scholarly journals Polyhydroxyalkanoate (PHA) Polymer Accumulation and pha Gene Expression in Phenazine (phz-) and Pyrrolnitrin (prn-) Defective Mutants of Pseudomonas chlororaphis PA23

Polymers ◽  
2018 ◽  
Vol 10 (11) ◽  
pp. 1203 ◽  
Author(s):  
Parveen Sharma ◽  
Riffat Munir ◽  
Jocelyn Plouffe ◽  
Nidhi Shah ◽  
Teresa Kievit ◽  
...  
Keyword(s):  
Gene Expression ◽  
Reducing Power ◽  
Pentose Phosphate ◽  
Pha Synthase ◽  
Pseudomonas Chlororaphis ◽  
Glucose Medium ◽  
Wild Type ◽  
Pha Production ◽  
Phenazine Production ◽  
Altered Gene

Pseudomonas chlororaphis PA23 was isolated from the rhizosphere of soybeans and identified as a biocontrol bacterium against Sclerotinia sclerotiorum, a fungal plant pathogen. This bacterium produces a number of secondary metabolites, including phenazine-1-carboxylic acid, 2-hydroxyphenazine, pyrrolnitrin (PRN), hydrogen cyanide, proteases, lipases and siderophores. It also synthesizes and accumulates polyhydroxyalkanoate (PHA) polymers as carbon and energy storage compounds under nutrient-limited conditions. Pseudomonads like P. chlororaphis metabolize glucose via the Entner-Doudoroff and Pentose Phosphate pathways, which provide precursors for phenazine production. Mutants defective in phenazine (PHZ; PA23-63), PRN (PA23-8), or both (PA23-63-1) accumulated higher concentrations of PHAs than the wild-type strain (PA23) when cultured in Ramsay’s Minimal Medium with glucose or octanoic acid as the carbon source. Expression levels of six pha genes, phaC1, phaZ, phaC2, phaD, phaF, and phaI, were compared with wild type PA23 by quantitative real time polymerase chain reaction (qPCR). The qPCR studies indicated that there was no change in levels of transcription of the PHA synthase genes phaC1 and phaC2 in the phz- (PA23-63) and phz- prn- (PA23-63-1) mutants in glucose medium. There was a significant increase in expression of phaC2 in octanoate medium. Transcription of phaD, phaF and phaI increased significantly in the phz- prn- (PA23-63-1) mutant. Mutations in regulatory genes like gacS, rpoS, and relA/spoT, which affect PHZ and PRN production, also resulted in altered gene expression. The expression of phaC1, phaC2, phaF, and phaI genes was down-regulated significantly in gacS and rpoS mutants. Thus, it appears that PHZ, PRN, and PHA production is regulated by common mechanisms. Higher PHA production in the phz- (PA23-63), prn- (PA23-8), and phz- prn- (PA23-63-1) mutants in octanoic medium could be correlated with higher expression of phaC2. Further, the greater PHA production observed in the phz- and prn- mutants was not due to increased transcription of PHA synthase genes in glucose medium, but due to more accessibility of carbon substrates and reducing power, which were otherwise used for the synthesis of PHZ and PRN.

scholarly journals Role of Sigma Factors in Controlling Global Gene Expression in Light/Dark Transitions in the Cyanobacterium Synechocystis sp. Strain PCC 6803

2007 ◽  
Vol 189 (21) ◽  
pp. 7829-7840 ◽  
Author(s):  
Tina C. Summerfield ◽  
Louis A. Sherman
Keyword(s):  
Gene Expression ◽  
Global Gene Expression ◽  
Growth Conditions ◽  
Day Length ◽  
Regulation Of Transcription ◽  
Wild Type ◽  
Protein Levels ◽  
In The Wild ◽  
Altered Gene ◽  
Pcc 6803

