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杨树GABA支路3个基因家族的鉴定和表达分析
陈炜, 成铁龙, 纪敬, 武妍妍, 谢田田, 江泽平, 史胜青
南京林业大学学报（自然科学版） ›› 2020, Vol. 44 ›› Issue (5) : 67-77.
PDF(4369 KB)
PDF(4369 KB)
杨树GABA支路3个基因家族的鉴定和表达分析
Identification of three gene families in the GABA shunt and their expression analysis in poplar
【目的】 γ-氨基丁酸(GABA)与植物的生长发育有着密切的联系。结合前期对GABA抑制杨树不定根发育的研究,对GABA支路基因家族的特征和表达模式进行研究,为进一步解释其在不定根发育中的作用奠定基础。【方法】从银白杨(Populus alba)×P. glandulosa ‘84K’(‘84K’杨)基因组中鉴定出GABA支路的PopGAD、PopGABA-T和PopSSADH 3个基因家族成员,并利用生物信息学方法分析其特征,以及与其他物种相关基因家族的亲缘关系;利用qRT-PCR研究外源GABA及其降解抑制剂vigabatrin(VGB)对各基因表达的影响。【结果】①PopGAD、PopGABA-T和PopSSADH 3个基因家族成员数量依次为6、2和2个,启动子序列中主要包含与光、激素和环境等响应相关元件。②系统进化分析表明,PopGAD家族中PopGAD6与家族其他成员亲缘关系较远;PopGABA-T和PopSSADH家族中的两个基因成员亲缘关系较近。③基因表达分析表明,不定根生长过程中GABA支路基因总体上在根中表达量高于茎和叶。外源GABA或VGB处理对PopGAD和PopGABA-T两个基因家族成员在根、茎和叶中的表达量影响程度不同,但对 PopSSADH基因家族的表达则无明显影响。【结论】 GABA支路3个基因家族在‘84K’杨树响应光、激素和环境胁迫等方面具有重要作用。外源GABA和VGB处理对PopGAD和PopGABA-T家族成员的影响较大,并且均在根中表达量最高,表明这两个基因家族在不定根生长过程中发挥着重要调控功能。这为深入解析GABA在树木不定根发生中的作用机制奠定了基础。
【Objective】Most prokaryotic and eukaryotic organisms contain γ-aminobutyric acid (GABA), a four-carbon non-protein amino acid, which plays a central role in carbon and nitrogen metabolism. The GABA also occurs in mammals, as it is a major inhibitory neurotransmitter; in addition, it is related to growth and development of plants. Based on our recent findings about the role of GABA in inhibiting poplar adventitious root growth, we studied the characteristics of three gene families in the GABA shunt and their expression patterns to provide a basis for the further investigation of GABA’s function in regulating adventitious root development. 【Method】 The members of three gene families, PopGAD, PopGABA-T and PopSSADH were identified from the complete genome sequence of Populus alba × P. glandulosa ‘84K’ (‘84K’ poplar). The gene structures, conserved motifs, physicochemical characters of proteins and promotor sequences were investigated by using bioinformatics as well as examination of the phylogenetic relationships in these gene families between the 11 species. Tissue-cultured poplar seedlings were cultivated for 12 days with different concentrations of GABA or VGB (GABA degradation inhibitor vigabatrin) treatments. The expression pattern of each gene family was analyzed in the different tissues and treatments with exogenous GABA and VGB by qRT-PCR. 【Result】 ① The PopGAD gene family has six members, the exon numbers are 5-7, and the protein molecular weights are 45.8-57.6 ku; the PopGABA-T gene family has two members, both have 19 exons, and their protein molecular weights are 56.5 ku; the PopSSADH gene family also has two members, there are 4 and 20 exons, respectively, and their protein molecular weights are 57.5 and 15.9 ku, respectively. The gene structure analysis revealed that: all gene members have introns and exons, and the same subfamily exhibits a similar exon-intron pattern; all proteins have three motifs; the promoter sequences of the three gene families all contain elements related to light-, hormone-and stress-responses. However, there were some differences. The promoters of PopGAD gene families have elements related to defense, stress response, and flavonoid biosynthesis. However, these elements are absent in PopGABA-T gene families, and PopSSADH gene families only have defense and stress response elements. ② The phylogenetic analysis of the 11 species showed the GAD gene family is divided into 10 clusters. PopGAD1, PopGAD2, PopGAD3 and PopGAD4, PopGAD5 have close relationships. PopGAD6, which has a less close relationship with other gene family members, belongs to cluster IV, whereas other members all belong to cluster I. Both PopGABA-T and PopSSADH gene families have three clusters. PopGABA-T1 and PopGABA-T2 come from a common ancestor belonging to cluster Ⅲ and PopSSADH1, PopSSADH2 have a close relationship and belong to cluster I. ③ The gene expression analysis showed that all genes of three family members in the GABA shunt, except for PopGAD3 and PopSSADH1, display a higher expression in roots than that in stems and leaves during adventitious root growth. Moreover, the expression of PopGAD1 and PopGAD2 is far beyond that of the other four family members whether or not exogenous GABA or VGB were applied. Exogenous GABA treatments up-regulated PopGAD1 and PopGAD2 in stems and leaves under certain concentrations during adventitious root growth but inhibited the expression of PopGABA-T1 in roots and stems. Exogenous VGB treatments significantly promoted PopGAD1 and PopGAD2 expression in leaves, but inhibited them in roots and stems. Exogenous VGB treatments inhibited both PopGABA-T members in roots. The expression of PopSSADH1 was at a very low level under both GABA and VGB treatments. The expression of PopSSADH2 was far beyond PopSSADH1 and was more inhibited in leaves than in the root and stem. 