ELONGATED HYPOCOTYL 5 mediates blue light-induced starch degradation in tomato

2020 ◽  
Author(s):  
Han Dong ◽  
Chaoyi Hu ◽  
Chaochao Liu ◽  
Jiachun Wang ◽  
Yanhong Zhou ◽  
...  
Keyword(s):  
Blue Light ◽  
Soluble Sugars ◽  
Light Signal ◽  
Starch Accumulation ◽  
Starch Degradation ◽  
Sugar Accumulation ◽  
Wild Type ◽  
Mobility Shift ◽  
Storage Carbohydrate ◽  
In The Wild

Abstract Starch is the major storage carbohydrate in plants, and its metabolism in chloroplasts depends mainly on light. However, the mechanism through which photoreceptors regulate starch metabolism in chloroplasts is unclear. In this study, we found that the cryptochrome 1a (CRY1a)-mediated blue light signal is critical for regulating starch accumulation by inducing starch degradation through the HY5 transcription factor in the chloroplasts in tomato. cry1a mutants and HY5-RNAi plants accumulated more starch and presented lower transcript levels of starch degradation-related genes in their leaves than did the wild-type (WT) plants. Blue light significantly induced the transcription of starch degradation-related genes in the wild-type and CRY1a- or HY5-overexpressing plants but had little effect in the cry1a and HY5-RNAi plants. Dual-luciferase assays, electrophoretic mobility shift assays (EMSA) and chromatin immunoprecipitation (ChIP)-qPCR revealed that HY5 could activate the starch degradation-related genes PWD, BAM1, BAM3, BAM8, MEX1 and DPE1 by directly binding to their promoters. Silencing of HY5 and these starch degradation-related genes in CRY1a-overexpressing plants led to increased accumulation of starch and decreased accumulation of soluble sugars. These findings presented here not only deepen our understanding of how light control starch degradation and sugar accumulation but also allow us to explore potential targets for improving crop quality.

scholarly journals Improvement of enzymatic saccharification in Arabidopsis thaliana by ectopic expression of the rice SUB1A-1 transcription factor

2015 ◽  
Author(s):  
Lizeth Núñez-López ◽  
Andrés Aguirre-Cruz ◽  
Blanca Estela Barrera-Figueroa ◽  
Julian Mario Peña-Castro
Keyword(s):  
Transcription Factor ◽  
Ectopic Expression ◽  
Enzymatic Saccharification ◽  
Biofuel Production ◽  
Starch Accumulation ◽  
Starch Degradation ◽  
Photosynthetic Parameters ◽  
Wild Type ◽  
Submergence Stress ◽  
Saccharification Yield

Saccharification of polysaccharides releases monosaccharides that can be used by ethanol-producing microorganisms in biofuel production. To improve plant biomass as a raw material for saccharification, factors controlling the accumulation and structure of carbohydrates must be identified. Rice SUB1A-1 is a transcription factor that represses the turnover of starch and postpones energy-consuming growth processes under submergence stress. Arabidopsis was employed to test if heterologous expression of SUB1A-1 or SUB1C-1 (a related gene) can be used to improve saccharification. Cellulolytic and amylolytic enzymatic treatments confirmed that SUB1A-1 transgenics had better saccharification yield than wild-type (Col-0), mainly from accumulated starch. This high saccharification yield was developmentally controlled since juvenile transgenic plants yielded 200-300% more glucose than Col-0. We measured photosynthetic parameters, starch granule microstructure, and transcript abundance of genes involved in starch degradation (SEX4, GWD1), juvenile transition (SPL3-5) and meristematic identity (FUL, SOC1) but found no differences to Col-0, indicating that starch accumulation may be controlled by down-regulation of CONSTANS and FLOWERING LOCUS T by SUB1A-1 as previously reported. SUB1A-1 transgenics also offered less resistance to deformation than wild-type concomitant to up-regulation of AtEXP2 expansin and BGL2 glucan-1,3,-beta-glucosidase. We conclude that heterologous SUB1A-1 expression can improve saccharification yield and softness, two traits needed in bioethanol production.