ABSTRACT We report on differential gene expression in the cyanobacterium Synechocystis sp. strain PCC 6803 after light-dark transitions in wild-type, ΔsigB, and ΔsigD strains. We also studied the effect of day length in the presence of glucose on a ΔsigB ΔsigE mutant. Our results indicated that the absence of SigB or SigD predominately altered gene expression in the dark or in the light, respectively. In the light, approximately 350 genes displayed transcript levels in the ΔsigD strain that were different from those of the wild type, with over 200 of these up-regulated in the mutant. In the dark, removal of SigB altered more than 150 genes, and the levels of 136 of these were increased in the mutant compared to those in the wild type. The removal of both SigB and SigE had a major impact on gene expression under mixotrophic growth conditions and resulted in the inability of cells to grow in the presence of glucose with 8-h light and 16-h dark cycles. Our results indicated the importance of group II σ factors in the global regulation of transcription in this organism and are best explained by using the σ cycle paradigm with the stochastic release model described previously (R. A. Mooney, S. A. Darst, and R. Landick, Mol. Cell 20:335-345, 2005). We combined our results with the total protein levels of the σ factors in the light and dark as calculated previously (S. Imamura, S. Yoshihara, S. Nakano, N. Shiozaki, A. Yamada, K. Tanaka, H. Takahashi, M. Asayama, and M. Shirai, J. Mol. Biol. 325:857-872, 2003; S. Imamura, M. Asayama, H. Takahashi, K. Tanaka, H. Takahashi, and M. Shirai, FEBS Lett. 554:357-362, 2003). Thus, we concluded that the control of global transcription is based on the amount of the various σ factors present and able to bind RNA polymerase.

scholarly journals The Ralstonia eutropha PhaR Protein Couples Synthesis of the PhaP Phasin to the Presence of Polyhydroxybutyrate in Cells and Promotes Polyhydroxybutyrate Production

2002 ◽  
Vol 184 (1) ◽  
pp. 59-66 ◽  
Author(s):  
Gregory M. York ◽  
JoAnne Stubbe ◽  
Anthony J. Sinskey
Keyword(s):  
Ralstonia Eutropha ◽  
Regulatory System ◽  
Negative Regulator ◽  
Pha Synthase ◽  
Deletion Strain ◽  
Wild Type ◽  
Heterologous System ◽  
Pathway Models ◽  
Pha Production ◽  
In Cells

ABSTRACT Polyhydroxyalkanoates (PHAs) are polyoxoesters that are produced by many bacteria and that accumulate as intracellular granules. Phasins (PhaP) are proteins that accumulate during PHA synthesis, bind PHA granules, and promote further PHA synthesis. Interestingly, PhaP accumulation seems to be strictly dependent on PHA synthesis, which is catalyzed by the PhaC PHA synthase. Here we have tested the effect of the Ralstonia eutropha PhaR protein on the regulation of PhaP accumulation. R. eutropha strains with phaR, phaC, and/or phaP deletions were constructed, and PhaP accumulation was measured by immunoblotting. The wild-type strain accumulated PhaP in a manner dependent on PHA production, and the phaC deletion strain accumulated no PhaP, as expected. In contrast, both the phaR and the phaR phaC deletion strains accumulated PhaP to higher levels than did the wild type. This result implies that PhaR is a negative regulator of PhaP accumulation and that PhaR specifically prevents PhaP from accumulating in cells that are not producing PHA. Transfer of the R. eutropha phaR, phaP, and PHA biosynthesis (phaCAB) genes into a heterologous system, Escherichia coli, was sufficient to reconstitute the PhaR/PhaP regulatory system, implying that PhaR both regulates PhaP accumulation and responds to PHA directly. Deletion of phaR caused a decrease in PHA yields, and a phaR phaP deletion strain exhibited a more severe PHA defect than a phaP deletion strain, implying that PhaR promotes PHA production and does this at least partially through a PhaP-independent pathway. Models for regulatory roles of PhaR in regulating PhaP and promoting PHA production are presented.

