264.7 cells, IRAK1 activity was almost completely blocked after LPS treatment, while providing a Syk-recovery gene to the knockout cells successfully restored IRAK1 expression. According to our measurements of IRAK1 mRNA levels, the transcriptional upregulation of IRAK1 was induced by LPS treatment between 4 and 60 min, and this can be suppressed in Syk knockout cells, providing an effect similar that that seen under piceatannol treatment. The overexpression of Syk reverses this effect and leads to a significantly higher IRAK1 mRNA level. Collectively, our results strongly suggest that Syk plays a critical role in regulating both the activity and transcriptional level of IRAK1.

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Diabetes decreases creatine kinase enzyme activity and mRNA level in the rat heart

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1989.257.4.e573 ◽

1989 ◽

Vol 257 (4) ◽

pp. E573-E577 ◽

Cited By ~ 6

Author(s):

B. K. Popovich ◽

K. R. Boheler ◽

W. H. Dillmann

Keyword(s):

Creatine Kinase ◽

Enzyme Activity ◽

Northern Blot Analysis ◽

Mrna Level ◽

Cdna Probe ◽

High Energy ◽

Diabetic Rats ◽

Mrna Levels ◽

High Energy Phosphates ◽

Contractile Performance

Several of the adenosinetriphosphatase enzymes that are responsible for cardiac muscle contraction rely on high-energy phosphates supplied by the creatine kinase (CK) system. Experimental diabetes mellitus has been shown to cause a decrease in the maximal contractile performance of the heart. We postulated that the decrease in contractile performance may be explained in part by a decrease in CK enzyme activity. To evaluate this possibility, we determined the level of CK activity and isoenzyme distribution in ventricular homogenates from normal, diabetic, and insulin-treated diabetic rats. We found that total CK activity was decreased by 35% in diabetic hearts and that a 66% reduction in the cardiac-specific MB isoenzyme occurs. Using a cDNA probe for CK-muscle (M) RNA in Northern blot analysis, we determined that a 61.1% decrease in CK-M mRNA occurs in diabetes. Chronic insulin therapy for 1 mo restores CK-M mRNA levels and enzyme activity. In conclusion, diabetes-induced CK enzyme decreases are mediated in part by a lower level of CK-M mRNA that codes for the major CK-M subunit protein. Decreased performance of the CK system may contribute to diabetic cardiomyopathy.

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β-Lactams Interfering with PBP1 Induce Panton-Valentine Leukocidin Expression by TriggeringsarAandrotGlobal Regulators of Staphylococcus aureus

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01401-10 ◽

2011 ◽

Vol 55 (7) ◽

pp. 3261-3271 ◽

Cited By ~ 37

Author(s):

Oana Dumitrescu ◽

Priya Choudhury ◽

Sandrine Boisset ◽

Cédric Badiou ◽

Michele Bes ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Antisense Rna ◽

Mrna Level ◽

Mrna Levels ◽

Beta Lactam ◽

Beta Lactams ◽

Content Type ◽

Regulatory Pathways ◽

Panton Valentine Leukocidin ◽

Valentine Leukocidin

ABSTRACTPrevious articles reported that beta-lactam antibiotics increase the expression ofStaphylococcus aureusPanton-Valentine leukocidin (PVL) by activating its transcription. We investigated the mechanisms underlying the inductor effect of beta-lactams on PVL expression by determining targets and regulatory pathways possibly implicated in this process. We measured PVL production in the presence of oxacillin (nonselective), imipenem (penicillin-binding protein 1 [PBP1] selective), cefotaxime (PBP2 selective), cefaclore (PBP3 selective), and cefoxitin (PBP4 selective).In vitro, we observed increased PVL production consistent withluk-PV mRNA levels that were 20 to 25 times higher for community-acquired methicillin-resistantS. aureus(CA-MRSA) cultures treated with PBP1-binding oxacillin and imipenem than for cultures treated with other beta-lactams or no antibiotic at all. This effect was also observedin vivo, with increased PVL mRNA levels in lung tissues from CA-MRSA-infected mice treated with imipenem but not cefoxitin. To confirm the involvement of PBP1 inhibition in this pathway, PBP1 depletion by use of an induciblepbp1antisense RNA showed a dose-dependent relationship between the level ofpbp1antisense RNA and theluk-PV mRNA level. Upon imipenem treatment of exponential-phase cultures, we observed an increasedsarAmRNA level after 30 min of incubation followed by a decreasedrotmRNA level after 1 to 4 h of incubation. Unlike theagrandsaeRSpositive regulators, which were nonessential for PVL induction by beta-lactams, thesarA(positive) androt(negative) PVL regulators were necessary for PVL induction by imipenem. Our results suggest that antibiotics binding to PBP1 increase PVL expression by modulatingsarAandrot, which are essential mediators of the inductor effect of beta-lactams on PVL expression.

