Developmental and Pathogen-Induced Activation of the Arabidopsis Acidic Chitinase Promoter (original) (raw)
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Developmental and Pathogen-lnduced Activation of the Arabidopsis Acidic Chitinase Promoter
1991
Expression of the Arabidopsis acidic chitinase promoter was investigated during plant development and in response to inoculation with fungal pathogens. A chimeric gene composed of 1129 bp of 5' upstream sequence from the acidic chitinase gene was fused to the 8-glucuronidase (GUS) coding region and used to transform Arabidopsis and tomato. Promoter activity was monitored by histochemical and quantitative assays of GUS activity. In healthy transgenic plants, the acidic chitinase promoter activity was restricted to roots, leaf vascular tissue, hydathodes, guard cells, and anthers, whereas GUS expression was induced in mesophyll cells surrounding lesions caused by Rhizoctonia solani infection of transgenic Arabidopsis. In transgenic tomato plants, GUS expression was induced around necrotic lesions caused by Alternaria solani and Phytophthora infestam. Expression of the acidic chitinase promoter-GUS transgene was weakly induced by infiltrating leaves with salicylic acid. Analysis of a series of 5' deletions of the acidic chitinase promoter in Arabidopsis indicated that the proximal 192 bp from the transcription initiation site was sufficient to establish both the constitutive and induced pattern of expression. Elements further upstream were involved in quantitative expression of the gene. The location of a negative regulatory element was indicated between -384 and -590 and positive regulatory elements between -1 129 and -590.
Activation of a Bean Chitinase Promoter in Transgenic Tobacco Plants by Phytopathogenic Fungi
THE PLANT CELL ONLINE, 1990
The temporal and spatial expression of a bean chitinase promoter has been investigated in response to fungal attack. Analysis of transgenic tobacco plants containing a chimeric gene composed of a 1.7-kilobase fragment carrying the chitinase 5B gene promoter fused to the coding region of the gus A gene indicated that the chitinase promoter is activated during attack by the fungal pathogens Botrytis cinerea, Rhizoctonia solani, and Sclerotium rolfsii. Although induction of 8-glucuronidase activity was observed in tissues that had not been exposed to these phytopathogens, the greatest induction occurred in and around the site of fungal infection. The increase in 8-glucuronidase activity closely paralleled the increase in endogenous tobacco chitinase activity produced in response to fungal infection. Thus, the chitinase 5B-gus A fusion gene may be used to analyze the cellular and molecular details of the activation of the host defense system during pathogen attack.
Expression of bacterial chitinase protein in tobacco leaves using two photosynthetic gene promoters
MGG Molecular & General Genetics, 1988
A bacterial chitinase gene from Serratia marcescens (chiA) was fused to (i) a promoter of the ribulose bisphosphate carboxylase small subunit (rbcS) gene and (ii) two different chlorophyll a/b binding protein (cab) gene promoters from petunia. The resulting constructions were introduced into Agrobaeterium Ti plasmid-based plant cell transformation vectors and used to generate multiple independent transgenic tobacco plants. ChiA mRNA and protein levels were measured in these plants. On average, the rbcS/ehiA fusion gave rise to threefold more ehiA mRNA than either cab/chiA fusion. We investigated the influence of sequences around the translational initiation ATG codon on the level of ChiA protein. The rbcS/chiA and cab/chiA fusions in which the sequence in the vicinity of the translational initiation codon is ACC ATGGC gave rise to transformants with higher levels of ChiA protein than those carrying a eab/chiA fusion with the sequence CAT ATGCG in the same region. This difference in translational efficiency is consistent with previous findings on preferred sequences in this region of the mRNA. In those transformants showing the highest level of ChiA expression, ChiA protein accumulated to about 0.25% of total soluble leaf protein. These plants contained significantly higher chitinase enzymatic activity than control plants.
THE PLANT CELL ONLINE, 1989
Expression of at least two genes from bean encoding the defense-related protein chitinase has been shown previously to be transcriptionally regulated by the phytohormone ethylene. We have determined the complete nucleotide sequence of one of these genes, the CH5B gene, which resides on a 4.7-kilobase fragment of bean genomic DNA. The structural gene consists of a single open reading frame and encodes the 301 amino acids of the mature protein and a 26-amino acid signal peptide. The CH5B gene has been introduced into tobacco plants using Agrobacterium Ti-plasmid vectors. Little or no expression of the bean gene was observed when transgenic tobacco plants were grown in air; however, exposure of these plants to an atmosphere containing 50 parts per million ethylene resulted in an approximately 20-fold to 50-fold increase in the leve1 of the bean chitinase mRNA. Ethylenedependent expression of a chimeric gene consisting of 1.6 kilobases of 5'-flanking DNA derived from the CH5B gene fused to the coding sequence of B-glucuronidase indicates that this region of the CH5B gene is sufficient for ethylene-regulated expression. Deletion analysis of the CH5B promoter region has allowed us to localize these DNA sequences to within a 228-base pair region situated between-422 and-195 upstream of the transcriptional start site. This region is characterized by two short DNA sequences that are exactly conserved in a second ethyleneregulated bean chitinase gene.
