Engineering a Homo-Ethanol Pathway inEscherichia coli: Increased Glycolytic Flux and Levels of Expression of Glycolytic Genes during Xylose Fermentation

ABSTRACT Replacement of the native fermentation pathway inEscherichia coli B with a homo-ethanol pathway fromZymomonas mobilis (pdc andadhB genes) resulted in a 30 to 50% increase in growth rate and glycolytic flux during the anaerobic fermentation of xylose. Gene array analysis was used as a tool to investigate differences in expression levels for the 30 genes involved in xylose catabolism in the parent (strain B) and the engineered strain (KO11). Of the 4,290 total open reading frames, only 8% were expressed at a significantly higher level in KO11 (P < 0.05). In contrast, over half of the 30 genes involved in the catabolism of xylose to pyruvate were expressed at 1.5-fold- to 8-fold-higher levels in KO11. For 14 of the 30 genes, higher expression was statistically significant at the 95% confidence level (xylAB, xylE, xylFG, xylR, rpiA, rpiB, pfkA, fbaA, tpiA, gapA, pgk, and pykA) during active fermentation (6, 12, and 24 h). Values at single time points for only four of these genes (eno, fbaA, fbaB, andtalA) were higher in strain B than in KO11. The relationship between changes in mRNA (cDNA) levels and changes in specific activities was verified for two genes (xylA andxylB) with good agreement. In KO11, expression levels and activities were threefold higher than in strain B for xylose isomerase (xylA) and twofold higher for xylulokinase (xylB). Increased expression of genes involved in xylose catabolism is proposed as the basis for the increase in growth rate and glycolytic flux in ethanologenic KO11.

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Determination of anti-angiogenic therapeutic targets by gene array analysis of differentially expressed angiogenic factors by malignant melanoma

Melanoma Research ◽

10.1097/00008390-200609001-00021 ◽

2006 ◽

Vol 16 (Supplement 1) ◽

pp. S13

Author(s):

L. Kwee ◽

C. Vermeulen ◽

S. Holwerda ◽

A.M.M. Eggermont ◽

T.L.M. ten Hagen

Keyword(s):

Malignant Melanoma ◽

Gene Array ◽

Therapeutic Targets ◽

Angiogenic Factors ◽

Differentially Expressed ◽

Array Analysis ◽

Gene Array Analysis

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Gene-Array Analysis of Glioma Cells After Treatment With an Anti-PKCα siRNA: A General Protocol

Ribozymes and siRNA protocols ◽

10.1385/1-59259-746-7:493 ◽

2004 ◽

pp. 493-500 ◽

Cited By ~ 1

Author(s):

Marianne Leirdal ◽

Mouldy Sioud

Keyword(s):

Gene Array ◽

Glioma Cells ◽

Array Analysis ◽

Gene Array Analysis

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Upregulation of collagens detected by gene array in a model of flow-induced pulmonary vascular remodeling

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00292.2001 ◽

2002 ◽

Vol 282 (2) ◽

pp. H414-H422 ◽

Cited By ~ 29

Author(s):

Meetha Medhora ◽

Michael Bousamra ◽

Daling Zhu ◽

Lewis Somberg ◽

Elizabeth R. Jacobs

Keyword(s):

Vascular Remodeling ◽

Gene Array ◽

Northern Analysis ◽

Western Blots ◽

Pulmonary Vascular Remodeling ◽

Cdna Sequences ◽

Young Pigs ◽

Expression Of Genes ◽

Human Genes ◽

Induced Changes

We recently reported localized increased pulmonary arterial resistance, neointimal lesions, and medial thickening induced by aortopulmonary anastomosis in young pigs. This model was used to investigate changes in expression of genes potentially involved in pulmonary vascular remodeling employing a high throughput Atlas Human Cardiovascular Array carrying ∼600 cardiovascular-related cDNA sequences. Data were confirmed by Northern analysis, Western blots, and histological examination. With the use of lower stringency conditions for hybridization, 56% of the 588 human genes on the array showed visible signal after autoradiography. Approximately 10% of the genes with visible hybridization were altered by shunt-induced high flow. Extracellular matrix and cell adhesion molecules were the most highly represented group of upregulated genes. To our knowledge, our data are the first to demonstrate flow-induced changes in gene expression using a combination of cross species cDNA arrays, homologous hybridization, immunospecific protein, and histology. Our observations expand the list of genes as putative candidates in pulmonary vascular remodeling and support the utility of cross-species microarray analysis in such applications.

