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1.
New Phytol ; 237(4): 1215-1228, 2023 02.
Article in English | MEDLINE | ID: mdl-36377104

ABSTRACT

Phosphate (Pi) availability is a major factor limiting plant growth and development. The key transcription factor controlling Pi-starvation response (PSR) is PHOSPHATE STARVATION RESPONSE 1 (PHR1) whose transcript levels do not change with changes in Pi levels. However, how PHR1 stability is regulated at the post-translational level is relatively unexplored in Arabidopsis thaliana. Inositol polyphosphates (InsPn) are important signal molecules that promote the association of stand-alone SPX domain proteins with PHR1 to regulate PSR. Here, we show that NITROGEN LIMITATION ADAPTATION (NLA) E3 ligase can associate with PHR1 through its conserved SPX domain and polyubiquitinate PHR1 in vitro. The association with PHR1 and its ubiquitination is enhanced by InsP6 but not by InsP5. Analysis of InsPn-related mutants and an overexpression plant shows PHR1 levels are more stable in itpk4-1 and vih2-4/VIH1amiRNA but less stable in ITPK4 overexpression plants. Under Pi-deficient conditions, nla seedlings contain high PHR1 levels, display long root hair and accumulate anthocyanin in shoots phenocopying PHR1 overexpression plants. By contrast, NLA overexpression plants phenocopy phr1 whose phenotypes are opposite to those of nla. Our results suggest NLA functions as a negative regulator of Pi response by modulating PHR1 stability and the NLA/PHR1 association depends on InsPn levels.


Subject(s)
Arabidopsis Proteins , Arabidopsis , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Gene Expression Regulation, Plant , Phosphates/metabolism , Polyphosphates/metabolism , Transcription Factors/metabolism , Ubiquitin-Protein Ligases/genetics , Ubiquitin-Protein Ligases/metabolism , Ubiquitination
2.
Plant Commun ; 3(5): 100348, 2022 09 12.
Article in English | MEDLINE | ID: mdl-35706355

ABSTRACT

As a key transcription factor in the brassinosteroid (BR) signaling pathway, the activity and expression of BES1 (BRI1-EMS-SUPPRESSOR 1) are stringently regulated. BES1 degradation is mediated by ubiquitin-related 26S proteasomal and autophagy pathways, which attenuate and terminate BR signaling; however, the opposing deubiquitinases (DUBs) are still unknown. Here, we showed that the ubp12-2w/13-3 double mutant phenocopies the BR-deficient dwarf mutant, suggesting that the two DUBs UBP12/UBP13 antagonize ubiquitin-mediated degradation to stabilize BES1. These two DUBs can trim tetraubiquitin with K46 and K63 linkages in vitro. UBP12/BES1 and UBP13/BES1 complexes are localized in both cytosol and nuclei. UBP12/13 can deubiquitinate polyubiquitinated BES1 in vitro and in planta, and UBP12 interacts with and deubiquitinates both inactive, phosphorylated BES1 and active, dephosphorylated BES1 in vivo. UBP12 overexpression in BES1OE plants significantly enhances cell elongation in hypocotyls and petioles and increases the ratio of leaf length to width compared with BES1OE or UBP12OE plants. Hypocotyl elongation and etiolation result from elevated BES1 levels because BES1 degradation is retarded by UBP12 in darkness or in light with BR. Protein degradation inhibitor experiments show that the majority of BES1 can be degraded by either the proteasomal or the autophagy pathway, but a minor BES1 fraction remains pathway specific. In conclusion, UBP12/UBP13 deubiquitinate BES1 to stabilize the latter as a positive regulator for BR responses.


Subject(s)
Arabidopsis Proteins , Arabidopsis , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Brassinosteroids/metabolism , DNA-Binding Proteins/metabolism , Endopeptidases/genetics , Endopeptidases/metabolism , Gene Expression Regulation, Plant , Signal Transduction , Ubiquitins/metabolism
3.
New Phytol ; 223(3): 1447-1460, 2019 08.
Article in English | MEDLINE | ID: mdl-31050353

