Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The phenylpropanoid pathway intermediate p-coumaric acid (4-CA) stimulates expression of the bean (Phaseolus vulgaris L.) chalcone synthase (malonyl-CoA:4-coumaroyl-CoA, EC 2.3.1.74) chs15 gene promoter in electroporated protoplasts of alfalfa (Medicago sativa L.). We have analyzed the effects of 5' deletions, mutations, and competition with promoter sequences in trans on the expression of a chs15 promoter-chloramphenicol acetyltransferase gene fusion in elicited alfalfa protoplasts. Two distinct sequence elements, the H-box (consensus CCTACC(N)7CT) and the G-box (CACGTG), are required for stimulation of the chs15 promoter by 4-CA. Furthermore, a 38-base-pair chs15 promoter sequence containing both cis elements conferred responsiveness to 4-CA on the cauliflower mosaic virus 35S minimal promoter. The H-box and G-box in combination establish the complex developmental pattern of chs15 expression and are also involved in stress induction. Hence, potential internal pathway regulation through feed-forward stimulation by 4-CA operates by modulation of the signal pathways for developmental and environmental regulation.
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PMID:Combination of H-box [CCTACC(N)7CT] and G-box (CACGTG) cis elements is necessary for feed-forward stimulation of a chalcone synthase promoter by the phenylpropanoid-pathway intermediate p-coumaric acid. 140 28

A chimeric gene construct containing a bean chalcone synthase (CHS) promoter fused to the chloramphenicol acetyltransferase (CAT) reporter gene was strongly expressed when electroporated into alfalfa protoplasts that were then exposed to a fungal elicitor. Low concentrations (5 x 10(-6) to 10(-4) M) of exogenously applied trans-cinnamic acid (CA), the first intermediate of the phenylpropanoid pathway, slightly stimulated elicitor-induced CAT expression, whereas high concentrations (greater than 10(-4) M) severely reduced expression to below the levels observed in the absence of elicitor. In contrast, trans-p-coumaric acid (4-CA, the second intermediate in the pathway) stimulated expression from the CHS promoter up to 4.5-fold at 5 x 10(-4) M. Expression of CAT driven by the promoters of other elicitor-inducible defense response genes was not markedly affected by CA or 4-CA. Stimulation of CHS promoter expression by low concentrations of CA and 4-CA was completely abolished by 5' deletion to position -130, but not -174. When the -180 to -130 region of the CHS15 promoter was coelectroporated into elicited protoplasts on a separate plasmid along with the intact -326 CHS-CAT construct, the decreased CAT expression as a function of CA or 4-CA concentration was consistent with the coelectroporated sequence competing in trans with the intact promoter for the binding of a factor(s) involved in the up regulation of CHS transcription by 4-CA and low concentrations of CA. Our data support the hypothesis that phenylpropanoid compounds may act as natural and specific regulators of plant gene expression and define the location of a cis-acting element in the CHS15 promoter involved in the induction by phenylpropanoid pathway intermediates.
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PMID:Phenylpropanoid pathway intermediates regulate transient expression of a chalcone synthase gene promoter. 182 Aug 22

A chimeric gene consisting of a bean (Phaseolus vulgaris L.) chalcone synthase (CHS) promoter fused to a bacterial chloramphenicol acetyltransferase (CAT) reporter gene was strongly expressed, and further induced by fungal elicitor, when electroporated into alfalfa (Medicago sativa L.) suspension cell protoplasts. Functional analysis of 5' deletions of the CHS promoter-CAT construct in these protoplasts indicated that the region between -326 and -130 contained both activator and silencer elements. Co-electroporation experiments confirmed that these cis-acting elements were binding sites for functionally active trans factors. In vitro DNase I footprinting revealed four potential binding sites for alfalfa suspension cell nuclear proteins between positions -326 and -130 of the CHS promoter. These sites mapped to regions shown to contain functional cis-acting elements on the basis of the deletion analysis. Three of these sites mapped to previously identified binding sites for bean nuclear proteins. Competition gel retardation analysis using oligonucleotide probes containing binding site sequences revealed sequence-specific binding of alfalfa nuclear proteins to an AT-rich element and a putative GT-1 factor consensus binding sequence. Our results define cis elements and their cognate trans factors functionally active in determining the quantitative expression of a defense response gene in a heterologous transient expression system.
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PMID:Stress responses in alfalfa (Medicago sativa L.). 8. Cis-elements and trans-acting factors for the quantitative expression of a bean chalcone synthase gene promoter in electroporated alfalfa protoplasts. 185 70