The global regulator GacS regulates biofilm formation in Pseudomonas chlororaphis O6 differently with carbon source

2014 ◽  
Vol 60 (3) ◽  
pp. 133-138 ◽  
Author(s):  
Ji Soo Kim ◽  
Yong Hwan Kim ◽  
Ju Yeon Park ◽  
Anne J. Anderson ◽  
Young Cheol Kim
Keyword(s):  
Carbon Source ◽  
Biofilm Formation ◽  
Defined Medium ◽  
Root Surface ◽  
Gray Mold ◽  
Systemic Resistance ◽  
Pseudomonas Chlororaphis ◽  
Wild Type ◽  
Tomato Leaf Mold ◽  
Phenazine Production

An aggressive root colonizer, Pseudomonas chlororaphis O6 produces various secondary metabolites that impact plant health. The sensor kinase GacS is a key regulator of the expression of biocontrol-related traits. Biofilm formation is one such trait because of its role in root surface colonization. This paper focuses on the effects of carbon source on biofilm formation. In comparison with the wild type, a gacS mutant formed biofilms at a reduced level with sucrose as the major carbon source but at much higher level with mannitol in the defined medium. Biofilm formation by the gacS mutant occurred without phenazine production and in the absence of normal levels of acyl homoserine lactones, which promote biofilms with other pseudomonads. Colonization of tomato roots was similar for the wild type and gacS mutant, showing that any differences in biofilm formation in the rhizosphere were not of consequence under the tested conditions. The reduced ability of the gacS mutant to induce systemic resistance against tomato leaf mold and tomato gray mold was consistent with a lack of production of effectors, such as phenazines. These results demonstrated plasticity in biofilm formation and root colonization in the rhizosphere by a beneficial pseudomonad.

scholarly journals Insertion of the retroposable element, jockey, near the Adh gene of Drosophila melanogaster is associated with altered gene expression

1996 ◽  
Vol 68 (3) ◽  
pp. 203-209 ◽  
Author(s):  
Lisa D. White ◽  
James W. Jacobson
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Insertion Mutation ◽  
Activity Levels ◽  
Mrna Levels ◽  
Wild Type ◽  
Reporter System ◽  
Wild Type Counterpart ◽  
Altered Gene

SummaryThe alcohol dehydrogenase (Adh) gene of Drosophila melanogaster is well suited to be a gene expression reporter system. Adh produces a measurable phenotype at both the enzyme and mRNA levels. We recovered a spontaneous transposable element (TE) insertion mutation near the Adh gene. The insertion is a truncated retroposable element, jockey, inserted upstream of the adult Adh enhancer region. Comparisons between the Adhjockey allele and its direct wild-type ancestral allele were made in an isogenic background (i.e. identical cis and trans factors). Differences in Adhjockey expression compared with the wild-type can be attributed solely to the presence of the jockey element. This jockey insertion results in a decrease in adult mRNA transcript levels in the Adhjockey homozygous lines relative to the wild-type counterpart and accounts for a correlated decrease in alcohol dehydrogenase (ADH) enzyme activity. The larval ADH activity levels are not detectably different.

Gene expression profile of mouse myocardium with transgenic overexpression of A1adenosine receptors

2002 ◽  
Vol 11 (2) ◽  
pp. 81-89 ◽  
Author(s):  
Amy R. Lankford ◽  
Anne M. Byford ◽  
Kevin J. Ashton ◽  
Brent A. French ◽  
Jae K. Lee ◽  
...  
Keyword(s):  
Gene Expression ◽  
Myocardial Ischemia ◽  
Nadh Dehydrogenase ◽  
Glucose Transporter ◽  
Caspase 8 ◽  
Rna Expression ◽  
Wild Type ◽  
Altered Gene Expression ◽  
Apoptotic Genes ◽  
Altered Gene

Transgenic mice with cardiac-specific overexpression of adenosine A1receptors (A1AR) have demonstrated metabolic and functional tolerance to myocardial ischemia. We utilized cDNA microarrays to test the hypothesis that the cardioprotective mechanism(s) of A1overexpression involves altered gene expression. Total RNA extracted from the left ventricles from A1transgenic ( n = 4) and wild-type ( n = 6) mice was hybridized to Affymetrix mgU74A chips. Comparison of RNA expression levels in transgenic to wild-type myocardium revealed ∼636 known genes with expression significantly altered by greater than 25%. We observed increased expressions of genes including NADH dehydrogenase, the GLUT4 glucose transporter, Na-K-ATPase, sarcolemmal KATPchannels, and Bcl-xl in A1AR-overexpressing hearts. We also observed decreased expression of pro-apoptotic genes including a 50% reduction in message level of caspase-8. Protein expression of GLUT4 and caspase-8 was also altered comparable to the differences in gene expression. These data illustrate genes with chronically altered patterns of expression in A1transgenic mouse myocardium that may be related to adenosine receptor overexpression-mediated cardioprotection.