Blue Light Enhanced Respiratory Activity Under Photosynthetic Conditions In Chlorella; A Mass Spectrometric Analysis

1992 ◽  
Vol 47 (11-12) ◽  
pp. 881-888 ◽  
Author(s):  
Klaus P. Bader ◽  
Georg H. Schmid ◽  
Günter Ruyters ◽  
Wolfgang Kowallikb
Keyword(s):  
Oxygen Uptake ◽  
Blue Light ◽  
Red Light ◽  
Wavelength Dependence ◽  
Mass Spectrometric ◽  
Mass Spectrometric Analysis ◽  
Spectrometric Analysis ◽  
Wild Type ◽  
Carbohydrate Degradation ◽  
In The Wild

Mass spectrometric analysis shows that blue light enhances oxygen uptake during photosynthesis in Chlorella fusca. Assays in which all of the normal 16O2 of air has been substituted by 18O2 permit discrimination between photosynthetic O2-evolution (measured as 16O2, i.e. mass 32) and O2-uptake (measured as 18O2, i.e. mass 36). A chlorophyll-free Chlorella kessleri mutant for which in earlier studies the occurrence of blue light enhanced oxidative carbohydrate degradation has been demonstrated (W. Kowallik, H. Gaffron, Planta 69, 9 2 -9 5 (1966); W. Kowallik, Ann. Rev. Plant Physiol. 33, 5 7 -7 2 (1982)) has been used for comparison in the present study. The light intensity dependencies of the observed effect seem to differ in mutant and wild type cells. In the mutant a fluence rate of 1 .5 -2 .0 (μE m -2·s-1 of blue light yields saturation, whereas in the wild type even ten times this value does not. A wavelength dependence of the effect measured with equal fluence rates at 422 nm, 457 nm, 488 nm, 555 nm and 649 nm shows maximal efficiency around 460 nm and no significant effect of red light. This agrees with earlier studies on the chlorophyll- free mutant. As a result of this correspondence, we think that the enhanced oxygen uptake during photosynthesis concerns oxidative carbohydrate degradation. The putative mechanism and significance of the observed blue light enhanced respiration in photosynthesizing Chlorella are discussed

scholarly journals Serratia marcescensarn, a PhoP-Regulated Locus Necessary for Polymyxin B Resistance

2014 ◽  
Vol 58 (9) ◽  
pp. 5181-5190 ◽  
Author(s):  
Quei Yen Lin ◽  
Yi-Lin Tsai ◽  
Ming-Che Liu ◽  
Wei-Cheng Lin ◽  
Po-Ren Hsueh ◽  
...  
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Response Regulator ◽  
Challenge Test ◽  
Polymyxin B ◽  
Resistant Bacteria ◽  
Wild Type ◽  
Mobility Shift ◽  
Content Type ◽  
In The Wild

ABSTRACTPolymyxins, which are increasingly being used to treat infections caused by multidrug-resistant bacteria, perform poorly againstSerratia marcescens. To investigate the underlying mechanisms, Tn5mutagenesis was performed and two mutants exhibiting increased polymyxin B (PB) susceptibility were isolated. The mutants were found to have Tn5inserted into thearnBandarnCgenes. In other bacteria,arnBandarnCbelong to the seven-genearnoperon, which is involved in lipopolysaccharide (LPS) modification. LPSs ofarnmutants had greater PB-binding abilities than that of wild-type LPS. Further, we identified PhoP, a bacterial two-component response regulator, as a regulator of PB susceptibility inS. marcescens. By the reporter assay, we found PB- and low-Mg2+-induced expression ofphoPandarnin the wild-type strain but not in thephoPmutant. Complementation of thephoPmutant with the full-lengthphoPgene restored the PB MIC and induction by PB and low Mg2+levels, as in the wild type. An electrophoretic mobility shift assay (EMSA) further demonstrated that PhoP bound directly to thearnpromoter. The PB challenge test confirmed that pretreatment with PB and low Mg2+levels protectedS. marcescensfrom a PB challenge in the wild-type strain but not in thephoPmutant. Real-time reverse transcriptase-PCR also indicated that PB serves as a signal to regulate expression ofugd, a gene required for LPS modification, inS. marcescensthrough a PhoP-dependent pathway. Finally, we found that PB-resistant clinical isolates displayed greater expression ofarnAupon exposure to PB than did susceptible isolates. This is the first report to describe the role ofS. marcescensarnin PB resistance and its modulation by PB and Mg2+through the PhoP protein.