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Biochemical and Genetic Analysis of 4-Hydroxypyridine Catabolism in Arthrobacter sp. Strain IN13

Microorganisms ◽

10.3390/microorganisms8060888 ◽

2020 ◽

Vol 8 (6) ◽

pp. 888

Author(s):

Justas Vaitekūnas ◽

Renata Gasparavičiūtė ◽

Jonita Stankevičiūtė ◽

Gintaras Urbelis ◽

Rolandas Meškys

Keyword(s):

Recombinant Expression ◽

Expression Profiles ◽

Peptide Mass Fingerprinting ◽

Sole Source ◽

Open Reading Frames ◽

Expression Of Genes ◽

Peptide Mass ◽

Genes Encoding ◽

Protein Expression Profiles ◽

Reading Frames

N-Heterocyclic compounds are widely spread in the biosphere, being constituents of alkaloids, cofactors, allelochemicals, and artificial substances. However, the fate of such compounds including a catabolism of hydroxylated pyridines is not yet fully understood. Arthrobacter sp. IN13 is capable of using 4-hydroxypyridine as a sole source of carbon and energy. Three substrate-inducible proteins were detected by comparing protein expression profiles, and peptide mass fingerprinting was performed using MS/MS. After partial sequencing of the genome, we were able to locate genes encoding 4-hydroxypyridine-inducible proteins and identify the kpi gene cluster consisting of 16 open reading frames. The recombinant expression of genes from this locus in Escherichia coli and Rhodococcus erytropolis SQ1 allowed an elucidation of the biochemical functions of the proteins. We report that in Arthrobacter sp. IN13, the initial hydroxylation of 4-hydroxypyridine is catalyzed by a flavin-dependent monooxygenase (KpiA). A product of the monooxygenase reaction is identified as 3,4-dihydroxypyridine, and a subsequent oxidative opening of the ring is performed by a hypothetical amidohydrolase (KpiC). The 3-(N-formyl)-formiminopyruvate formed in this reaction is further converted by KpiB hydrolase to 3-formylpyruvate. Thus, the degradation of 4-hydroxypyridine in Arthrobacter sp. IN13 was analyzed at genetic and biochemical levels, elucidating this catabolic pathway.

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Loss of Function inEscherichia coliExposed to Environmentally Relevant Concentrations of Benzalkonium Chloride

Applied and Environmental Microbiology ◽

10.1128/aem.02417-18 ◽

2018 ◽

Vol 85 (4) ◽

Cited By ~ 4

Author(s):

Sarah Forbes ◽

Nicola Morgan ◽

Gavin J. Humphreys ◽

Alejandro Amézquita ◽

Hitesh Mistry ◽

...

Keyword(s):

Escherichia Coli ◽

Growth Rate ◽

Biofilm Formation ◽

Benzalkonium Chloride ◽

Loss Of Function ◽

Free Medium ◽

Content Type ◽

Expression Of Genes ◽

Repeated Passage

ABSTRACTAssessing the risk of resistance associated with biocide exposure commonly involves exposing microorganisms to biocides at concentrations close to the MIC. With the aim of representing exposure to environmental biocide residues,Escherichia coliMG1655 was grown for 20 passages in the presence or absence of benzalkonium chloride (BAC) at 100 ng/liter and 1,000 ng/liter (0.0002% and 0.002% of the MIC, respectively). BAC susceptibility, planktonic growth rates, motility, and biofilm formation were assessed, and differentially expressed genes were determined via transcriptome sequencing. Planktonic growth rate and biofilm formation were significantly reduced (P< 0.001) following BAC adaptation, while BAC minimum bactericidal concentration increased 2-fold. Transcriptomic analysis identified 289 upregulated and 391 downregulated genes after long-term BAC adaptation compared with the respective control organism passaged in BAC-free medium. When the BAC-adapted bacterium was grown in BAC-free medium, 1,052 genes were upregulated and 753 were downregulated. Repeated passage solely in biocide-free medium resulted in 460 upregulated and 476 downregulated genes compared with unexposed bacteria. Long-term exposure to environmentally relevant BAC concentrations increased the expression of genes associated with efflux and reduced the expression of genes associated with outer-membrane porins, motility, and chemotaxis. This was manifested phenotypically through the loss of function (motility). Repeated passage in a BAC-free environment resulted in the upregulation of multiple respiration-associated genes, which was reflected by increased growth rate. In summary, repeated exposure ofE. colito BAC residues resulted in significant alterations in global gene expression that were associated with minor decreases in biocide susceptibility, reductions in growth rate and biofilm formation, and loss of motility.IMPORTANCEExposure to very low concentrations of biocides in the environment is a poorly understood risk factor for antimicrobial resistance. Repeated exposure to trace levels of the biocide benzalkonium chloride (BAC) resulted in loss of function (motility) and a general reduction in bacterial fitness but relatively minor decreases in susceptibility. These changes were accompanied by widespread changes in theEscherichia colitranscriptome. These results demonstrate the importance of including phenotypic characterization in studies designed to assess the risks of biocide exposure.