Altered lung gene expression in CCSP-null mice suggests immunoregulatory roles for Clara cells

2001 ◽  
Vol 281 (6) ◽  
pp. L1523-L1530 ◽  
Author(s):  
T. M. Watson ◽  
S. D. Reynolds ◽  
G. W. Mango ◽  
I.-M. Boe ◽  
J. Lund ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expressed Sequence Tag ◽  
Immunoglobulin A ◽  
Secretory Protein ◽  
Clara Cell ◽  
Lymphoid Tissues ◽  
Clara Cells ◽  
Wild Type ◽  
Environmental Agents ◽  
Altered Gene

Clara cell secretory protein (CCSP) is one of the most abundant proteins present in airway lining fluid of mammals. In an effort to elucidate the function of CCSP, we established CCSP-null [CCSP(−/−)] mice and demonstrated altered sensitivity to various environmental agents including oxidant pollutants and microorganisms. Although CCSP deficiency itself may be central to the observed changes in environmental susceptibility, altered lung gene expression associated with CCSP deficiency may contribute to the observed phenotype. To determine whether CCSP deficiency results in altered lung gene expression, high-density cDNA microarrays were used to profile gene expression in the total lung RNA of wild-type and CCSP(−/−) mice. Genes that were differentially expressed between wild-type and CCSP(−/−) mice included a previously nonannotated expressed sequence tag (EST W82219 ) and immunoglobulin A (IgA), both of which were elevated with CCSP deficiency. mRNA expression of EST W82219 and IgA was localized in the lungs of wild-type and CCSP(−/−) mice to airway Clara cells and peribronchial lymphoid tissues, respectively. We conclude that CCSP deficiency is associated with 1) altered gene expression in Clara cells of the conducting airway epithelium and 2) alterations to peribronchial B lymphocytes. These findings identify new roles for Clara cells and their secretions in airway homeostasis.

scholarly journals Genetic engineering of Pseudomonas chlororaphis Lzh-T5 to enhance production of trans-2,3-dihydro-3-hydroxyanthranilic acid

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kaiquan Liu ◽  
Ling Li ◽  
Wentao Yao ◽  
Wei Wang ◽  
Yujie Huang ◽  
...  
Keyword(s):  
Shikimate Pathway ◽  
Pentose Phosphate ◽  
Genetically Engineered ◽  
Regulatory Genes ◽  
Pseudomonas Chlororaphis ◽  
Chiral Materials ◽  
Key Genes ◽  
Phenazine Production ◽  
Hydroxyanthranilic Acid ◽  
Natural Peptides

AbstractTrans-2,3-dihydro-3-hydroxyanthranilic acid (DHHA) is a cyclic β-amino acid used for the synthesis of non-natural peptides and chiral materials. And it is an intermediate product of phenazine production in Pseudomonas spp. Lzh-T5 is a P. chlororaphis strain isolated from tomato rhizosphere found in China. It can synthesize three antifungal phenazine compounds. Disruption the phzF gene of P. chlororaphis Lzh-T5 results in DHHA accumulation. Several strategies were used to improve production of DHHA: enhancing the shikimate pathway by overexpression, knocking out negative regulatory genes, and adding metal ions to the medium. In this study, three regulatory genes (psrA, pykF, and rpeA) were disrupted in the genome of P. chlororaphis Lzh-T5, yielding 5.52 g/L of DHHA. When six key genes selected from the shikimate, pentose phosphate, and gluconeogenesis pathways were overexpressed, the yield of DHHA increased to 7.89 g/L. Lastly, a different concentration of Fe3+ was added to the medium for DHHA fermentation. This genetically engineered strain increased the DHHA production to 10.45 g/L. According to our result, P. chlororaphis Lzh-T5 could be modified as a microbial factory to produce DHHA. This study laid a good foundation for the future industrial production and application of DHHA.