scholarly journals Shifting carbon flux from non-transient starch to lipid allows oil accumulation in transgenic tobacco leaves

2020 ◽  
Author(s):  
Kevin L. Chu ◽  
Lauren M. Jenkins ◽  
Sally R. Bailey ◽  
Shrikaar Kambhampati ◽  
Somnath Koley ◽  
...  
Keyword(s):  
Transgenic Tobacco ◽  
Carbon Flux ◽  
Soluble Sugars ◽  
Dry Weight ◽  
Starch Accumulation ◽  
Leaf Biomass ◽  
Wild Type ◽  
Oil Accumulation ◽  
Tobacco Leaves ◽  
Source Sink

AbstractPlant leaf biomass is composed predominantly of carbohydrate and protein with less than 5% dry weight allocated to lipid and less than 1% of total lipid in the form of triacylglycerols (TAGs). The combined overexpression of multiple genes involved in different aspects of TAG synthesis and stabilization can result in TAG accumulation to over 30% dry weight in tobacco leaves, presumably requiring many metabolic adjustments within plant cells. The metabolic consequences to the combined source and sink capacities of high oil accumulating transgenic tobacco leaves compared to wild-type were inspected across development and photoperiod by utilizing foliar biomass components and 13CO2 flux through central carbon intermediates. Lipid biosynthesis was investigated through assessment of acyl-acyl carrier protein (ACP) pools using a recently derived quantification method that was extended to accommodate isotopic labeling. Lipids accumulated stepwise over plant development in the high-oil leaves, with 13CO2-labeling studies confirming increased carbon flux to lipids. The large increase in lipid content was concurrent with a decrease in foliar starch, with limited contribution from non-sucrose soluble sugars, indicating a redirection of carbon from starch to lipids. Starch accumulated non-transiently with plant age in wild-type leaves, suggesting an inherent capacity for a developmentally-regulated carbon sink in tobacco leaves that may have enabled the programmed altered carbon partitioning to lipids in transgenics. These studies provide insight into the metabolic plasticity of dual source-sink leaves over development and may in part explain recent successful leaf lipid engineering efforts in tobacco.One sentence summaryEngineering high oil accumulation in tobacco leaves is enabled by inherent source-sink plasticity associated with non-transient foliar starch accumulation over development.

scholarly journals Improvement of enzymatic saccharification in Arabidopsis thaliana by ectopic expression of the rice SUB1A-1 transcription factor

2015 ◽  
Author(s):  
Lizeth Núñez-López ◽  
Andrés Aguirre-Cruz ◽  
Blanca Estela Barrera-Figueroa ◽  
Julian Mario Peña-Castro
Keyword(s):  
Transcription Factor ◽  
Ectopic Expression ◽  
Enzymatic Saccharification ◽  
Biofuel Production ◽  
Starch Accumulation ◽  
Starch Degradation ◽  
Photosynthetic Parameters ◽  
Wild Type ◽  
Submergence Stress ◽  
Saccharification Yield