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DICER1 and PRKRA in Colon Adenocarcinoma

Biomarker Insights ◽

10.4137/bmi.s600 ◽

2008 ◽

Vol 3 ◽

pp. BMI.S600 ◽

Cited By ~ 6

Author(s):

S. Chiosea ◽

M. Acquafondata ◽

J. Luo ◽

SF. Kuan ◽

RR. Seethala

Keyword(s):

Rna Interference ◽

Gene Array ◽

Colon Adenocarcinoma ◽

Clinicopathologic Features ◽

Array Analysis ◽

Gene Array Analysis ◽

Microrna Profile ◽

The Status ◽

And Function ◽

Dicer1 Expression

Differential microRNA expression in colon adenocarcinoma (CA) was previously reported. MicroRNA biogenesis and function requires a set of proteins designated as the microRNA machinery, which includes DICER1 and PRKRA. Loss of heterozygosity at 14q32.13 DICER1 locus was detected in up to 60% of CA cases. The in silico gene array analysis of CA showed down-regulation of DICER1 and an up-regulation of PRKRA. Immunohistochemically, DICER1 expression was abnormal in 65% of CA (95 of 147 cases). PRKRA was deregulated in 70% of CA (32 of 46 cases). Expression of DICER1 and PRKRA was correlated with clinicopathologic features of CA. DICER1 up-regulation was seen more commonly in women. Only 10 of 46 cases immunostained for both DICER1 and PRKRA showed normal levels of both DICER1 and PRKRA. Microsatellite status of 32 cases was determined. Microsatellite instable cases showed DICER1 up-regulation more commonly when compared to microsatellite stable cases; however, this trend was not statistically significant. Abnormal DICER1 and/or PRKRA expression might explain the observed changes in microRNA profile. The status of the endogenous DICER1 and PRKRA in CA may help to predict the response to future RNA interference-based therapy.

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Stability of gene expression in postmortem brain revealed by cDNA gene array analysis

Brain Research ◽

10.1016/s0006-8993(02)02644-6 ◽

2002 ◽

Vol 942 (1-2) ◽

pp. 120-123 ◽

Cited By ~ 29

Author(s):

Stacey A. Trotter ◽

Louis B. Brill II ◽

James P. Bennett

Keyword(s):

Gene Expression ◽

Gene Array ◽

Postmortem Brain ◽

Array Analysis ◽

Gene Array Analysis

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5 PROPYLENE GLYCOL FEEDING SUPPLEMENTATION MODIFIES INSULIN-LIKE GROWTH FACTOR SYSTEM GENE EXPRESSION IN CUMULUS–OOCYTE COMPLEXES AND THE EXPRESSION OF SELECTED CANDIDATE GENES IN EMBRYOS PRODUCED IN VITRO IN FEED-RESTRICTED HEIFERS

Reproduction Fertility and Development ◽

10.1071/rdv27n1ab5 ◽

2015 ◽

Vol 27 (1) ◽

pp. 95

Author(s):

G. Gamarra ◽

C. Ponsart ◽

S. Lacaze ◽

F. Nuttinck ◽

P. Mermillod ◽

...

Keyword(s):