【Conclusion】 In the GABA shunt, six PopGAD members, two PopGABA-T members and two PopSSADH members were identified from ‘84K’ poplar. The bioinformatics analysis showed that three gene families in the GABA shunt have crucial roles in poplar response to light, hormone and environmental stresses. Both GABA and VGB treatments had a greater impact on the expression of both PopGAD and PopGABA-T families, which have higher expression levels in roots, indicating that these two families may play an important role in regulating poplar adventitious root growth, which would lay a foundation for further deciphering the functions of GABA in adventitious root development in trees.
基因家族 / GABA支路 / γ-氨基丁酸(GABA) / 进化分析 / 组织特异性表达 / 杨树
gene family / GABA shunt / γ-aminobutyric acid (GABA) / phylogenetic analysis / tissue specific expression / poplar
| [1] |
施征, 史胜青, 钟传飞 , 等. γ-氨基丁酸在植物抗逆生理及调控中的作用[J]. 生命科学研究, 2007,11(S1):57-61.
|
| [2] |
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| [3] |
gamma-aminobutyric acid (GABA) is a four-carbon non-protein amino acid presented in a wide range of organisms. In this study, a suppression subtractive hybridization (SSH) library was constructed using roots of a legume shrub, Caragana intermedia, with the combined treatment of 300 mm NaCl and 300 mm NaCl + 10 mm GABA. We obtained 224 GABA-regulated unique expressed sequence tags (ESTs) including signal transduction, transcriptional regulation, hormone biosynthesis, reactive oxygen species (ROS) and polyamine metabolism, etc. The key H(2)O(2)-generated genes, NADPH oxidase (CaGR60), peroxidase (CaGR61) and amine oxidase (CaGR62), were regulated at the mRNA level by 10 mm GABA, which clearly inhibited H(2)O(2) accumulation brought about by NaCl stress in roots and leaves with the observation of 3,3'-diaminobenzidine (DAB) staining. Similarly, 10 mm GABA also regulated the expression of 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase (ACO) genes (CaGR30 and CaGR31) and ethylene production in NaCl-treated roots. Surprisingly, these H(2)O(2)-generated genes were enhanced at the mRNA level by a lower concentration of GABA, at 0.25 mm, but not other alternative nitrogen sources, and endogenous GABA accumulated largely just by the application of GABA at either concentration. Our results further proved that GABA, as a signal molecule, participates in regulating the expression of genes in plants under salt stress.
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| [4] |
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| [5] |
Plant aluminum-activated malate transporters (ALMTs) are currently classified as anion channels; they are also known to be regulated by diverse signals, leading to a range of physiological responses. Gamma-aminobutyric acid (GABA) regulation of anion flux through ALMT proteins requires a specific amino acid motif in ALMTs that shares similarity with a GABA binding site in mammalian GABAA receptors. Here, we explore why TaALMT1 activation leads to a negative correlation between malate efflux and endogenous GABA concentrations ([GABA]i) in both wheat (Triticum aestivum) root tips and in heterologous expression systems. We show that TaALMT1 activation reduces [GABA]i because TaALMT1 facilitates GABA efflux but GABA does not complex Al(3+) TaALMT1 also leads to GABA transport into cells, demonstrated by a yeast complementation assay and via (14)C-GABA uptake into TaALMT1-expressing Xenopus laevis oocytes; this was found to be a general feature of all ALMTs we examined. Mutation of the GABA motif (TaALMT1(F213C)) prevented both GABA influx and efflux, and resulted in no correlation between malate efflux and [GABA]i We conclude that ALMTs are likely to act as both GABA and anion transporters in planta. GABA and malate appear to interact with ALMTs in a complex manner to regulate each other's transport, suggestive of a role for ALMTs in communicating metabolic status.
