scholarly journals Characterization and Analysis of Anthocyanin-Related Genes in Wild-Type Blueberry and the Pink-Fruited Mutant Cultivar ‘Pink Lemonade’: New Insights into Anthocyanin Biosynthesis

Agronomy ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1296
Author(s):  
Jose V. Die ◽  
Richard W. Jones ◽  
Elizabeth L. Ogden ◽  
Mark K. Ehlenfeldt ◽  
Lisa J. Rowland
Keyword(s):  
Fruits And Vegetables ◽  
Anthocyanin Biosynthesis ◽  
Mechanistic Explanation ◽  
Comparative Sequence Analysis ◽  
Wild Type ◽  
Flavonoid Pathway ◽  
Transient Expression Assay ◽  
Gene Encoding ◽  
In The Wild ◽  
Anthocyanin Production

Blueberries are one of the richest sources of antioxidants, such as anthocyanins, among fruits and vegetables. Anthocyanin mutants, like the pink-fruited cultivar ‘Pink Lemonade’, are valuable resources for investigating anthocyanin biosynthesis in blueberries. In this study, we examined expression of flavonoid pathway genes during fruit development in wild-type, blue-fruited blueberries using quantitative real-time PCR. Expression was also compared between wild-type and the pink-fruited ‘Pink Lemonade’. This revealed significantly lower expression in ‘Pink Lemonade’ than in wild-type of nearly all the structural genes examined suggesting that a transcriptional regulator of the pathway was affected. Hence, we compared expression of three known regulatory genes and found that the gene encoding the transcription factor MYB1 was expressed at a significantly lower level in ‘Pink Lemonade’ than in the wild-type. To validate the capacity of this MYB1 to regulate the transcription of anthocyanin genes in blueberries, a transient expression assay was conducted. Results indicated MYB1 overexpression enhanced anthocyanin production. Comparative sequence analysis between wild-type and mutant MYB1 variants found differences in highly conserved features suggesting a mechanistic explanation for the mutant phenotype. Collectively, the results presented here contribute to a better understanding of mechanisms regulating anthocyanin biosynthesis in Vaccinium.

scholarly journals Expression and function of the cdgD gene, encoding a CHASE–PAS-DGC-EAL domain protein, in Azospirillum brasilense

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
José Francisco Cruz-Pérez ◽  
Roxana Lara-Oueilhe ◽  
Cynthia Marcos-Jiménez ◽  
Ricardo Cuatlayotl-Olarte ◽  
María Luisa Xiqui-Vázquez ◽  
...  
Keyword(s):  
Azospirillum Brasilense ◽  
Type Strain ◽  
Wild Type Strain ◽  
Soil Conditions ◽  
Wild Type ◽  
Gene Encoding ◽  
Wheat Roots ◽  
Genes Encoding ◽  
In The Wild ◽  
Plant Growth Promoting

AbstractThe plant growth-promoting bacterium Azospirillum brasilense contains several genes encoding proteins involved in the biosynthesis and degradation of the second messenger cyclic-di-GMP, which may control key bacterial functions, such as biofilm formation and motility. Here, we analysed the function and expression of the cdgD gene, encoding a multidomain protein that includes GGDEF-EAL domains and CHASE and PAS domains. An insertional cdgD gene mutant was constructed, and analysis of biofilm and extracellular polymeric substance production, as well as the motility phenotype indicated that cdgD encoded a functional diguanylate protein. These results were correlated with a reduced overall cellular concentration of cyclic-di-GMP in the mutant over 48 h compared with that observed in the wild-type strain, which was recovered in the complemented strain. In addition, cdgD gene expression was measured in cells growing under planktonic or biofilm conditions, and differential expression was observed when KNO3 or NH4Cl was added to the minimal medium as a nitrogen source. The transcriptional fusion of the cdgD promoter with the gene encoding the autofluorescent mCherry protein indicated that the cdgD gene was expressed both under abiotic conditions and in association with wheat roots. Reduced colonization of wheat roots was observed for the mutant compared with the wild-type strain grown in the same soil conditions. The Azospirillum-plant association begins with the motility of the bacterium towards the plant rhizosphere followed by the adsorption and adherence of these bacteria to plant roots. Therefore, it is important to study the genes that contribute to this initial interaction of the bacterium with its host plant.