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Alternative splicing of chicken fibronectin in embryos and in normal and transformed cells.

Molecular and Cellular Biology ◽

10.1128/mcb.7.12.4297 ◽

1987 ◽

Vol 7 (12) ◽

pp. 4297-4307 ◽

Cited By ~ 110

Author(s):

P A Norton ◽

R O Hynes

Keyword(s):

Alternative Splicing ◽

Mrna Level ◽

Receptor Expression ◽

Transcriptional Level ◽

Cdna Clones ◽

Mrna Levels ◽

Oncogenic Transformation ◽

Rous Sarcoma ◽

Alternatively Spliced ◽

V Region

To study the alternative splicing of fibronectin during embryogenesis and oncogenic transformation, we isolated cDNA clones of chicken fibronectin. The partial amino acid sequence deduced from sequencing of these clones showed that, overall, chicken fibronectin is approximately 80% identical with mammalian fibronectins. However, two of the three known regions of alternative splicing differed from this average. The V region was significantly more divergent, and RNA from embryonic chicken liver showed a pattern of V exon splicing which was distinct from that seen in human or rat fibronectins. In contrast, the EIIIB segment was very highly conserved (96%). As in mammals, this segment and another (EIIIA) were alternatively spliced in a cell-type-specific fashion. EIIIA+ and EIIIB+ species were almost absent in liver but predominated in total embryo RNA at all times from 2.5 to 11 days postfertilization. We also examined the possible contributions of fibronectin splicing and integrin receptor expression to the loss of fibronectin on oncogenic transformation. We detected little change in fibronectin splicing, other than a slight increase in representation of EIIIB+ species in fibroblasts after transformation by Rous sarcoma virus. It was also established that the overall reduction in fibronectin mRNA level observed after transformation was not accompanied by a decrease in integrin mRNA levels, indicating that fibronectin and integrin receptors are not coordinately regulated at the transcriptional level.

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The Expression of mRNA and Enzyme Activity of Deoxycytidine- and Deoxyguanosine Kinases in Cell from Patients with Untreated B-Cell Chronic Lymphocytic Leukemia.

Blood ◽

10.1182/blood.v104.11.4378.4378 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4378-4378

Author(s):

Kourosh Lotfi ◽

Karin Karlsson ◽

Gunnar Juliusson ◽

Curt Petersona ◽

Staffan Eriksson ◽

...

Keyword(s):

Enzyme Activity ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Enzyme Activities ◽