ABSTRACT

Nitrogen deficiency (-N) in plants triggers leaf senescence which is regulated by the transcription factor ORE1. Little is known about post-translational regulation of ORE1 in this process. Here, we show that UBP12/UBP13 (ubiquitin-specific protease 12/13) antagonize the action of NLA (nitrogen limitation adaptation) E3 ligase to maintain ORE1 homeostasis. In vitro pull-down and in vivo co-immunoprecipitation assays demonstrated specific binding between UBP12/UBP13 and ORE1. We further analyzed in various genotypes total Chl content and expression levels of senescence-related genes under -N conditions. We found that UBP12/UBP13 can deubiquitinate polyubiquitinated ORE1 in vitro and increase the stability of ORE1 in vivo in MG132/cycloheximide-chase experiments. Plants overexpressing UBP12/UBP13 display accelerated leaf senescence which is reversed by the ore1 mutation. By contrast, the senescence phenotype of plants overexpressing ORE1 is exacerbated by UBP12/UBP13 overexpression. The expression of senescence-related genes tracks the senescence phenotype. ORE1 protein levels can be elevated by UBP12/UBP13 overexpression but decreased in ubp12-2w/13-3. In conclusion, UBP12/UBP13 deubiquitinate ORE1 to stabilize this transcription factor and promote its activity as a positive regulator for leaf senescence under -N conditions. Our study shows that UBP12/UBP13 counteracts the effect of NLA E3 ligase to accelerate leaf senescence under nitrogen starvation.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Endopeptidases/metabolism , Nitrogen/deficiency , Plant Leaves/enzymology , Plant Leaves/growth & development , Transcription Factors/metabolism , Arabidopsis/genetics , Genotype , Models, Biological , Mutation/genetics , Phenotype , Polyubiquitin/metabolism , Protein Binding , Protein Stability , Ubiquitination
4.
BMC Genomics ; 19(1): 532, 2018 Jul 13.
Article in English | MEDLINE | ID: mdl-30005603

ABSTRACT

BACKGROUND: Nitrogen (N) is a key macronutrient essential for plant growth, and its availability has a strong influence on crop development. The application of synthetic N fertilizers on crops has increased substantially in recent decades; however, the applied N is not fully utilized due to the low N use efficiency of crops. To overcome this limitation, it is important to understand the genome-wide responses and functions of key genes and potential regulatory factors in N metabolism. RESULTS: Here, we characterized changes in the rice (Oryza sativa) transcriptome, including genes, newly identified putative long non-coding RNAs (lncRNAs), and microRNAs (miRNAs) and their target mRNAs in response to N starvation using four different transcriptome approaches. Analysis of rice genes involved in N metabolism and/or transport using strand-specific RNA-Seq identified 2588 novel putative lncRNA encoding loci. Analysis of previously published RNA-Seq datasets revealed a group of N starvation-responsive lncRNAs showing differential expression under other abiotic stress conditions. Poly A-primed sequencing (2P-Seq) revealed alternatively polyadenylated isoforms of N starvation-responsive lncRNAs and provided precise 3' end information on the transcript models of these lncRNAs. Analysis of small RNA-Seq data identified N starvation-responsive miRNAs and down-regulation of miR169 family members, causing de-repression of NF-YA, as confirmed by strand-specific RNA-Seq and qRT-PCR. Moreover, we profiled the N starvation-responsive down-regulation of root-specific miRNA, osa-miR444a.4-3p, and Degradome sequencing confirmed MADS25 as a novel target gene. CONCLUSIONS: In this study, we used a combination of multiple RNA-Seq analyses to extensively profile the expression of genes, newly identified lncRNAs, and microRNAs in N-starved rice roots and shoots. Data generated in this study provide an in-depth understanding of the regulatory pathways modulated by N starvation-responsive miRNAs. The results of comprehensive, large-scale data analysis provide valuable information on multiple aspects of the rice transcriptome, which may be useful in understanding the responses of rice plants to changes in the N supply status of soil.


Subject(s)
Nitrogen/deficiency , Oryza/genetics , RNA, Untranslated/genetics , Transcriptome , Gene Expression Profiling , Gene Expression Regulation, Plant , MicroRNAs/genetics , MicroRNAs/metabolism , Oryza/metabolism , Phosphates/metabolism , Plant Roots/genetics , Plant Roots/metabolism , Plant Shoots/genetics , Plant Shoots/metabolism , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , RNA, Untranslated/metabolism , Sequence Analysis, RNA , Stress, Physiological
5.
Plant Cell ; 29(6): 1406-1424, 2017 Jun.
Article in English | MEDLINE | ID: mdl-28536144

ABSTRACT

The transcription factor MYC2 has emerged as a master regulator of jasmonate (JA)-mediated responses as well as crosstalk among different signaling pathways. The instability of MYC2 is in part due to the action of PUB10 E3 ligase, which can polyubiquitinate this protein. Here, we show that polyubiquitinated MYC2 can be deubiquitinated by UBP12 and UBP13 in vitro, suggesting that the two deubiquitinating enzymes can counteract the effect of PUB10 in vivo. Consistent with this view, UBP12 and UBP13 associate with MYC2 in the nucleus. Transgenic Arabidopsis thaliana plants deficient in UBP12 and UBP13 show accelerated decay of MYC2 and are hyposensitive to JA, whereas plants overexpressing UBP12 or UBP13 have prolonged MYC2 half-life and are hypersensitive to JA Our results suggest that there is a genetic link between UBP12, UBP13, and MYC2. Our results identify UBP12 and UBP13 as additional positive regulators of JA responses and suggest that these enzymes likely act by stabilizing MYC2.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Arabidopsis/metabolism , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , Cyclopentanes/pharmacology , Endopeptidases/metabolism , Oxylipins/pharmacology , Arabidopsis/drug effects , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/genetics , Endopeptidases/genetics , Gene Expression Regulation, Plant/drug effects , Gene Expression Regulation, Plant/genetics , Plants, Genetically Modified/drug effects , Plants, Genetically Modified/enzymology , Plants, Genetically Modified/genetics , Plants, Genetically Modified/metabolism
6.
Plant Biotechnol J ; 15(6): 754-764, 2017 Jun.
Article in English | MEDLINE | ID: mdl-27892643