Gene Expression ◽

Growth Rate ◽

Growth Factor ◽

Propylene Glycol ◽

Dietary Supplementation ◽

Cumulus Cells ◽

Oestrous Cycle ◽

Expression Levels ◽

Igf System

Dietary supplementation with propylene glycol (PG) increases the rate of grade 1 embryos produced from feed restricted females (Gamarra et al. 2014 Reprod. Fertil. Dev.). The aim of this study was to evaluate if a PG feeding supplement could modify the expression profile of selected candidate genes that are important for in vitro embryo development and the gene expression patterns of the insulin-like growth factor (IGF) system in oocytes and cumulus cells in feed-restricted heifers. Feed-restricted heifers (n = 16, growth rate of 600 g day–1) received a single daily drench of 400 mL of water (group restricted, R) from Day 1 to Day 9 of a first synchronized oestrous cycle followed by 400 mL of PG from Day 1 to Day 9 of the second synchronized oestrous cycle (group restricted + PG, RPG). Ovum pick-up (OPU) was performed following superovulation, on Day 5 of the oestrous cycle to produce embryos in vitro and on Day 9 without superovulation to obtain oocytes and cumulus cells. The same protocol was used in control animals (n = 6, growth rate of 800 g day–1). Real-time PCR was used to determine the relative abundance of genes involved in lipid metabolism and storage (PLIN2, SCD), energy metabolism (ATP5A1, GLUT1), membrane permeability (AQP3), epigenetic marks (DNMT3a), apoptosis (BAX, TP53), and protein processing (HSPA9B) in grade 1 blastocysts, IGF1, IGF1R, IGFBP2, IGFBP4 in cumulus cells, and IGF1R and IGFBP2 in oocytes. Mann-Whitney nonparametric tests were performed to analyse gene expression results. The expression of PLIN2, ATP5A1, GLUT1, AQP3, DNMT3a, BAX, and HSPA9B were decreased in embryos collected from restricted compared with control animals. The expression levels of these genes were restored when females were supplemented with PG. The expression of TP53 and SCD were not affected. In cumulus cells, the expression levels of IGF1, IGF1R, and IGFBP4 were decreased in restricted compared with control animals. The expression levels of IGF1 and IGF1R were restored with PG supplementation. No differences were observed for the IGFBP2 gene. In the oocytes, no differences were observed for the expression levels of IGF1R and IGFBP2 genes. In conclusion, this work shows for the first time that feed restriction and dietary supplementation by PG in heifers produced changes in gene expression in blastocysts and modified the pattern of the IGF system in cumulus cells. These results suggest the existence of an epigenetic regulation induced by PG during follicular growth, which can regulate the level of gene expression up to the blastocyst stage. In general, PG supplementation of feed-restricted donors restored gene expression at the levels observed after normal feeding.

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Identification of odorant-binding protein genes inGaleruca daurica(Coleoptera: Chrysomelidae) and analysis of their expression profiles

Bulletin of Entomological Research ◽

10.1017/s0007485317000402 ◽

2017 ◽

Vol 107 (4) ◽

pp. 550-561 ◽

Cited By ~ 9

Author(s):

L. Li ◽

Y.-T. Zhou ◽

Y. Tan ◽

X.-R. Zhou ◽

B.-P. Pang

Keyword(s):

Amino Acid ◽

Molecular Mechanisms ◽

Insect Pests ◽

Expression Profiles ◽

Open Reading Frames ◽

Amino Acid Residues ◽

Expression Levels ◽

Molecular Weights ◽

Related Proteins ◽

Odorant Binding

AbstractOdorant-binding proteins (OBPs) play a fundamental role in insect olfaction. In recent years,Galeruca daurica(Joannis) (Coleoptera: Chrysomelidae) has become one of the most important insect pests in the Inner Mongolian grasslands of China. This pest only feeds on the species ofAlliumplants, implying the central role of olfaction in its search for specific host plants. However, the olfaction-related proteins have not been investigated in this beetle. In this study, we identified 29 putative OBP genes, namely GdauOBP1–29, from the transcriptome database ofG. dauricaassembled in our laboratory by using RNA-Seq. All 29 genes had the full-length open reading frames except GdauOBP29, encoding proteins in length from 119 to 202 amino acids with their predicted molecular weights from 12 to 22 kDa with isoelectric points from 3.88 to 8.84. Predicted signal peptides consisting of 15–22 amino acid residues were found in all except GdauOBP6, GdauOBP13 and GdauOBP29. The amino acid sequence identity between the 29 OBPs ranged 8.33–71.83%. GdauOBP1–12 belongs to the Classic OBPs, while the others belong with the Minus-C OBPs. Phylogenetic analysis indicated that GdauOBPs are the closest to CbowOBPs fromColaphellus bowringi. RT-PCR and qRT-PCR analyses showed that all GdauOBPs were expressed in adult antennae, 11 of which with significant differences in their expression levels between males and females. Most GdauOBPs were also expressed in adult heads (without antennae), thoraxes, abdomens, legs and wings. Moreover, the expression levels of the GdauOBPs varied during the different development stages ofG. dauricawith most GdauOBPs expressed highly in the adult antennae but scarcely in eggs and pupae. These results provide insights for further research on the molecular mechanisms of chemical communications inG. daurica.

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