Saccharification of polysaccharides releases monosaccharides that can be used by ethanol-producing microorganisms in biofuel production. To improve plant biomass as a raw material for saccharification, factors controlling the accumulation and structure of carbohydrates must be identified. Rice SUB1A-1 is a transcription factor that represses the turnover of starch and postpones energy-consuming growth processes under submergence stress. Arabidopsis was employed to test if heterologous expression of SUB1A-1 or SUB1C-1 (a related gene) can be used to improve saccharification. Cellulolytic and amylolytic enzymatic treatments confirmed that SUB1A-1 transgenics had better saccharification yield than wild-type (Col-0), mainly from accumulated starch. This high saccharification yield was developmentally controlled since juvenile transgenic plants yielded 200-300% more glucose than Col-0. We measured photosynthetic parameters, starch granule microstructure, and transcript abundance of genes involved in starch degradation (SEX4, GWD1), juvenile transition (SPL3-5) and meristematic identity (FUL, SOC1) but found no differences to Col-0, indicating that starch accumulation may be controlled by down-regulation of CONSTANS and FLOWERING LOCUS T by SUB1A-1 as previously reported. SUB1A-1 transgenics also offered less resistance to deformation than wild-type concomitant to up-regulation of AtEXP2 expansin and BGL2 glucan-1,3,-beta-glucosidase. We conclude that heterologous SUB1A-1 expression can improve saccharification yield and softness, two traits needed in bioethanol production.

scholarly journals DevA, a GntR-Like Transcriptional Regulator Required for Development in Streptomyces coelicolor

2006 ◽  
Vol 188 (14) ◽  
pp. 5014-5023 ◽  
Author(s):  
Paul A. Hoskisson ◽  
Sebastien Rigali ◽  
Kay Fowler ◽  
Kim C. Findlay ◽  
Mark J. Buttner
Keyword(s):  
Fluorescent Protein ◽  
Streptomyces Coelicolor ◽  
Aerial Mycelium ◽  
Developmental Cycle ◽  
Wild Type ◽  
Mobility Shift ◽  
S1 Nuclease ◽  
Aerial Hyphae ◽  
In The Wild

ABSTRACT The gram-positive filamentous bacterium Streptomyces coelicolor has a complex developmental cycle with three distinct phases: growth of the substrate mycelium, development of reproductive structures called aerial hyphae, and differentiation of these aerial filaments into long chains of exospores. During a transposon mutagenesis screen, we identified a novel gene (devA) required for proper development. The devA mutant produced only rare aerial hyphae, and those that were produced developed aberrant spore chains that were much shorter than wild-type chains and had misplaced septa. devA encodes a member of the GntR superfamily, a class of transcriptional regulators that typically respond to metabolite effector molecules. devA forms an operon with the downstream gene devB, which encodes a putative hydrolase that is also required for aerial mycelium formation on R5 medium. S1 nuclease protection analysis showed that transcription from the single devA promoter was temporally associated with vegetative growth, and enhanced green fluorescent protein transcriptional fusions showed that transcription was spatially confined to the substrate hyphae in the wild type. In contrast, devAB transcript levels were dramatically upregulated in a devA mutant and the devA promoter was also active in aerial hyphae and spores in this background, suggesting that DevA might negatively regulate its own production. This suggestion was confirmed by gel mobility shift assays that showed that DevA binds its own promoter region in vitro.

scholarly journals The T-DNA Oncogene A4-orf8 from Agrobacterium rhizogenes A4 Induces Abnormal Growth in Tobacco

2005 ◽  
Vol 18 (3) ◽  
pp. 205-211 ◽  
Author(s):  
Marie Umber ◽  
Bernadette Clément ◽  
Léon Otten
Keyword(s):  
Agrobacterium Rhizogenes ◽  
Soluble Sugars ◽  
Starch Accumulation ◽  
Wild Type ◽  
Abnormal Growth ◽  
Regenerated Plants ◽  
Physical Linkage ◽  
Dark Green ◽  
Characteristic Growth ◽  
Fad Binding