ABSTRACT

Drought has a serious impact on agriculture worldwide. A plant's ability to adapt to rhizosphere drought stress requires reprogramming of root growth and development. Although physiological studies have documented the root adaption for tolerance to the drought stress, underlying molecular mechanisms is still incomplete, which is essential for crop engineering. Here, we identified OsNAC6-mediated root structural adaptations, including increased root number and root diameter, which enhanced drought tolerance. Multiyear drought field tests demonstrated that the grain yield of OsNAC6 root-specific overexpressing transgenic rice lines was less affected by drought stress than were nontransgenic controls. Genome-wide analyses of loss- and gain-of-function mutants revealed that OsNAC6 up-regulates the expression of direct target genes involved in membrane modification, nicotianamine (NA) biosynthesis, glutathione relocation, 3'-phophoadenosine 5'-phosphosulphate accumulation and glycosylation, which represent multiple drought tolerance pathways. Moreover, overexpression of NICOTIANAMINE SYNTHASE genes, direct targets of OsNAC6, promoted the accumulation of the metal chelator NA and, consequently, drought tolerance. Collectively, OsNAC6 orchestrates novel molecular drought tolerance mechanisms and has potential for the biotechnological development of high-yielding crops under water-limiting conditions.


Subject(s)
Oryza/metabolism , Plant Proteins/metabolism , Plant Roots/metabolism , Transcription Factors/metabolism , Azetidinecarboxylic Acid/analogs & derivatives , Azetidinecarboxylic Acid/metabolism , Biotechnology , Droughts , Gene Expression Regulation, Plant/genetics , Gene Expression Regulation, Plant/physiology , Genome-Wide Association Study , Oryza/genetics , Plant Proteins/genetics , Plant Roots/genetics , Plants, Genetically Modified/genetics , Plants, Genetically Modified/metabolism , Transcription Factors/genetics
7.
Plant Physiol ; 172(1): 575-88, 2016 09.
Article in English | MEDLINE | ID: mdl-27382137

ABSTRACT

Plant responses to drought stress require the regulation of transcriptional networks via drought-responsive transcription factors, which mediate a range of morphological and physiological changes. AP2/ERF transcription factors are known to act as key regulators of drought resistance transcriptional networks; however, little is known about the associated molecular mechanisms that give rise to specific morphological and physiological adaptations. In this study, we functionally characterized the rice (Oryza sativa) drought-responsive AP2/ERF transcription factor OsERF71, which is expressed predominantly in the root meristem, pericycle, and endodermis. Overexpression of OsERF71, either throughout the entire plant or specifically in roots, resulted in a drought resistance phenotype at the vegetative growth stage, indicating that overexpression in roots was sufficient to confer drought resistance. The root-specific overexpression was more effective in conferring drought resistance at the reproductive stage, such that grain yield was increased by 23% to 42% over wild-type plants or whole-body overexpressing transgenic lines under drought conditions. OsERF71 overexpression in roots elevated the expression levels of genes related to cell wall loosening and lignin biosynthetic genes, which correlated with changes in root structure, the formation of enlarged aerenchyma, and high lignification levels. Furthermore, OsERF71 was found to directly bind to the promoter of OsCINNAMOYL-COENZYME A REDUCTASE1, a key gene in lignin biosynthesis. These results indicate that the OsERF71-mediated drought resistance pathway recruits factors involved in cell wall modification to enable root morphological adaptations, thereby providing a mechanism for enhancing drought resistance.


Subject(s)
Droughts , Gene Expression Regulation, Plant , Oryza/genetics , Plant Proteins/genetics , Plant Roots/genetics , Transcription Factors/genetics , Adaptation, Physiological/genetics , Gene Expression Profiling/methods , Microscopy, Confocal , Oryza/anatomy & histology , Plant Proteins/metabolism , Plant Roots/anatomy & histology , Plants, Genetically Modified , Reverse Transcriptase Polymerase Chain Reaction , Transcription Factors/metabolism
8.
Plant Sci ; 241: 199-210, 2015 Dec.
Article in English | MEDLINE | ID: mdl-26706071