Lymphocytic Leukemia ◽

Phase Iii ◽

Mrna Levels ◽

Purine Analogues ◽

Important Place ◽

Response To Chemotherapy

Abstract Chronic lymphocytic leukemia (CLL) is the most common leukemia in Europe and North America. The purine analogues fludarabine and cladribine have earned an important place in the treatment of CLL. Yet, the alkylating agent chlorambucil still remains as cornerstone of treatment. A gradual shift towards the use of purine analogues as the central component has become apparent however it is of greatest importance to strictly compare the efficacy of fludarabine and cladribine to have a basis for future clinical studies with these agents. In this study 59 patients included in the international Phase III Trial for untreated B-cell CLL, were randomized in order to evaluate the efficacy of chlorambucil, fludarabine, and cladribine, as primary treatment of patients with symptomatic B-cell CLL. Both fludarabine and cladribine are prodrugs and must be phosphorylated intracellularly to monophosphates by the nuclear/cytosol enzyme deoxycytidine kinase (dCK) and possibly by the mitochondrial enzyme deoxyguanosine kinase (dGK). DCK plays a pivotal roll in the activation of fludarabine and cladribine and it has previously been reported that resistance to these drugs is mainly due to deficiency of the dCK. Before treatment, peripheral blood cells were isolated and the activity of dCK and dGK enzymes were analyzed in patient cells together with the dCK and dGK mRNA levels using the real-time quantitative PCR method. The dGK activity was considerably lower (6 fold) than the dCK activity. The enzyme activities in samples from CLL patients varied about 20 and 30 folds for dCK and dGK, respectively. The mRNA expression for dCK and dGK in patients showed also a large inter individual variability from 0,011 to 0,189 and 0,001 to 0,154 respectively. There was no correlation between enzyme activity and mRNA levels in the studied CLL patients. We could not found any spliced mRNA dCK variant in our materiel or any mutation, which could explain the discrepancy between dCK activity, and mRNA levels. In conclusion, the results suggest that dCK and dGK expression is regulated at the (post) translational level in leukemic cells. However, studies of the relationship between enzyme activities and/or mRNA levels with clinical response to chemotherapy are underway and will be reported.

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Candida glabrata Med3 Plays a Role in Altering Cell Size and Budding Index To Coordinate Cell Growth

Applied and Environmental Microbiology ◽

10.1128/aem.00781-18 ◽

2018 ◽

Vol 84 (15) ◽

Cited By ~ 1

Author(s):

Hui Liu ◽

Lulin Kong ◽

Yanli Qi ◽

Xiulai Chen ◽

Liming Liu

Keyword(s):

Cell Growth ◽

Cell Size ◽

Candida Glabrata ◽

Mrna Level ◽

Growth Defect ◽

Mrna Levels ◽

Acetyl Coa ◽

Content Type ◽

Quantitative Expression ◽

Transcriptomics Data

ABSTRACT Candida glabrata is a promising microorganism for the production of organic acids. Here, we report deletion and quantitative-expression approaches to elucidate the role of C. glabrata Med3AB (CgMed3AB), a subunit of the mediator transcriptional coactivator, in regulating cell growth. Deletion of CgMed3AB caused an 8.6% decrease in final biomass based on growth curve plots and 10.5% lower cell viability. Based on transcriptomics data, the reason for this growth defect was attributable to changes in expression of genes involved in pyruvate and acetyl-coenzyme A (CoA)-related metabolism in a Cgmed3abΔ strain. Furthermore, the mRNA level of acetyl-CoA synthetase was downregulated after deleting Cgmed3ab, resulting in 22.8% and 21% lower activity of acetyl-CoA synthetase and cellular acetyl-CoA, respectively. Additionally, the mRNA level of CgCln3, whose expression depends on acetyl-CoA, was 34% lower in this strain. As a consequence, the cell size and budding index in the Cgmed3abΔ strain were both reduced. Conversely, overexpression of Cgmed3ab led to 16.8% more acetyl-CoA and 120% higher CgCln3 mRNA levels, as well as 19.1% larger cell size and a 13.3% higher budding index than in wild-type cells. Taken together, these results suggest that CgMed3AB regulates cell growth in C. glabrata by coordinating homeostasis between cellular acetyl-CoA and CgCln3. IMPORTANCE This study demonstrates that CgMed3AB can regulate cell growth in C. glabrata by coordinating the homeostasis of cellular acetyl-CoA metabolism and the cell cycle cyclin CgCln3. Specifically, we report that CgMed3AB regulates the cellular acetyl-CoA level, which induces the transcription of Cgcln3, finally resulting in alterations to the cell size and budding index. In conclusion, we report that CgMed3AB functions as a wheel responsible for driving cellular acetyl-CoA metabolism, indirectly inducing the transcription of Cgcln3 and coordinating cell growth. We propose that Mediator subunits may represent a vital regulatory target modulating cell growth in C. glabrata.

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