The related orf8 and iaaM T-DNA genes from Agrobacterium are each composed of two distinct parts. The 5′ parts (called Norf8 or NiaaM) encode a 200-amino-acid (aa) sequence with homology to various T-DNA oncoproteins such as RolB, RolC, and 6b. The 3′ parts (Corf8 or CiaaM) encode a 550-aa sequence with homology to IaaM proteins from Pseudomonas and Pantoea spp. Whereas iaaM genes encode flavin adenine dinucleotide (FAD)-dependent tryptophan 2-monooxygenases that catalyze the synthesis of indole-3-acetamide (IAM), A4-orf8 from Agrobacterium rhizogenes A4 does not. Plants expressing a 2x35S-A4-Norf8 construct accumulate soluble sugars and starch. We now have regenerated plants that express the full-size 2x35S-A4-orf8 and the truncated 2x35S-A4-Corf8 gene. 2x35S-A4-Corf8 plants accumulate starch and show reduced growth like 2x35SA4-Norf8 plants but, in addition, display a novel set of characteristic growth modifications. These consist of leaf hypertrophy and hyperplasia (blisters); thick, dark-green leaves; thick stems; and swollen midveins. Mutations in the putative FAD-binding site of A4-Orf8 did not affect the blister syndrome. Plants expressing 2x35S-A4-Corf8 had a normal phenotype but contained less starch and soluble sugars than did wild-type plants. When 2x35S-A4-Corf8 plants were crossed to starch-accumulating 2x35S-A4-Norf8 plants with reduced growth, A4-Corf8 partially restored growth and reduced starch accumulation. A4-Corf8xA4-Norf8 crosses did not lead to the blister syndrome, suggesting that this requires physical linkage of the A4-NOrf8 and A4-COrf8 sequences.

scholarly journals Salicylic Acid-Mediated Diacylglycerol/Triacylglycerol Conversion Affects the Freezing Tolerance of Arabidopsis

2021 ◽  
Author(s):  
Le Xu ◽  
Jun Wu ◽  
Yancui Zhao ◽  
Huaqiong Liu ◽  
Wenying Zhang ◽  
...  
Keyword(s):  
Salicylic Acid ◽  
Freezing Tolerance ◽  
Diacylglycerol Acyltransferase ◽  
Freezing Stress ◽  
Sugar Accumulation ◽  
Wild Type ◽  
Lipid Profiling ◽  
Opposite Trend ◽  
In The Wild ◽  
Higher Sensitivity

Abstract Diacylglycerol (DAG) is likely converted to triacylglycerol (TAG) by the enzyme diacylglycerol acyltransferase (DGAT), and this conversion is important in freezing tolerance of Arabidopsis. The phytohormone salicylic acid (SA) plays important roles in the chilling and freezing tolerance of plants. In our study, we analysed the chilling phenotype, proline and sugar accumulation, phytohormone measurement, and lipid profiling of dgat1 mutants during chilling or freezing stress. We found that dgat1-1 mutants exhibited higher sensitivity to long exposure to cold stress and showed lower proline and sugar accumulation under cold acclimation conditions. The freezing-sensitive phenotype of dgat1 mutants can be ameliorated by mutations of key salicylic acid (SA) signalling components SAG101, EDS1, and PAD4 through phenotyping analysis of double mutants. Dgat1 mutants accumulated more SA, ABA, JA-Ile (jasmonate isoleucine) and OPDA (12- oxyphytodienoic acid) during freezing stress and after recovery. In addition, the DAG/TAG content in the SA-deficient mutant sid2 was lower than that in the wild type, while the SA-excessive accumulated mutant siz1 showed the opposite trend. In summary, SA could mediate the freezing tolerance of Arabidopsis by regulating the ratio of DAG and TAG, which influences the integrity of the membrane.

scholarly journals Transcriptional Control of the Iron-ResponsivefxbA Gene by the Mycobacterial Regulator IdeR

1999 ◽  
Vol 181 (11) ◽  
pp. 3402-3408 ◽  
Author(s):  
Olivier Dussurget ◽  
Juliano Timm ◽  
Manuel Gomez ◽  
Benjamin Gold ◽  
Shengwei Yu ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Regulatory Region ◽  
Wild Type ◽  
Mobility Shift ◽  
Positive Role ◽  
Dnase I Footprinting ◽  
In The Wild ◽  
Essential Enzyme ◽  
Gel Mobility Shift