ABSTRACT

The mechanisms of plant response and adaptation to drought stress require the regulation of transcriptional networks via the induction of drought-responsive transcription factors. Nuclear Factor Y (NF-Y) transcription factors have aroused interest in roles of plant drought stress responses. However, the molecular mechanism of the NF-Y-induced drought tolerance is not well understood. Here, we functionally analyzed two rice NF-YA genes, OsNF-YA7 and OsNF-YA4. Expression of OsNF-YA7 was induced by drought stress and its overexpression in transgenic rice plants improved their drought tolerance. In contrast, OsNF-YA4 expression was not increased by drought stress and its overexpression in transgenic rice plants did not affect their sensitivity to drought stress. OsNF-YA4 expression was highly induced by the stress-related hormone abscisic acid (ABA), while OsNF-YA7 was not, indicating that OsNF-YA7 mediates drought tolerance in an ABA-independent manner. Analysis of the OsNF-YA7 promoter revealed three ABA-independent DRE/CTR elements and RNA-seq analysis identified 48 genes downstream of OsNFYA7 action putatively involved in the OsNF-YA7-mediated drought tolerance pathway. Taken together, our results suggest an important role for OsNF-YA7 in rice drought stress tolerance.


Subject(s)
Abscisic Acid/metabolism , Droughts , Gene Expression Regulation, Plant , Oryza/physiology , Plant Proteins/genetics , Transcription Factors/genetics , CCAAT-Binding Factor , Oryza/genetics , Plant Proteins/metabolism , Plants, Genetically Modified/genetics , Plants, Genetically Modified/physiology , Stress, Physiological , Transcription Factors/metabolism
9.
Planta ; 237(1): 211-24, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23007553

ABSTRACT

To be effective in crop biotechnology applications, gene promoters need to be stably active over sequential generations in a population of single-copy transgenic lines. Most of the stress-inducible promoters characterized in plants thus far have been analyzed at early (T0, T1 or T2) generations and/or by testing only a small number of transgenic lines. In our current study, we report our analysis of OsNCED3, a stress-inducible rice promoter involved in ABA biosynthesis, in various organs and tissues of transgenic rice plants over the T(2-4) homozygous generations. The transgene copy numbers in the lines harboring the OsNCED3:gfp construct were determined and six single- and two double-copy transgenic lines were analyzed for promoter activity in comparison with the Wsi18, a stress-inducible promoter previously characterized. The exogenous promoter activities were found to be significantly enhanced in the roots and leaves, whereas zero or low levels of activity were evident in grains and flowers, under drought and high-salinity conditions. The highest induction levels of gfp transcripts in the OsNCED3:gfp plants upon drought treatments were 161- and 93-fold in leaves and roots, respectively, and these levels were comparable with those of gfp transcripts in the Wsi18:gfp plants. A comparison of the promoter activities between the T2-T4 plants revealed that comparable activity levels were maintained over these three homozygous generations with no evidence of silencing. Thus, our results provide the OsNCED3 promoter that is stress-inducible in a whole rice plant except for in the aleurones and endosperm and stably active over three generations.


Subject(s)
Oryza/genetics , Plant Proteins/genetics , Plants, Genetically Modified/genetics , Promoter Regions, Genetic/genetics , Droughts , Flowers/genetics , Flowers/metabolism , Gene Expression Regulation, Plant/drug effects , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Microscopy, Confocal , Microscopy, Fluorescence , Oryza/metabolism , Plant Leaves/genetics , Plant Leaves/metabolism , Plant Proteins/metabolism , Plant Roots/genetics , Plant Roots/metabolism , Plants, Genetically Modified/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Sodium Chloride/pharmacology , Stress, Physiological , Time Factors , Water/pharmacology
10.
Plant Biotechnol J ; 11(1): 101-14, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23094910

ABSTRACT

Drought conditions are among the most serious challenges to crop production worldwide. Here, we report the results of field evaluations of transgenic rice plants overexpressing OsNAC5, under the control of either the root-specific (RCc3) or constitutive (GOS2) promoters. Field evaluations over three growing seasons revealed that the grain yield of the RCc3:OsNAC5 and GOS2:OsNAC5 plants were increased by 9%-23% and 9%-26% under normal conditions, respectively. Under drought conditions, however, RCc3:OsNAC5 plants showed a significantly higher grain yield of 22%-63%, whilst the GOS2:OsNAC5 plants showed a reduced or similar yield to the nontransgenic (NT) controls. Both the RCc3:OsNAC5 and GOS2:OsNAC5 plants were found to have larger roots due to an enlarged stele and aerenchyma at flowering stage. Cell numbers per cortex layer and stele of developing roots were higher in both transgenic plants than NT controls, contributing to the increase in root diameter. The root diameter was enlarged to a greater extent in the RCc3:OsNAC5, suggesting the importance of this phenotype for enhanced drought tolerance. Microarray experiments identified 25 up-regulated genes by more than three-fold (P < 0.01) in the roots of both transgenic lines. Also identified were 19 and 18 up-regulated genes that are specific to the RCc3:OsNAC5 and GOS2:OsNAC5 roots, respectively. Of the genes specifically up-regulated in the RCc3:OsNAC5 roots, GLP, PDX, MERI5 and O-methyltransferase were implicated in root growth and development. Our present findings demonstrate that the root-specific overexpression of OsNAC5 enlarges roots significantly and thereby enhances drought tolerance and grain yield under field conditions.