scholarly journals Chlorobium tepidum Mutant Lacking Bacteriochlorophyll c Made by Inactivation of the bchK Gene, Encoding Bacteriochlorophyll c Synthase

2002 ◽  
Vol 184 (12) ◽  
pp. 3368-3376 ◽  
Author(s):  
Niels-Ulrik Frigaard ◽  
Ginny D. Voigt ◽  
Donald A. Bryant
Keyword(s):  
Photosynthetic Reaction Center ◽  
Chlorobium Tepidum ◽  
Wild Type ◽  
Bacteriochlorophyll C ◽  
Gene Encoding ◽  
Thin Sections ◽  
Insertional Inactivation ◽  
In The Wild ◽  
Green Sulfur ◽  
Isoprenoid Quinones

ABSTRACT The gene encoding bacteriochlorophyll (BChl) c synthase was identified by insertional inactivation in the photosynthetic green sulfur bacterium Chlorobium tepidum and was named bchK. The bchK mutant of C. tepidum was rusty-orange in color and completely lacked BChl c. Because of the absence of the BChl c antenna, the mutant grew about seven times slower than the wild type at light intensities that were limiting to the wild type (<90 μmol m−2 s−1). Various pheophorbides, which probably represent precursors of BChl c which had lost magnesium, accumulated in the mutant cells. A small fraction of these pheophorbides were apparently esterified by the remaining chlorophyll (Chl) a and BChl a synthases in cells. The amounts of BChl a, Chl a, isoprenoid quinones, carotenoids, Fenna-Matthews-Olson protein, and chlorosome envelope protein CsmA were not significantly altered on a cellular basis in the mutant compared to in the wild type. This suggests that the BChl a antennae, photosynthetic reaction centers, and remaining chlorosome components were essentially unaffected in the mutant. Electron microscopy of thin sections revealed that the mutant lacked normal chlorosomes. However, a fraction containing vestigial chlorosomes, denoted “carotenosomes,” was partly purified by density centrifugation; these structures contained carotenoids, isoprenoid quinones, and a 798-nm-absorbing BChl a species that is probably protein associated. Because of the absence of the strong BChl c absorption found in the wild type, the bchK mutant should prove valuable for future analyses of the photosynthetic reaction center and of the roles of BChl a in photosynthesis in green bacteria. An evolutionary implication of our findings is that the photosynthetic ancestor of green sulfur bacteria could have evolved without chlorosomes and BChl c and instead used only BChl a-containing proteins as the major light-harvesting antennae.

scholarly journals Overproduction of l-Lysine from Methanol by Methylobacillus glycogenes Derivatives Carrying a Plasmid with a Mutated dapA Gene

2001 ◽  
Vol 67 (7) ◽  
pp. 3064-3070 ◽  
Author(s):  
Hiroaki Motoyama ◽  
Hiroshi Yano ◽  
Yoko Terasaki ◽  
Hideharu Anazawa
Keyword(s):  
Amino Acid ◽  
Specific Activity ◽  
Test Tube ◽  
Amino Acid Sequences ◽  
Nutritional Requirement ◽  
Wild Type ◽  
Gene Encoding ◽  
Lysine Production ◽  
Obligate Methylotroph ◽  
In The Wild

ABSTRACT The dapA gene, encoding dihydrodipicolinate synthase (DDPS) partially desensitized to inhibition by l-lysine, was cloned from an l-threonine- andl-lysine-coproducing mutant of the obligate methylotrophMethylobacillus glycogenes DHL122 by complementation of the nutritional requirement of an Escherichia coli dapAmutant. Introduction of the dapA gene into DHL122 and AL119, which is the parent of DHL122 and an l-threonine producing mutant, elevated the specific activity of DDPS 20-fold andl-lysine production 2- to 3-fold with concomitant reduction of l-threonine in test tube cultures. AL119 containing thedapA gene produced 8 g of l-lysine per liter in a 5-liter jar fermentor from methanol as a substrate. Analysis of the nucleotide sequence of the dapA gene shows that it encodes a peptide with an M r of 30,664 and that the encoded amino acid sequence is extensively homologous to those of other organisms. In order to study the mutation that occurred in DHL122, the dapA genes of the wild type and AL119 were cloned and sequenced. Comparison of the nucleotide sequences of the dapA genes revealed that the amino acid at residue 88 was F in DHL122 whereas it was L in the wild type and AL119, suggesting that this amino acid alteration that occurred in DHL122 caused the partial desensitization of DDPS to the inhibition byl-lysine. The similarity in the amino acid sequences of DDPS in M. glycogenes and other organisms suggests that the mutation of the dapA gene in DHL122 is located in the region concerned with interaction of the allosteric effector,l-lysine.