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| [6] |
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| [7] |
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| [8] |
史胜青, 齐力旺, 肖文发 , 等. 外源GABA对NaCl胁迫下中间锦鸡儿幼苗乙烯生成的调控作用[J]. 林业科学, 2008,44(9):26-30.
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| [9] |
Mitochondrial respiration and tricarboxylic acid (TCA) cycle activity are required during salt stress in plants to provide ATP and reductants for adaptive processes such as ion exclusion, compatible solute synthesis and reactive oxygen species (ROS) detoxification. However, there is a poor mechanistic understanding of how salinity affects mitochondrial metabolism, particularly respiratory substrate source. To determine the mechanism of respiratory changes under salt stress in wheat leaves, we conducted an integrated analysis of metabolite content, respiratory rate and targeted protein abundance measurements. Also, we investigated the direct effect of salt on mitochondrial enzyme activities. Salt-treated wheat leaves exhibit higher respiration rate and extensive metabolite changes. The activity of the TCA cycle enzymes pyruvate dehydrogenase complex and the 2-oxoglutarate dehydrogenase complex were shown to be directly salt-sensitive. Multiple lines of evidence showed that the gamma-aminobutyric acid (GABA) shunt was activated under salt treatment. During salt exposure, key metabolic enzymes required for the cyclic operation of the TCA cycle are physiochemically inhibited by salt. This inhibition is overcome by increased GABA shunt activity, which provides an alternative carbon source for mitochondria that bypasses salt-sensitive enzymes, to facilitate the increased respiration of wheat leaves.
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| [10] |
gamma-Aminobutyric acid (GABA) accumulates in many plant species in response to environmental stress. However, the physiological function of GABA or its metabolic pathway (GABA shunt) in plants remains largely unclear. Here, the genes, including glutamate decarboxylases (SlGADs), GABA transaminases (SlGABA-Ts) and succinic semialdehyde dehydrogenase (SlSSADH), controlling three steps of the metabolic pathway of GABA, were studied through virus-induced gene silencing approach in tomato. Silencing of SlGADs (GABA biosynthetic genes) and SlGABA-Ts (GABA catabolic genes) led to increased accumulation of reactive oxygen species (ROS) as well as salt sensitivity under 200 mm NaCl treatment. Targeted quantitative analysis of metabolites revealed that GABA decreased and increased in the SlGADs- and SlGABA-Ts-silenced plants, respectively, whereas succinate (the final product of GABA metabolism) decreased in both silenced plants. Contrarily, SlSSADH-silenced plants, also defective in GABA degradation process, showed dwarf phenotype, curled leaves and enhanced accumulation of ROS in normal conditions, suggesting the involvement of a bypath for succinic semialdehyde catabolism to gamma-hydroxybutyrate as reported previously in Arabidopsis, were less sensitive to salt stress. These results suggest that GABA shunt is involved in salt tolerance of tomato, probably by affecting the homeostasis of metabolites such as succinate and gamma-hydroxybutyrate and subsequent ROS accumulation under salt stress.
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| [11] |
Environmental constraints challenge cell homeostasis and thus require a tight regulation of metabolic activity. We have previously reported that the gamma-aminobutyric acid (GABA) metabolism is crucial for Arabidopsis salt tolerance as revealed by the NaCl hypersensitivity of the GABA transaminase (GABA-T, At3g22200) gaba-t/pop2-1 mutant. In this study, we demonstrate that GABA-T deficiency during salt stress causes root and hypocotyl developmental defects and alterations of cell wall composition. A comparative genome-wide transcriptional analysis revealed that expression levels of genes involved in carbon metabolism, particularly sucrose and starch catabolism, were found to increase upon the loss of GABA-T function under salt stress conditions. Consistent with the altered mutant cell wall composition, a number of cell wall-related genes were also found differentially expressed. A targeted quantitative analysis of primary metabolites revealed that glutamate (GABA precursor) accumulated while succinate (the final product of GABA metabolism) significantly decreased in mutant roots after 1 d of NaCl treatment. Furthermore, sugar concentration was twofold reduced in gaba-t/pop2-1 mutant roots compared with wild type. Together, our results provide strong evidence that GABA metabolism is a major route for succinate production in roots and identify GABA as a major player of central carbon adjustment during salt stress.