Subject(s)
Droughts , Oryza/growth & development , Oryza/genetics , Plant Roots/growth & development , Plant Roots/genetics , Seeds/growth & development , Transcription Factors/metabolism , Gene Expression Regulation, Plant , Genes, Plant , Genetic Variation , Genotype , Phenotype , Plants, Genetically Modified , Promoter Regions, Genetic , Seeds/genetics , Stress, Physiological
11.
Planta ; 235(6): 1397-408, 2012 Jun.
Article in English | MEDLINE | ID: mdl-22212906

ABSTRACT

We have previously characterized the constitutively active promoters of the APX, PGD1 and R1G1B genes in rice (Park et al. 2010 in J Exp Bot 61:2459-2467). To have potential crop biotechnology applications, gene promoters must be stably active over many generations. In our current study, we report our further detailed analysis of the APX, PGD1 and R1G1B gene promoters in various organs and tissues of transgenic rice plants for three (T3₋5) homozygous generations. The copy numbers in 37 transgenic lines that harbor promoter:gfp constructs were determined and promoter activities were measured by real-time qPCR. Analysis of the 37 lines revealed that 15 contained a single copy of one of the three promoter:gfp chimeric constructs. The promoter activity levels were generally higher in multi-copy lines, whereas variations in these levels over the T3₋5 generations studied were observed to be smaller in single-copy than in multi-copy lines. The three promoters were further found to be highly active in the whole plant body at both the vegetative and reproductive stages of plant growth, with the exception of the APX in the ovary and R1G1B in the pistil and filaments where zero or very low levels of activity were detected. Of note, the spatial activities of the PGD1 promoter were found to be strikingly similar to those of the ZmUbi1, a widely used constitutive promoter. Our comparison of promoter activities between T3, T4 and T5 plants revealed that the APX, PGD1 and R1G1B promoters maintained their activities at comparable levels in leaves and roots over three homozygous generations and are therefore potentially viable alternative promoters for crop biotechnology applications.


Subject(s)
Gene Expression Regulation, Plant , Genes, Plant/genetics , Homozygote , Organ Specificity/genetics , Oryza/genetics , Promoter Regions, Genetic , Gene Dosage/genetics , Green Fluorescent Proteins/metabolism , Plant Leaves/genetics , Plant Proteins/genetics , Plant Proteins/metabolism , Plant Roots/genetics , Plants, Genetically Modified , RNA, Messenger/genetics , RNA, Messenger/metabolism
12.
Transgenic Res ; 20(1): 153-63, 2011 Feb.
Article in English | MEDLINE | ID: mdl-20490919

ABSTRACT

There is currently a shortage of efficient promoters for stress-inducible gene expression, especially in monocotyledonous crops. Here, we report analysis of the rice Wsi18 promoter, a member of the group 3 Lea family, in transgenic rice plants. The abundance of Wsi18 mRNA increased in leaf tissues within 2 h of exposure to NaCl or abscisic acid (ABA) and within 6 h of exposure to drought, but there was no transcript increase in response to low-temperature conditions. Wsi18 mRNA accumulated in the roots similarly to in the leaves, but at a faster rate. The promoter was linked to the GFP reporter gene, transformed into rice, and its activity was analyzed in transgenic plants at all stages of plant growth from calli, vegetative tissues, flowers, and to dry seeds, both before and after stress treatment. The activity of the promoter was significantly increased in the whole plant body, including flowers, on exposure of plants to stress conditions, with very low levels of basal activity in all tissues. Moreover, the promoter was found to be predominantly active in the whole grain, including endosperm, embryo, and aleurone layer during seed development. Together, we have identified and analyzed the Wsi18 promoter and found a previously undescribed characteristic-a stress-inducible property in the whole plant body with activity in the whole grain during seed development.