scholarly journals A Combination of Three Mutations, dcd,pyrH, and cdd, Establishes Thymidine (Deoxyuridine) Auxotrophy in thyA+ Strains of Salmonella typhimurium

1998 ◽  
Vol 180 (22) ◽  
pp. 5891-5895 ◽  
Author(s):  
Nevan J. Krogan ◽  
Michelle L. Zaharik ◽  
Jan Neuhard ◽  
Rod A. Kelln
Keyword(s):  
Salmonella Typhimurium ◽  
Genetic Background ◽  
Kinase Activity ◽  
Null Mutation ◽  
Wild Type ◽  
Gene Encoding ◽  
Deoxycytidine Deaminase ◽  
In The Wild ◽  
Ump Kinase ◽  
Thymidine Auxotrophy

ABSTRACT The dum gene of Salmonella typhimurium was originally identified as a gene involved in dUMP synthesis (C. F. Beck et al., J. Bacteriol. 129:305–316, 1977). In the genetic background used in their selection, the joint acquisition of adcd (dCTP deaminase) and a dum mutation established a condition of thymidine (deoxyuridine) auxotrophy. In this study, we show that dum is identical to pyrH, the gene encoding UMP kinase. The level of UMP kinase activity in thedum mutant was found to be only 30% of that observed for the dum + strain. Thymidine prototrophy was restored to the original dum dcd mutant (KP1361) either by transduction using a pyrH + donor or by complementation with either of twopyrH +-carrying plasmids. Thymidine auxotrophy could be reconstructed in the dum + derivative (KP1389) by the introduction of a mutant pyrH allele. To define the minimal mutational complement necessary to produce thymidine auxotrophy in thyA + strains, adcd::Km null mutation was constructed. In the wild-type background, dcd::Km alone or in combination with a pyrH (dum) mutation did not result in a thymidine requirement. A third mutation, cdd(cytidine-deoxycytidine deaminase), was required together with thedcd and pyrH mutations to impart thymidine auxotrophy.

scholarly journals Pleiotropic Effects of Inactivating a Carboxyl-Terminal Protease, CtpA, in Borrelia burgdorferi

2004 ◽  
Vol 186 (7) ◽  
pp. 2074-2084 ◽  
Author(s):  
Yngve Östberg ◽  
James A. Carroll ◽  
Marija Pinne ◽  
Jonathan G. Krum ◽  
Patricia Rosa ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Higher Plants ◽  
Bacterial Species ◽  
Immunoblot Analysis ◽  
Pleiotropic Effect ◽  
Wild Type ◽  
Gene Encoding ◽  
Two Dimensional Gel Electrophoresis ◽  
In The Wild ◽  
Carboxyl Terminal