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| [12] |
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| [13] |
Glutamate decarboxylase (GAD) converts L-glutamate to gamma-aminobutyric acid (GABA), which is a non-protein amino acid present in all organisms. Plant GADs carry a C-terminal extension that binds to Ca(2+)/calmodulin (CaM) to modulate enzyme activity. However, rice possesses two distinct types of GAD, OsGAD1 and OsGAD2. Although they both have a C-terminal extension, the former peptide contains an authentic CaM-binding domain (CaMBD), which is common to dicotyledonous plants, while the latter does not. Therefore, the role of the C-terminal extension in functional expression of OsGAD2 was investigated. An in vitro enzyme assay using recombinant OsGAD2 proteins revealed low activity in the presence or absence of Ca(2+)/CaM. However, a truncated version of GAD2 (OsGAD2DeltaC) had over 40-fold higher activity than wild-type GAD at physiological pH. These two DNA constructs were introduced simultaneously into rice calli via Agrobacterium to establish transgenic cell lines. Free amino acids were isolated from several lines for each construct to determine GABA content. Calli overexpressing OsGAD2 and OsGAD2DeltaC had about 6-fold and 100-fold the GABA content of wild-type calli, respectively. Regenerated OsGAD2DeltaC rice plants had aberrant phenotypes such as dwarfism, etiolated leaves, and sterility. These data suggest that the C-terminal extension of OsGAD2 plays a role as a strong autoinhibitory domain, and that truncation of this domain causes the enzyme to act constitutively, with higher activity both in vitro and in vivo.
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| [14] |
Glutamate decarboxylase (GAD) catalyzes the decarboxylation of glutamate to CO2 and gamma-aminobutyrate (GABA). GAD is ubiquitous in prokaryotes and eukaryotes, but only plant GAD has been shown to bind calmodulin (CaM). Here, we assess the role of the GAD CaM-binding domain in vivo. Transgenic tobacco plants expressing a mutant petunia GAD lacking the CaM-binding domain (GADdeltaC plants) exhibit severe morphological abnormalities, such as short stems, in which cortex parenchyma cells fail to elongate, associated with extremely high GABA and low glutamate levels. The morphology of transgenic plants expressing the full-length GAD (GAD plants) is indistinguishable from that of wild-type (WT) plants. In WT and GAD plant extracts, GAD activity is inhibited by EGTA and by the CaM antagonist trifluoperazine, and is associated with a CaM-containing protein complex of approximately 500 kDa. In contrast, GADdeltaC plants lack normal GAD complexes, and GAD activity in their extracts is not affected by EGTA and trifluoperazine. We conclude that CaM binding to GAD is essential for the regulation of GABA and glutamate metabolism, and that regulation of GAD activity is necessary for normal plant development. This study is the first to demonstrate an in vivo function for CaM binding to a target protein in plants.
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| [15] |
GABA (gamma-aminobutyric acid), a non-protein amino acid, is a signaling factor in many organisms. In plants, GABA is known to accumulate under a variety of stresses. However, the consequence of GABA accumulation, especially in vegetative tissues, remains poorly understood. Moreover, gene expression changes as a consequence of GABA accumulation in plants are largely unknown. The pop2 mutant, which is defective in GABA catabolism and accumulates GABA, is a good model to examine the effects of GABA accumulation on plant development. Here, we show that the pop2 mutants have pollen tube elongation defects in the transmitting tract of pistils. Additionally, we observed growth inhibition of primary root and dark-grown hypocotyl, at least in part due to cell elongation defects, upon exposure to exogenous GABA. Microarray analysis of pop2-1 seedlings grown in GABA-supplemented medium revealed that 60% of genes whose expression decreased encode secreted proteins. Besides, functional classification of genes with decreased expression in the pop2-1 mutant showed that cell wall-related genes were significantly enriched in the microarray data set, consistent with the cell elongation defects observed in pop2 mutants. Our study identifies cell elongation defects caused by GABA accumulation in both reproductive and vegetative tissues. Additionally, our results show that genes that encode secreted and cell wall-related proteins may mediate some of the effects of GABA accumulation. The potential function of GABA as a growth control factor under stressful conditions is discussed.