Pycnospore germination fluid of Mycosphaerella pinodes, a fungus pathogenic on pea, contains both an elicitor and a suppressor of the accumulation of pisatin, a major phytoalexin of pea. Transcription of the genes encoding key enzymes in the biosynthesis of pisatin, namely PAL (a gene encoding phenylalanine ammonia-lyase) and CHS (a gene encoding chalcone synthase), was shown to be activated upon the treatment of pea epicotyl tissues with the fungal elicitor and suppressed upon treatment with the fungal suppressor. To investigate the mechanisms underlying activation and suppression of plant defense genes by signal molecules secreted by a fungal pathogen and other stresses, such as ultraviolet (UV) light, we constructed chimeric genes composed of the 5'-flanking regions of two members of the PSPAL family (the genes encoding phenylalanine ammonia-lyase in Pisum sativum) fused to a bacterial gene for chloramphenicol acetyltransferase. Then, the cis-regulatory elements necessary for elicitor-mediated activation and suppressor-mediated suppression were examined in pea protoplasts. Functional analysis of 5' nested deletions of PSPAL1 and PSPAL2 suggested that an enhancer-like element is located in the TATA-distal region (-2,196 to -406) in PSPAL2. A cis-acting element(s) responsible for elicitor-mediated activation was found in the TATA-proximal region (-340 to -95 in PSPAL1; -406 to -158 in PSPAL2), in which the consensus sequence motifs known as box 1, box 2 and box 4 [Yamada et al. (1992) Plant Cell Physiol. 33: 715, Lois et al. (1989) EMBO J. 8: 1641] were present in close proximity. Furthermore, both promoters were activated by UV light but were partially suppressed in response to the fungal suppressor.
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PMID:Functional analysis of the promoters of phenylalanine ammonia-lyase genes in pea. 798 63

To investigate the mechanisms underlying activation of plant defenses against microbial attack we have studied elicitor regulation of a chimeric gene comprising the 5' flanking region of a defense gene encoding the phytoalexin biosynthetic enzyme chalcone synthase fused to a bacterial chloramphenicol acetyltransferase gene. Glutathione or fungal elicitor caused a rapid, marked but transient expression of the chimeric gene electroporated into soybean protoplasts. The response closely resembled that of endogenous chalcone synthase genes in suspension cultured cells. Functional analysis of 5' deletions suggests that promoter activity is determined by an elicitor-regulated activator located between the "TATA box" and nucleotide position -173 and an upstream silencer between -173 and -326. These cis-acting elements function in the transduction of the elicitation signal to initiate elaboration of an inducible defense response.
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PMID:Glutathione and fungal elicitor regulation of a plant defense gene promoter in electroporated protoplasts. 1659 81

Protoplasts derived from cell suspensions of alfalfa (Medicago sativa L.) responded to treatment with fungal elicitor (FE) by an increase in endogenous chalcone synthase (CHS) activity but were unresponsive to reduced glutathione (GSH). Preexposure of protoplasts to polyethylene glycol and electroporation resulted in strong responsiveness to GSH but little change in responsiveness to FE. Protoplasts from suspension cultures which had been subcultured more than 12 times lost responsiveness to GSH, but not FE, as assessed by measuring expression of a chimeric gene containing a bean CHS promoter linked to a bacterial chloramphenicol acetyltransferase (CAT) reporter gene. In protoplasts in which putative cis-acting CHS promoter sequences had been coelectroporated in trans with the intact CHS promoter-CAT construct, the extent of CAT expression depended upon the elicitor used (FE or GSH), the age (number of times subcultured) of the cells from which the protoplasts were isolated, and the nature of the coelectroporated CHS promoter sequence. For example, a region of the CHS promoter from -326 to -141 behaved as a trans-activator when coelectroporated with the CAT construct into unelicited protoplasts isolated from newly initiated cell suspensions, but the same region acted as a trans-silencer in the same protoplasts in the presence of FE. This silencer activity was much reduced in GSH-treated protoplasts. The results suggest that there are differences in the signal transduction pathways for elicitation of CHS transcription by FE and GSH, which involve previously indentified cis-elements in the CHS promoter.
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PMID:Stress Responses in Alfalfa (Medicago sativa L.): VI. Differential Responsiveness of Chalcone Synthase Induction to Fungal Elicitor or Glutathione in Electroporated Protoplasts. 1666 19

We have investigated conditions for the uptake and expression of chimeric genes in protoplasts of alfalfa (Medicago sativa L.). Constructs containing the bacterial reporter gene chloramphenicol acetyltransferase (CAT) under the control of either the cauliflower mosaic virus 35S promoter or a bean chalcone synthase (CHS) promoter were introduced into protoplasts by electroporation in the presence of polyethyleneglycol. The extent of expression in the absence of added inducers depended on the conditions for isolation, electroporation and subsequent culture of the protoplasts. Expression of the CHS promoter construct was increased on exposure of the protoplasts to a fungal elicitor or reduced glutathione. The relative levels of induced expression in relation to either basal expression or the type of elicitor used depended on the age of the suspension cultures from which the protoplasts were isolated. Electroporation of protoplasts with a construct from which bean CHS antisense transcripts were synthesized under the control of the 35S promoter resulted in the inhibition of appearance of elicitor-induced endogenous alfalfa CHS activity. The suitability of the alfalfa protoplast system for analysis and potential identification of defense response genes is discussed.
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PMID:Stress responses in alfalfa (Medicago sativa L.) IV. Expression of defense gene constructs in electroporated suspension cell protoplasts. 2422 76