ABSTRACT A gene encoding a putative carboxyl-terminal protease (CtpA), an unusual type of protease, is present in the Borrelia burgdorferi B31 genome. The B. burgdorferi CtpA amino acid sequence exhibits similarities to the sequences of the CtpA enzymes of the cyanobacterium Synechocystis sp. strain PCC 6803 and higher plants and also exhibits similarities to the sequences of putative CtpA proteins in other bacterial species. Here, we studied the effect of ctpA gene inactivation on the B. burgdorferi protein expression profile. Total B. burgdorferi proteins were separated by two-dimensional gel electrophoresis, and the results revealed that six proteins of the wild type were not detected in the ctpA mutant and that nine proteins observed in the ctpA mutant were undetectable in the wild type. Immunoblot analysis showed that the integral outer membrane protein P13 was larger and had a more acidic pI in the ctpA mutant, which is consistent with the theoretical change in pI for P13 not processed at the carboxyl terminus. Matrix-assisted laser desorption ionization—time of flight data indicated that in addition to P13, the BB0323 protein may serve as a substrate for carboxyl-terminal processing by CtpA. Complementation analysis of the ctpA mutant provided strong evidence that the observed effect on proteins depended on inactivation of the ctpA gene alone. We show that CtpA in B. burgdorferi is involved in the processing of proteins such as P13 and BB0323 and that inactivation of ctpA has a pleiotropic effect on borrelial protein synthesis. To our knowledge, this is the first analysis of both a CtpA protease and different substrate proteins in a pathogenic bacterium.

scholarly journals Sucrose Enhances Anthocyanin Accumulation in Torenia by Promoting Expression of Anthocyanin Biosynthesis Genes

Horticulturae ◽  
2021 ◽  
Vol 7 (8) ◽  
pp. 219
Author(s):  
Aung Htay Naing ◽  
Junping Xu ◽  
Kyeung Il Park ◽  
Mi Young Chung ◽  
Chang Kil Kim
Keyword(s):  
Transcription Factors ◽  
Transgenic Plants ◽  
Plant Growth ◽  
Anthocyanin Biosynthesis ◽  
Anthocyanin Accumulation ◽  
Wild Type ◽  
Anthocyanin Production

We examined the effects of different sucrose concentrations (3%, 5%, and 7%) on anthocyanin accumulation and plant growth in wild type (WT) and transgenic (T2) torenia cultivar “Kauai Rose” overexpressing the anthocyanin regulatory transcription factors B-Peru + mPAP1 or RsMYB1. Sucrose increased anthocyanin production in both WT and transgenic plants, with higher anthocyanin production in transgenic plants compared to WT plants. Higher sucrose concentrations increased production of anthocyanin in transgenic and WT plants, with increased anthocyanin production associated with increased expression of anthocyanin biosynthesis genes. Higher sucrose concentrations reduced growth of WT and transgenic plants. Our results indicate that sucrose enhances anthocyanin production in torenia by regulating anthocyanin biosynthesis genes.

scholarly journals Galactose Metabolism by Streptococcus mutans

2004 ◽  
Vol 70 (10) ◽  
pp. 6047-6052 ◽  
Author(s):  
Jacqueline Abranches ◽  
Yi-Ywan M. Chen ◽  
Robert A. Burne
Keyword(s):  
Streptococcus Mutans ◽  
Wild Type Strain ◽  
Sugar Metabolism ◽  
Growth Defect ◽  
Wild Type ◽  
Phosphotransferase System ◽  
Galactose Metabolism ◽  
Gene Encoding ◽  
Leloir Pathway ◽  
In The Wild

ABSTRACT The galK gene, encoding galactokinase of the Leloir pathway, was insertionally inactivated in Streptococcus mutans UA159. The galK knockout strain displayed only marginal growth on galactose, but growth on glucose or lactose was not affected. In strain UA159, the sugar phosphotransferase system (PTS) for lactose and the PTS for galactose were induced by growth in lactose and galactose, although galactose PTS activity was very low, suggesting that S. mutans does not have a galactose-specific PTS and that the lactose PTS may transport galactose, albeit poorly. To determine if the galactose growth defect of the galK mutant could be overcome by enhancing lactose PTS activity, the gene encoding a putative repressor of the operon for lactose PTS and phospho-β-galactosidase, lacR, was insertionally inactivated. A galK and lacR mutant still could not grow on galactose, although the strain had constitutively elevated lactose PTS activity. The glucose PTS activity of lacR mutants grown in glucose was lower than in the wild-type strain, revealing an influence of LacR or the lactose PTS on the regulation of the glucose PTS. Mutation of the lacA gene of the tagatose pathway caused impaired growth in lactose and galactose, suggesting that galactose can only be efficiently utilized when both the Leloir and tagatose pathways are functional. A mutation of the permease in the multiple sugar metabolism operon did not affect growth on galactose. Thus, the galactose permease of S. mutans is not present in the gal, lac, or msm operons.