Subject(s)
Gene Expression Regulation, Plant , Heat-Shock Response , Oryza/genetics , Plant Leaves/metabolism , Plant Proteins/metabolism , Plants, Genetically Modified/genetics , Promoter Regions, Genetic/genetics , Abscisic Acid/pharmacology , Base Sequence , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Molecular Sequence Data , Plant Leaves/genetics , Plant Proteins/chemistry , Plant Proteins/genetics , Promoter Regions, Genetic/physiology , RNA, Messenger/genetics , RNA, Messenger/metabolism , Seeds/growth & development , Sodium Chloride/pharmacology
13.
Planta ; 232(3): 743-54, 2010 Aug.
Article in English | MEDLINE | ID: mdl-20567981

ABSTRACT

There are few efficient promoters for use with stress-inducible gene expression in plants, and in particular for monocotyledonous crops. Here, we report the identification of six genes, Rab21, Wsi18, Lea3, Uge1, Dip1, and R1G1B that were induced by drought stress in rice microarray experiments. Gene promoters were linked to the gfp reporter and their activities were analyzed in transgenic rice plants throughout all stages of plant growth, from dry seeds to vegetative tissues to flowers, both before and after drought treatments. In fold induction levels, Rab21 and Wsi18 promoters ranged from 65- and 36-fold in leaves to 1,355- and 492-fold in flowers, respectively, whereas Lea3 and Uge1 were higher in leaves, but lower in roots and flowers, as compared with Rab21 and Wsi18. Dip1 and R1G1B promoters had higher basal levels of activity under normal growth conditions in all tissues, resulting in smaller fold-induction levels than those of the others. In drought treatment time course, activities of Dip1 and R1G1B promoters rapidly increased, peaked at 2 h, and remained constant until 8 h, while that of Lea3 slowly yet steadily increased until 8 h. Interestingly, Rab21 activity increased rapidly and steadily in response to drought stress until expression peaked at 8 h. Thus, we have isolated and characterized six rice promoters that are all distinct in fold induction, tissue specificity, and induction kinetics under drought conditions, providing a variety of drought-inducible promoters for crop biotechnology.


Subject(s)
Droughts , Oryza/genetics , Plants, Genetically Modified/genetics , Promoter Regions, Genetic , Blotting, Southern , Gene Expression Regulation, Plant , Genes, Plant , Oryza/growth & development , Plants, Genetically Modified/growth & development , Polymerase Chain Reaction
14.
Plant Physiol ; 153(1): 185-97, 2010 May.
Article in English | MEDLINE | ID: mdl-20335401

ABSTRACT

Drought poses a serious threat to the sustainability of rice (Oryza sativa) yields in rain-fed agriculture. Here, we report the results of a functional genomics approach that identified a rice NAC (an acronym for NAM [No Apical Meristem], ATAF1-2, and CUC2 [Cup-Shaped Cotyledon]) domain gene, OsNAC10, which improved performance of transgenic rice plants under field drought conditions. Of the 140 OsNAC genes predicted in rice, 18 were identified to be induced by stress conditions. Phylogenic analysis of the 18 OsNAC genes revealed the presence of three subgroups with distinct signature motifs. A group of OsNAC genes were prescreened for enhanced stress tolerance when overexpressed in rice. OsNAC10, one of the effective members selected from prescreening, is expressed predominantly in roots and panicles and induced by drought, high salinity, and abscisic acid. Overexpression of OsNAC10 in rice under the control of the constitutive promoter GOS2 and the root-specific promoter RCc3 increased the plant tolerance to drought, high salinity, and low temperature at the vegetative stage. More importantly, the RCc3:OsNAC10 plants showed significantly enhanced drought tolerance at the reproductive stage, increasing grain yield by 25% to 42% and by 5% to 14% over controls in the field under drought and normal conditions, respectively. Grain yield of GOS2:OsNAC10 plants in the field, in contrast, remained similar to that of controls under both normal and drought conditions. These differences in performance under field drought conditions reflect the differences in expression of OsNAC10-dependent target genes in roots as well as in leaves of the two transgenic plants, as revealed by microarray analyses. Root diameter of the RCc3:OsNAC10 plants was thicker by 1.25-fold than that of the GOS2:OsNAC10 and nontransgenic plants due to the enlarged stele, cortex, and epidermis. Overall, our results demonstrated that root-specific overexpression of OsNAC10 enlarges roots, enhancing drought tolerance of transgenic plants, which increases grain yield significantly under field drought conditions.


Subject(s)
Droughts , Oryza/metabolism , Plant Proteins/metabolism , Stress, Physiological , Transcription Factors/metabolism , Amino Acid Sequence , Biomass , Gene Expression Profiling , Molecular Sequence Data , Oryza/genetics , Oryza/growth & development , Plant Proteins/genetics , Plant Roots/metabolism , Transcription Factors/genetics , Up-Regulation
15.
Plant J ; 59(5): 764-76, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19453457