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| [16] |
In plants, as in most eukaryotes, glutamate decarboxylase catalyses the synthesis of GABA. The Arabidopsis genome contains five glutamate decarboxylase genes and one of these genes (glutamate decarboxylase1; i.e. GAD1 ) is expressed specifically in roots. By isolating and analyzing three gad1 T-DNA insertion alleles, derived from two ecotypes, we investigated the potential role of GAD1 in GABA production. We also analyzed a promoter region of the GAD1 gene and show that it confers root-specific expression when fused to reporter genes. Phenotypic analysis of the gad1 insertion mutants revealed that GABA levels in roots were drastically reduced compared with those in the wild type. The roots of the wild type contained about sevenfold more GABA than roots of the mutants. Disruption of the GAD1 gene also prevented the accumulation of GABA in roots in response to heat stress. Our results show that the root-specific calcium/calmodulin-regulated GAD1 plays a major role in GABA synthesis in plants under normal growth conditions and in response to stress.
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| [17] |
gamma-Aminobutyric acid (GABA) is a non-proteinogenic amino acid that is found in uni- and multi-cellular organisms and is involved in many aspects of plant life cycle. GABA metabolism occurs by the action of evolutionary conserved enzymes that constitute the GABA shunt, bypassing two steps of the TCA cycle. The central position of GABA in the interface between plant carbon and nitrogen metabolism is well established. In parallel, there is evidence to support a role for GABA as a signaling molecule in plants. Here we cover some of the recent findings on GABA metabolism and signaling in plants and further suggest that the metabolic and signaling aspects of GABA may actually be inseparable.
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| [18] |
Much of the recent work on the gamma-aminobutyrate (GABA) shunt in plants has concentrated on stress/pest-associated and signalling roles. However, fifty years after the structural elucidation of the pathway, aspects of its regulation and even of its biological significance remain largely obscure. Here, we assess the importance of GABA metabolism in plants, reviewing relevant biological circumstances and taking advantage of high-throughput data accessibility and computational approaches. We discuss the premise that GABA metabolism plays a major role in carbon and nitrogen primary metabolism. We further evaluate technological developments that will likely allow us to address the quantitative importance of this shunt within the biological processes to which it contributes.
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Gamma-aminobutyric acid transaminase (GABA-T) catalyses the breakdown of GABA to succinic semialdehyde. In this report, three GABA-T isoforms were identified in the tomato (Solanum lycopersicum L.) plant. The deduced amino acid sequences of the three isoforms are highly similar over most of their coding regions with the exception of their N-terminal regions. Transient expression of the individual full-length GABA-T isoforms fused to the green fluorescent protein in tobacco suspension-cultured cells revealed their distinct subcellular localizations to the mitochondrion, plastid or cytosol, and that the specific targeting of the mitochondrion- and plastid-localized isoforms is mediated by their predicted N-terminal presequences. Removal of the N-terminal targeting presequences from the mitochondrion and plastid GABA-T isoforms yielded good recovery of the soluble recombinant proteins in Escherichia coli when they were co-expressed with the GroES/EL molecular chaperone complex. Activity assays indicated that all three recombinant isoforms possess both pyruvate- and glyoxylate-dependent GABA-T activities, although the mitochondrial enzyme has a specific activity that is significantly higher than that of its plastid and cytosolic counterparts. Finally, differential expression patterns of the three GABA-T isoforms in reproductive tissues, but not vegetative tissues, suggest unique roles for each enzyme in developmental processes. Overall, these findings, together with recent information about rice and pepper GABA-Ts, indicate that the subcellular distribution of GABA-T in the plant kingdom is highly variable.
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| [20] |
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| [21] |
gamma-aminobutyrate-transaminase (EC 2.6.1.19) catalyzes the first step of the catabolism of gamma-aminobutyric acid (GABA), a non-protein amino acid well-known to accumulate in plant in response to environmental stimuli. Recent studies reinforce more and more the role of its metabolism in carbon and/or nitrogen metabolisms and as a signalling molecule in developmental processes. Here we investigated the effects of inhibition of gamma-aminobutyrate-transaminase (GABA-T) in seedlings of Brassica napus, using vinyl-GABA (VGB) as a specific inhibitor of GABA-T to prevent enzyme activity. Root growth was reduced by 44% in VGB-treated seedlings but was less inhibited when VGB was associated with exogenous GABA and was not reduced with exogenous GABA alone. Measurements of GABA content in seedlings grown on VGB, GABA or VGB + GABA demonstrated that GABA level in root was not linked with the root length reduction, suggesting that GABA was not the sole component acting in root growth inhibition. Besides, metabolic profiling revealed that in root, VGB-treatment caused a twofold increase in content of almost all amino acids, except for alanine whose content was 19-fold higher than in control. In order to test the involvement of alanine accumulation on growth we studied the effects of exogenous alanine. High alanine content slightly reduced root growth suggesting that VGB-induced alanine accumulation was not responsible for root length reduction. We conclude that root growth inhibition in plants whose GABA catabolism was impaired could result at least partly from the disruption of the primary metabolism as a whole rather than direct effect of GABA on cellular growth process.