Haem-linked interactions in horseradish peroxidase revealed by spectroscopic analysis of the Phe-221→Met mutant

2001 ◽  
Vol 353 (2) ◽  
pp. 181-191 ◽  
Author(s):  
Barry D. HOWES ◽  
Nigel C. VEITCH ◽  
Andrew T. SMITH ◽  
Christopher G. WHITE ◽  
Giulietta SMULEVICH
Keyword(s):  
Horseradish Peroxidase ◽  
Enzyme Stability ◽  
Solvent Accessibility ◽  
Imidazole Ring ◽  
Axial Ligand ◽  
Wild Type ◽  
Gene Encoding ◽  
Alkaline Transition ◽  
In The Wild ◽  
Benzhydroxamic Acid

A gene encoding a Phe-221-to-Met substitution in the haem enzyme horseradish peroxidase has been constructed and expressed in Escherichia coli. In the wild-type enzyme the side chain of Phe-221 is tightly stacked against the imidazole ring of His-170, which provides the only axial ligand to the haem iron atom. The Phe-221 → Met enzyme is active, and forms characteristic complexes with typical peroxidase ligands (CO, cyanide, fluoride), and with benzhydroxamic acid. Significant differences between the mutant and wild-type enzymes can be detected spectroscopically. These include a change in the Fe(III) resting state of the enzyme to an unusual quantum mechanically mixed-spin haem species, a marked decrease in the pKa of the alkaline transition and a reduction in enzyme stability at alkaline pH for both Fe(III) and Fe(II) forms. The perturbation of the haem pocket in the mutant can be attributed to several factors, including the increased steric freedom and solvent accessibility of the His-170 ligand, as indicated by 1H-NMR data, and the loss of the πŐπ interaction between His-170 and Phe-221.

scholarly journals Increased Production of Zeaxanthin and Other Pigments by Application of Genetic Engineering Techniques toSynechocystis sp. Strain PCC 6803

2000 ◽  
Vol 66 (1) ◽  
pp. 64-72 ◽  
Author(s):  
Delphine Lagarde ◽  
Laurent Beuf ◽  
Wim Vermaas
Keyword(s):  
Mutant Strain ◽  
Fold Increase ◽  
Carotenoid Biosynthesis ◽  
Carotenoid Content ◽  
Wild Type ◽  
Gene Encoding ◽  
Genes Encoding ◽  
In The Wild ◽  
Β Carotene ◽  
Pcc 6803

ABSTRACT The psbAII locus was used as an integration platform to overexpress genes involved in carotenoid biosynthesis inSynechocystis sp. strain PCC 6803 under the control of the strong psbAII promoter. The sequences of the genes encoding the yeast isopentenyl diphosphate isomerase (ipi) and theSynechocystis β-carotene hydroxylase (crtR) and the linked Synechocystis genes coding for phytoene desaturase and phytoene synthase (crtP andcrtB, respectively) were introduced intoSynechocystis, replacing the psbAII coding sequence. Expression of ipi, crtR, andcrtP and crtB led to a large increase in the corresponding transcript levels in the mutant strains, showing that the psbAII promoter can be used to drive transcription and to overexpress various genes in Synechocystis. Overexpression of crtP and crtB led to a 50% increase in the myxoxanthophyll and zeaxanthin contents in the mutant strain, whereas the β-carotene and echinenone contents remained unchanged. Overexpression of crtR induced a 2.5-fold increase in zeaxanthin accumulation in the corresponding overexpressing mutant compared to that in the wild-type strain. In this mutant strain, zeaxanthin becomes the major pigment (more than half the total amount of carotenoid) and the β-carotene and echinenone amounts are reduced by a factor of 2. However, overexpression of ipi did not result in a change in the carotenoid content of the mutant. To further alter the carotenoid content of Synechocystis, the crtOgene, encoding β-carotene ketolase, which converts β-carotene to echinenone, was disrupted in the wild type and in the overexpressing strains so that they no longer produced echinenone. In this way, by a combination of overexpression and deletion of particular genes, the carotenoid content of cyanobacteria can be altered significantly.

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