ABSTRACT

We have previously isolated a rice gene encoding a histone deacetylase, OsHDAC1, and observed that its transgenic overexpression increases seedling root growth. To identify the transcriptional repression events that occur as a result of OsHDAC1 overexpression (OsHDAC1(OE)), a global profiling of root-expressed genes was performed on OsHDAC1(OE) or HDAC inhibitor-treated non-transgenic (NT) roots, in comparison with untreated NT roots. We selected 39 genes that are induced and repressed in HDAC inhibitor-treated NT and OsHDAC1(OE) roots, compared with NT roots, respectively. Interestingly, OsNAC6, a member of the NAM-ATAF-CUC (NAC) family, was identified as a key component of the OsHDAC1 regulon, and was found to be epigenetically repressed by OsHDAC1 overexpression. The root phenotype of OsNAC6 knock-out seedlings was observed to be similar to that of the OsHDAC1(OE) seedlings. Conversely, the root phenotype of the OsNAC6 overexpressors was similar to that of the OsHDAC1 knock-out seedlings. These observations indicate that OsHDAC1 negatively regulates the OsNAC6 gene that primarily mediates the alteration in the root growth of the OsHDAC1(OE) seedlings. Chromatin immunoprecipitation assays of the OsNAC6 promoter region using antibodies specific to acetylated histones H3 and H4 revealed that OsHDAC1 epigenetically represses the expression of OsNAC6 by deacetylating K9, K14 and K18 on H3 and K5, K12 and K16 on H4.


Subject(s)
Histone Deacetylases/metabolism , Oryza/genetics , Plant Proteins/metabolism , Plant Roots/growth & development , Epigenesis, Genetic , Gene Expression Regulation, Plant , Histone Deacetylases/genetics , Oligonucleotide Array Sequence Analysis , Oryza/enzymology , Oryza/growth & development , Plant Proteins/genetics , Plant Roots/enzymology , Plant Roots/genetics , RNA, Plant/genetics , Seedlings/enzymology , Seedlings/genetics , Seedlings/growth & development
16.
Plant Physiol ; 150(3): 1368-79, 2009 Jul.
Article in English | MEDLINE | ID: mdl-19429605

ABSTRACT

Transcription factors with an APETELA2 (AP2) domain have been implicated in various cellular processes involved in plant development and stress responses. Of the 139 AP2 genes predicted in rice (Oryza sativa), we identified 42 genes in our current study that are induced by one or more stress conditions, including drought, high salinity, low temperature, and abscisic acid. Phylogenic analysis of these 42 stress-inducible AP2 genes revealed the presence of six subgroups (I-VI) with distinct signature motifs. Two genes, AP37 and AP59, representing subgroups I and II, respectively, were functionally characterized. Both genes were found to be induced upon 2 h of exposure to drought and high-salinity conditions but to differ in their expression profile upon exposure to low temperature and abscisic acid. The overexpression of AP37 and AP59 in rice under the control of the constitutive promoter OsCc1 increased the tolerance to drought and high salinity at the vegetative stage. Increased tolerance to low temperatures was observed only in OsCc1:AP37 plants. More importantly, the OsCc1:AP37 plants showed significantly enhanced drought tolerance in the field, which increased grain yield by 16% to 57% over controls under severe drought conditions, yet exhibited no significant difference under normal growth conditions. In contrast, grain yield in OsCc1:AP59 plants in the field was reduced by 23% to 43% compared with controls under both normal and drought stress conditions. Microarray experiments identified 10 and 38 genes that are up-regulated by AP37 and AP59, respectively, in addition to 37 genes that are commonly induced by both factors. Our results suggest that the AP37 gene has the potential to improve drought tolerance in rice without causing undesirable growth phenotypes.


Subject(s)
Oryza/metabolism , Plant Proteins/physiology , Transcription Factors/physiology , Amino Acid Sequence , Droughts , Molecular Sequence Data , Oryza/genetics , Oryza/growth & development , Plant Proteins/genetics , Plant Proteins/metabolism , Sequence Alignment , Stress, Psychological , Transcription Factors/genetics , Transcription Factors/metabolism
17.
Article in Korean | WPRIM (Western Pacific) | ID: wpr-152882

ABSTRACT

BACKGROUND: Diffusion tensor MRI (DTI) is a new imaging technique and enables us to analyze the structural damage of fiber pathways and to monitor the time course of Wallerian degeneration of the pyramidal tract in stroke patients. We used DTI to investigate structural changes of the infarct area and the associated descending corticospinal tract in patients with subcortical infarct. METHODS: We examined 24 consecutive patients who presented with acute single cerebral infarct in the subcortical area and who also had undergone an MRI study within 7 days after symptom onset. Clinical outcome was assessed using the National Institutes of Health Stroke Scale (NIHSS) at admission, 7 days, 14 days and 30 days and modified Rankin Scale (mRS) at admission and 30 days. Each of the indices was achieved by post processing the acquired DTI data and correlated with the NIHSS. RESULTS: In infarct region, fractional anisotropy (FA) was significantly decreased compared with matched-contralateral regions (0.39 vs. 0.53, p<0.001). In the distal to the infarct, FA was significantly decreased at internal capsule (0.62 vs. 0.64, p=0.019), not at pons (0.51 vs. 0.53, p=0.103). The decrease of anisotropy at infarct region correlated positively with the NIHSS at 7, 14 and 30 days and mRS at 30 days after stroke, but the decrease of anisotropy at internal capsule did not correlate with the NIHSS. CONCLUSIONS: This study shows the potential of DTI to detect and monitor the structural degeneration of fiber pathways and to establish the prognosis in patients with acute subcortical cerebral infarct.