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The gamma-aminobutyrate (GABA) shunt is a metabolic pathway that bypasses two steps of the tricarboxylic acid cycle, and it is present in both prokaryotes and eukaryotes. In plants the pathway is composed of the calcium/calmodulin-regulated cytosolic enzyme glutamate decarboxylase and the mitochondrial enzymes GABA transaminase and succinic-semialdehyde dehydrogenase (SSADH). The activity of the GABA shunt in plants is rapidly enhanced in response to various biotic and abiotic stresses. However the physiological role of this pathway remains obscure. To elucidate its role in plants, we analyzed Arabidopsis T-DNA knockout mutants of SSADH, the ultimate enzyme of the pathway. Four alleles of the ssadh mutation were isolated, and these exhibited a similar phenotype. When exposed to white light (100 micromol of photons per m2 per s), they appear dwarfed with necrotic lesions. Detailed spectrum analysis revealed that UV-B has the most adverse effect on the mutant phenotype, whereas photosynthetic active range light has a very little effect. The ssadh mutants are also sensitive to heat, as they develop necrosis when submitted to such stress. Moreover, both UV and heat cause a rapid increase in the levels of hydrogen peroxide in the ssadh mutants, which is associated with enhanced cell death. Surprisingly, our study also shows that trichomes are hypersensitive to stresses in ssadh mutants. Our work establishes a role for the GABA shunt in preventing the accumulation of reactive oxygen intermediates and cell death, which appears to be essential for plant defense against environmental stress.
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gamma-Aminobutyric acid (GABA) influences plant growth, but little is known about how this metabolite regulates adventitious root (AR) development. Here, we investigate the effects of GABA on ARs using poplar lines overexpressing glutamate decarboxilase 2 (GAD2) and by treating poplar stem cuttings with exogenous GABA or vigabatrin (VGB; a specific GABA transaminase inhibitor). Endogenous GABA accumulation not only inhibited AR growth, but it also suppressed or delayed AR formation. Anatomical observations revealed that the GABA and VGB treatments resulted in a 1 d delay in the formation of AR primordia and the appearance of ARs. This delay coincided with changes in primary metabolism, including transient increases in hexose and amino acid levels. GABA-dependent changes in the expression of genes related to hormone synthesis and signalling, as well as analysis of hormone levels revealed that ethylene-dependent pathways were decreased at the earliest stage of AR formation. In contrast, auxin and abscisic acid were increased at 1-5 d as well as GA4 over a 5 d period of AR formation. These results demonstrate that GABA plays a crucial role in AR development. Evidence is presented demonstrating that GABA can interact with hormone-related pathways as well as carbon/nitrogen metabolism. These findings also elucidate the functions of GABA in plant development.
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| [25] |
Poplar 84K (Populus alba x P. tremula var. glandulosa) is a fast-growing poplar hybrid. Originated in South Korea, this hybrid has been extensively cultivated in northern China. Due to the economic and ecological importance of this hybrid and high transformability, we now report the de novo sequencing and assembly of a male individual of poplar 84K using PacBio and Hi-C technologies. The final reference nuclear genome (747.5 Mb) has a contig N50 size of 1.99 Mb and a scaffold N50 size of 19.6 Mb. Complete chloroplast and mitochondrial genomes were also assembled from the sequencing data. Based on similarities to the genomes of P. alba var. pyramidalis and P. tremula, we were able to identify two subgenomes, representing 356 Mb from P. alba (subgenome A) and 354 Mb from P. tremula var. glandulosa (subgenome G). The phased assembly allowed us to detect the transcriptional bias between the two subgenomes, and we found that the subgenome from P. tremula displayed dominant expression in both 84K and another widely used hybrid, P. tremula x P. alba. This high-quality poplar 84K genome will be a valuable resource for poplar breeding and for molecular biology studies.