ABSTRACT Exochelin is the primary extracellular siderophore ofMycobacterium smegmatis, and the iron-regulatedfxbA gene encodes a putative formyltransferase, an essential enzyme in the exochelin biosynthetic pathway (E. H. Fiss, Y. Yu, and W. R. Jacobs, Jr., Mol. Microbiol. 14:557–569, 1994). We investigated the regulation of fxbA by the mycobacterial IdeR, a homolog of the Corynebacterium diphtheriae iron regulator DtxR (M. P. Schmitt, M. Predich, L. Doukhan, I. Smith, and R. K. Holmes, Infect. Immun. 63:4284–4289, 1995). Gel mobility shift experiments showed that IdeR binds to the fxbA regulatory region in the presence of divalent metals. DNase I footprinting assays indicated that IdeR binding protects a 28-bp region containing a palindromic sequence of the fxbA promoter that was identified in primer extension assays. fxbA regulation was measured in M. smegmatis wild-type and ideR mutant strains containing fxbA promoter-lacZ fusions. These experiments confirmed that fxbA expression is negatively regulated by iron and showed that inactivation of ideRresults in iron-independent expression of fxbA. However, the levels of its expression in the ideR mutant were approximately 50% lower than those in the wild-type strain under iron limitation, indicating an undefined positive role of IdeR in the regulation of fxbA.

scholarly journals Mua (HP0868) Is a Nickel-Binding Protein That Modulates Urease Activity in Helicobacter pylori

mBio ◽  
2011 ◽  
Vol 2 (2) ◽  
Author(s):  
Stéphane L. Benoit ◽  
Robert J. Maier
Keyword(s):  
Helicobacter Pylori ◽  
Urease Activity ◽  
Binding Protein ◽  
Total Protein Content ◽  
Nickel Concentration ◽  
Wild Type ◽  
Mobility Shift ◽  
Content Type ◽  
Significant Difference ◽  
In The Wild

ABSTRACTA novel mechanism aimed at controlling urease expression inHelicobacter pyloriin the presence of ample nickel is described. Higher urease activities were observed in anhp0868mutant (than in the wild type) in cells supplemented with nickel, suggesting that the HP0868 protein (herein named Mua formodulator ofureaseactivity) represses urease activity when nickel concentrations are ample. The increase in urease activity in the Δmuamutant was linked to an increase in urease transcription and synthesis, as shown by quantitative real-time PCR, SDS-PAGE, and immunoblotting against UreAB. Increased urease synthesis was also detected in a ΔmuaΔnikRdouble mutant strain. The Δmuamutant was more sensitive to nickel toxicity but more resistant to acid challenge than was the wild-type strain. Pure Mua protein binds 2 moles of Ni2+per mole of dimer. Electrophoretic mobility shift assays did not reveal any binding of Mua to theureApromoter or other selected promoters (nikR,arsRS,5′ ureB-sRNAp). Previous yeast two-hybrid studies indicated that Mua and RpoD may interact; however, only a weak interaction was detected via cross-linking with pure components and this could not be verified by another approach. There was no significant difference in the intracellular nickel level between wild-type andmuamutant cells. Taken together, our results suggest the HP0868 gene product represses urease transcription when nickel levels are high through an as-yet-uncharacterized mechanism, thus counterbalancing the well-described NikR-mediated activation.IMPORTANCEUrease is a nickel-containing enzyme that buffers both the cytoplasm and the periplasm ofHelicobacter pyloriby converting urea into ammonia and carbon dioxide. The enzyme is the most abundant protein inH. pylori, accounting for an estimated 10% of the total protein content of the cell, and it is essential for early colonization and virulence. Numerous studies have focused on the transcription of the structuralureABgenes and its control by the regulatory proteins NikR and ArsR. Here we propose that urease transcription is under the control of another Ni-binding protein besides NikR, the Mua (HP0868) protein. Our results suggest that the Mua protein represses urease transcription when nickel levels are high. This mechanism would counterbalance the NikR-mediated activation of urease and ensure that, in the presence of a high nickel concentration, urease activation is limited and does not lead to massive production of detrimental ammonia.

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