Subject(s)
Humans , Anisotropy , Cerebral Infarction , Diffusion , Internal Capsule , Magnetic Resonance Imaging , Pons , Prognosis , Pyramidal Tracts , Stroke , Wallerian Degeneration
18.
Article in English | WPRIM (Western Pacific) | ID: wpr-54779

ABSTRACT

A 58-year-old woman presented with an acute embolic occlusion of the distal basilar artery. She underwent angioplasty and intra-arterial thrombolysis. Angiography performed after recanalization revealed a single perforating thalamic artery. A nonenhanced CT scan carried out immediately after the procedure revealed hyperdense lesions in the bilateral paramedian portions of the thalami, which disappeared on the 24-hour follow-up CT scan. Three months later, the patient improved to functional independence, but had some memory dysfunction and vertical gaze palsy. This case suggests that contrast enhancement or extravasation can occur in the thalamus after intra-arterial thrombolysis performed to recanalize a basilar artery occlusion.


Subject(s)
Female , Humans , Middle Aged , Angioplasty , Basilar Artery/diagnostic imaging , Contrast Media , Intracranial Embolism and Thrombosis/diagnostic imaging , Thalamus/blood supply , Thrombolytic Therapy , Tomography, X-Ray Computed
19.
Article in Korean | WPRIM (Western Pacific) | ID: wpr-117058

ABSTRACT

PURPOSE: To assess the usefulness of diffusion-weighted MR imaging (DWI) and apparent diffusion coefficient (ADC) in the initial and follow-up studies of patients with neuro-Behcet's disease. MATERIALS AND METHODS: Six patients diagnosed with neuro-Behcet's disease were the subjects of this study. Initial and follow-up MR imaging were obtained in all six patients. Initial and follow-up DWI were also obtained in four of the six patients, with only an initial DWI in the other two. The DWI were obtained using multi-shot echo planar imaging, on a 1.5T MR unit, with two gradient steps (b values of 0, 1000 sec/mm2). The ADC value and ADC maps were obtained using commercial software. The locations and signal intensities of the lesions were analyzed on conventional MRI and DWI, respectively. The ADC values of the lesions were calculated on the initial and follow-up DWI, and compared those of lesions in the normal contralateral regions. RESULTS:The initial DWI showed iso-signal intensities in four of the six patients, with high signal intensities in the other two. In five of the six patients, including three of the four that showed isosignal intensities and the two that showed high signal intensities on the initial DWI, the ADC values of the involved lesions were higher than those of the normal contralateral regions. In three of four that showed isosignal intensities, the ADC values of the lesions were decreased and normalized on the follow-up DWI. CONCLUSION: Obtaining DWI and ADC values in patients with neuro-Behcet's disease may be helpful in the understanding of pathophysiology and differential diagnosis of this disease.


Subject(s)
Humans , Diagnosis, Differential , Diffusion , Echo-Planar Imaging , Follow-Up Studies , Magnetic Resonance Imaging
20.
Article in Korean | WPRIM (Western Pacific) | ID: wpr-91264

ABSTRACT

PURPOSE: This study was designed to evaluate the usefulness of 3T-TOF MR angiography (3T-TOF MRA) compared with transcranial Doppler sonography (TCD) and conventional angiography (CA) in patients with suspected cerebral infarction. MATERIALS AND METHODS: Fifty four patients with clinical symptoms of cerebral infarction were involved in this study, and had undergone 3T-TOF MRA and TCD, with CA in 11 patients. On the basis of divisions of the carotid artery, four groups were designated: group I, both vertebral arteries and basilar artery; group II, segment between 2 cm below bifurcation of common carotid artery and genu portion of internal carotid artery; group III, segment between petrous portion of internal carotid artery and bifurcation of anterior and middle cerebral artery; group IV, from bifurcation of anterior and middle cerebral artery to thier distal branches. Two radiologists retrospectively reviewed the vascular imaging and stenosis in 3T-TOF MRA, TCD, and CA. RESULTS: A total of 432 arteries, 108 in each group, were available. The assessment of vascular imaging quality in 3T-TOF MRA is scored 2.98, 2.96, 2.91, 2.88 in 4 groups, respectively. Agreement among 3TTOF MR angiography, TCD, and CA was high. CONCLUSION: 3T-TOF MR angiography may be useful method for the assessment of stenotic lesions of cranial vasculature in patients with cerebral infarction.


Subject(s)
Humans , Angiography , Arteries , Basilar Artery , Carotid Arteries , Carotid Artery, Common , Carotid Artery, Internal , Cerebral Infarction , Constriction, Pathologic , Middle Cerebral Artery , Retrospective Studies , Ultrasonography, Doppler, Transcranial , Vertebral Artery
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