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| [26] |
Programs to import, manage and align sequences and to analyse the properties of DNA, RNA and proteins are essential for every biological laboratory. This review describes two different freeware (BioEdit and pDRAW for MS Windows) and a commercial program (Sequencher for MS Windows and Apple MacOS). Bioedit and Sequencher offer functions such as sequence alignment and editing plus reading of sequence trace files. pDRAW is a very comfortable visualisation tool with a variety of analysis functions. While Sequencher impresses with a very user-friendly interface and easy-to-use tools, BioEdit offers the largest and most customisable variety of tools. The strength of pDRAW is drawing and analysis of single sequences for priming and restriction sites and virtual cloning. It has a database function for user-specific oligonucleotides and restriction enzymes.
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| [27] |
Uptake and translocation of cationic nutrients play essential roles in physiological processes including plant growth, nutrition, signal transduction, and development. Approximately 5% of the Arabidopsis genome appears to encode membrane transport proteins. These proteins are classified in 46 unique families containing approximately 880 members. In addition, several hundred putative transporters have not yet been assigned to families. In this paper, we have analyzed the phylogenetic relationships of over 150 cation transport proteins. This analysis has focused on cation transporter gene families for which initial characterizations have been achieved for individual members, including potassium transporters and channels, sodium transporters, calcium antiporters, cyclic nucleotide-gated channels, cation diffusion facilitator proteins, natural resistance-associated macrophage proteins (NRAMP), and Zn-regulated transporter Fe-regulated transporter-like proteins. Phylogenetic trees of each family define the evolutionary relationships of the members to each other. These families contain numerous members, indicating diverse functions in vivo. Closely related isoforms and separate subfamilies exist within many of these gene families, indicating possible redundancies and specialized functions. To facilitate their further study, the PlantsT database (http://plantst.sdsc.edu) has been created that includes alignments of the analyzed cation transporters and their chromosomal locations.
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| [28] |
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| [29] |
The ExPASy (the Expert Protein Analysis System) World Wide Web server (http://www.expasy.org), is provided as a service to the life science community by a multidisciplinary team at the Swiss Institute of Bioinformatics (SIB). It provides access to a variety of databases and analytical tools dedicated to proteins and proteomics. ExPASy databases include SWISS-PROT and TrEMBL, SWISS-2DPAGE, PROSITE, ENZYME and the SWISS-MODEL repository. Analysis tools are available for specific tasks relevant to proteomics, similarity searches, pattern and profile searches, post-translational modification prediction, topology prediction, primary, secondary and tertiary structure analysis and sequence alignment. These databases and tools are tightly interlinked: a special emphasis is placed on integration of database entries with related resources developed at the SIB and elsewhere, and the proteomics tools have been designed to read the annotations in SWISS-PROT in order to enhance their predictions. ExPASy started to operate in 1993, as the first WWW server in the field of life sciences. In addition to the main site in Switzerland, seven mirror sites in different continents currently serve the user community.
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| [30] |
任丽, 董京祥, 杨洋 , 等. 白桦BpTCP7基因启动子的克隆及表达分析[J]. 南京林业大学学报(自然科学版), 2019,43(1):32-38.
|
| [31] |
|
| [32] |
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| [33] |
Promoters play key roles in conferring temporal, spatial, chemical, developmental, or environmental regulation of gene expression. Promoters that are subject to specific regulations are useful for manipulating foreign gene expression in plant cells, tissues, or organs with desirable patterns and under controlled conditions, and have been important for both basic research and applications in agriculture biotechnology. Recent advances in genomics technologies have greatly facilitated identification and study of promoters in a genome scale with high efficiency. Previously we have generated a large T-DNA tagged rice mutant library (TRIM), in which the T-DNA was designed with a gene/promoter trap system, by placing a promoter-less GUS gene next to the right border of T-DNA. GUS activity screens of this library offer in situ and in planta identifications and analyses of promoter activities in their native configurations in the rice genome. In the present study, we systematically performed GUS activity screens of the rice mutant library for genes/promoters constitutively, differentially, or specifically active in vegetative and reproductive tissues. More than 8,200 lines have been screened, and 11% and 22% of them displayed GUS staining in vegetative tissues and in flowers, respectively. Among the vegetative tissue active promoters, the ratio of leaf active versus root active is about 1.6. Interestingly, all the flower active promoters are anther active, but with varied activities in different flower tissues. To identify tissue specific ABA/stress up-regulated promoters, we compared microarray data of ABA/stress induced genes with those of tissue-specific expression determined by promoter trap GUS staining. Following this approach, we showed that the peroxidase 1 gene promoter was ABA up-regulated by 4 fold within 1 day of exposure to ABA and its expression is lateral root specific. We suggest that this be an easy bioinformatics approach in identifying tissue/cell type specific promoters that are up-regulated by hormones or other factors.