scholarly journals Genetic Engineering of Pseudomonas Chlororaphis Lzh-T5 to Enhance Production of Trans-2,3-Dihydro-3-Hydroxyanthranilic Acid

2020 ◽  
Author(s):  
Ling Li ◽  
Zhenghua Li ◽  
Xuehong Zhang ◽  
Wei Wang ◽  
Yujie Huang ◽  
...  
Keyword(s):  
Metal Ions ◽  
Shikimate Pathway ◽  
Pentose Phosphate ◽  
Genetically Engineered ◽  
Regulatory Genes ◽  
Pseudomonas Chlororaphis ◽  
Key Genes ◽  
Phenazine Production ◽  
Hydroxyanthranilic Acid ◽  
Natural Peptides

Abstract Background: Trans-2,3-dihydro-3-hydroxyanthranilic acid (DHHA) is a cyclic β-amino acid used for the synthesis of non-natural peptides and chiral materials. It is an intermediate product of phenazine production in Pseudomonas spp . Lzh-T5 is a P. chlororaphis strain isolated from tomato rhizosphere found in China. It can synthesize three antifungal phenazine compounds. Results: Disrupting the phzF gene of P. chlororaphis Lzh-T5 results in DHHA accumulation. Several strategies were used to improve production of DHHA: enhancing the shikimate pathway by overexpression, knocking out negative regulatory genes, and adding metal ions to the medium. In this study, three regulatory genes ( psrA , pykF, and rpeA ) were-disrupted in the genome of P. chlororaphis Lzh-T5, yielding 4.55 g/L of DHHA. When six key genes selected from the shikimate, pentose phosphate, and gluconeogenesis pathways were overexpressed, the yield of DHHA increased to 6.89g/L. Fe 3+ was added to the medium for DHHA fermentation. This genetically engineered strain increased the DHHA production to 10.45g/L. Conclusions: P. chlororaphis Lzh-T5 could be modified as a microbial factory to produce DHHA by inactivating phzF , disrupting negative regulatory genes, overexpressing key genes, and adding metal ions to medium for fermentation.

scholarly journals Mice Deficient for Testis-Brain RNA-Binding Protein Exhibit a Coordinate Loss of TRAX, Reduced Fertility, Altered Gene Expression in the Brain, and Behavioral Changes

2003 ◽  
Vol 23 (18) ◽  
pp. 6419-6434 ◽  
Author(s):  
Vargheese Chennathukuzhi ◽  
Joel M. Stein ◽  
Ted Abel ◽  
Stacy Donlon ◽  
Shicheng Yang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Mitotic Cell ◽  
Seminiferous Tubules ◽  
Factor X ◽  
Wild Type ◽  
Altered Gene Expression ◽  
Altered Gene

ABSTRACT Testis-brain RNA-binding protein (TB-RBP), the mouse orthologue of the human protein Translin, is a widely expressed and highly conserved protein with proposed functions in chromosomal translocations, mitotic cell division, and mRNA transport and storage. To better define the biological roles of TB-RBP, we generated mice lacking TB-RBP. Matings between heterozygotes gave rise to viable, apparently normal homozygous mutant mice at a normal Mendelian ratio. The TB-RBP-related and -interacting protein Translin-associated factor X was reduced to 50% normal levels in heterozygotes and was absent in TB-RBP-null animals. The null mice were 10 to 30% smaller than their wild-type or heterozygote littermates at birth and remained so to about 6 to 9 months of age, showed normal B- and T-cell development, and accumulated visceral fat. TB-RBP-null male mice were fertile and sired offspring but had abnormal seminiferous tubules and reduced sperm counts. Null female mice were subfertile and had reduced litter sizes. Microarray analysis of total brain RNA from null and wild-type mice revealed an altered gene expression profile with the up-regulation of 14 genes and the down-regulation of 217 genes out of 12,473 probe sets. Numerous neurotransmitter receptors and ion channels, including γ-aminobutyric acid A receptor α1 and glutamate receptor α3, were strongly down-regulated. Behavioral abnormalities were also seen. Compared to littermates, the TB-RBP-null mice appeared docile and exhibited reduced Rota-Rod performance.

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