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| [34] |
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| [35] |
BACKGROUND: Glutamate decarboxylase (GAD), as a key enzyme in the gamma -aminobutyric acid (GABA) shunt, catalyzes the decarboxylation of L-glutamate to form GABA. This pathway has attracted much interest because of its roles in carbon and nitrogen metabolism, stress responses, and signaling in higher plants. The aim of this study was to isolate and characterize genes encoding GADs from Caragana intermedia, an important nitrogen-fixing leguminous shrub. METHODS: Two full-length cDNAs encoding GADs (designated as CiGAD1 and CiGAD2) were isolated and characterized. Multiple alignment and phylogenetic analyses were conducted to evaluate their structures and identities to each other and to homologs in other plants. Tissue expression analyses were conducted to evaluate their transcriptional responses to stress (NaCl, ZnSO4, CdCl2, high/low temperature, and dehydration) and exogenous abscisic acid. RESULTS: The CiGADs contained the conserved PLP domain and calmodulin (CaM)-binding domain in the C-terminal region. The phylogenetic analysis showed that they were more closely related to the GADs of soybean, another legume, than to GADs of other model plants. According to Southern blotting analysis, CiGAD1 had one copy and CiGAD2-related genes were present as two copies in C. intermedia. In the tissue expression analyses, there were much higher transcript levels of CiGAD2 than CiGAD1 in bark, suggesting that CiGAD2 might play a role in secondary growth of woody plants. Several stress treatments (NaCl, ZnSO4, CdCl2, high/low temperature, and dehydration) significantly increased the transcript levels of both CiGADs, except for CiGAD2 under Cd stress. The CiGAD1 transcript levels strongly increased in response to Zn stress (74.3-fold increase in roots) and heat stress (218.1-fold increase in leaves). The transcript levels of both CiGADs significantly increased as GABA accumulated during a 24-h salt treatment. Abscisic acid was involved in regulating the expression of these two CiGADs under salt stress. DISCUSSION: This study showed that two CiGADs cloned from C. intermedia are closely related to homologs in another legume, soybean. CiGAD2 expression was much higher than that of CiGAD1 in bark, indicating that CiGAD2 might participate in the process of secondary growth in woody plants. Multiple stresses, interestingly, showed that Zn and heat stresses had the strongest effects on CiGAD1 expression, suggesting that CiGAD1 plays important roles in the responses to Zn and heat stresses. Additionally, these two genes might be involved in ABA dependent pathway during stress. This result provides important information about the role of GADs in woody plants' responses to environmental stresses.
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| [36] |
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| [37] |
Glutamate decarboxylase (GAD) catalyzes the conversion of L-glutamate to gamma-aminobutyric acid (GABA). A full-length cDNA encoding GAD (designated as PgGAD) was isolated and characterized from the root of Panax ginseng C. A. Meyer. The length cDNA of PgGAD was 1881 bp and contained a 1491 bp open reading frame (ORF) encoding a glutamate decarboxylase protein of 496 amino acids, possessing a Ser-X-X-Lys active site, which belongs to the GAD group. The deduced amino acid sequence of the PgGAD was classified in the plant GAD family and has 76-85% high similarity with other plants as like petunia, Arabidopsis, tomato. Secondary structure of PgGAD was predicted by using SOPMA software program. Southern blot analysis of genomic DNA suggests that, there is more than one copy of the PgGAD gene. The organ specific gene expression pattern also studied in P. ginseng seedlings, in which the stem showed elevated expression than root, leaf, bud and rhizomes. Along with this, we also confirmed the gene expression of PgGAD under various abiotic stresses like temperature stress, osmotic stress, anoxia, oxidative stress, and mechanical damage. Temporal analysis of gene expression except exposure of oxidative stress revealed an enhanced expression after each stresses. The enzyme activity of PgGAD was stimulated to 2-fold under cold stress.
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| [38] |
单雪萌, 杨克彬, 史晶晶 , 等. 毛竹GeBP转录因子家族的全基因组鉴定和表达分析[J]. 南京林业大学学报(自然科学版), 2020,44(3):41-48.
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