Plant Biotechnology Reports, 2013
Fusarium wilt is a soil-borne disease causing substantial yield losses in various crops and vegetables. We have previously reported the synthetic chitinase (NIC) gene (1.2 kb), in which codon usage of fungus, replaced with that of plant, conferred resistance against Botrytis cinerea. In this study, the NIC or GUS gene was linked to two rootspecific promoters, LjNRT2 or AtNRT2.1 (nitrate transporter 2), derived from Lotus japonica and Arabidopsis thaliana, respectively. Transgenic tobacco lines expressing LjNRT2-GUS and LjNRT2-NIC, and tomato lines expressing AtNRT2.1-NIC, were produced by Agrobacteriummediated transformation. GUS histochemical staining was observed in vascular regions of the roots but was conspicuously absent in the leaves of transgenic plants. Western blot analysis showed the production of NIC proteins in the roots but not in the leaves of transgenic tobacco and tomato lines. These results indicate that LjNRT2 and At-NRT2.1 promoters expressed transgenes in a root-specific manner. When in vitro whole plant resistance assay against Fusarium oxysporum was conducted, transgenic plants showed increased levels of resistance compared to nontransgenic plants. Antifungal activities of the root extract against spore germination of F. oxysporum showed lower CFU (colony-forming unit) than those of the leaf extract. Root colonization assay against F. oxysporum showed much lower CFU values in the roots of transgenic plants than in those of non-transgenic plants. These results suggest that NIC gene triggered by the root-specific promoters successfully expressed only in the roots and conferred increased levels of resistance against the root pathogen, F. oxysporum.
FEBS Letters, 1997
Endochitinases are widely distributed among higher plants, including a number of important crop species. They are generally considered to be involved in plant defence against potential pathogens. We have cloned a class IV chitinase gene {AtchitIV) from Arabidopsis thaliana. Southern blot analysis allowed the detection of two cross-hybridising genes in the A. thaliana genome. AtchitIV transcripts are detected in seedpods, but not in roots, inflorescence stems, leaves and flowers of healthy plants. The transcripts accumulated very rapidly in leaves after inoculation with Xanthomonas campestris. Maximum mRNA accumulation was reached one hour after infection and decreased to very low levels 72 hours after induction. This result suggests an involvement of AtchitIV in the initial events of the hypersensitive reaction. Nevertheless, A. thaliana plants transformed with the gus gene under the control of a class IV chitinase bean promoter, showed GUS activity in seed embryos. These data, together with the constitutive expression of the endogenous gene in the seedpods, points to additional physiological roles for this protein.
Effect of chitinase antisense RNA expression on disease susceptibility of Arabidopsis plants
Plant Molecular Biology, 1994
Chitinases accumulate in higher plants upon pathogen attack are capable of hydrolyzing chitin-containing fungal cell walls and are thus implicated as part of the plant defense response to fungal pathogens. To evaluate the relative role of the predominate chitinase (class I, basic enzyme) ofArabidopsis thaliana in disease resistance, transgenic Arabidopsis plants were generated that expressed antisense RNA to the class I chitinase. Young plants or young leaves of some plants expressing antisense RNA had < 10~o of the chitinase levels of control plants. In the oldest leaves of these antisense plants, chitinase levels rose to 37-90~o of the chitinase levels relative to vector control plants, most likely because of accumulation and storage of the enzyme in vacuoles. The rate of infection by the fungal pathogen Botrytis cinerea was measured in detached leaves containing 7-15 ~o of the chitinase levels of control plants prior to inoculation. Antisense RNA was not effective in suppressing induced chitinase expression upon infection as chitinase levels increased in antisense leaves to 47~o of levels in control leaves within 24 hours after inoculation. Leaves from antisense plants became diseased at a slightly faster rate than leaves from control plants, but differences were not significant due to high variability. Although the tendency to increased susceptibility in antisense plants suggests that chitinases may slow the growth of invading fungal pathogens, the overall contribution of chitinase to the inducible defense reponses in Arabidopsis remains unclear.
Biotechnology reports (Amsterdam, Netherlands), 2017
Chitinases are the hydrolytic enzymes which belong to the pathogenesis-related (PR) protein family and play an important role not only in plant defense but also in various abiotic stresses. However, only a limited number of chitinase genes have been characterised in B. juncea. In this study, we have characterised B. juncea class IV chitinase gene (accession no EF586206) in response to fungal infection, salicylic acid (SA), jasmonic acid (JA) treatments and wounding. Gene expression studies revealed that the transcript levels of Bjchitinase (BjChp) gene increases significantly both in local and distal tissues after Alternaria infection. Bjchitinase gene was also induced by jasmonic acid and wounding but moderately by salicylic acid. A 2.5 kb class IV chitinase promoter of this gene was isolated from B. juncea by Genome walking (accession no KF055403.1). In-silico analysis of this promoter revealed a number of conserved cis-regulatory elements related to defense, wounding and signalli...
Molecular Plant-microbe Interactions, 2004
The Arabidopsis glutathione S-transferase GSTF8 promoter directs root-specific responses to stress. In this study, the response of this promoter to plant infection with Rhizoctonia solani was investigated using a luciferase reporter system. Arabidopsis seedlings harboring the GSTF8:luciferase construct were monitored in vivo for bioluminescence following infection with R. solani. Although the reporter gene was induced in infected roots, the response differed markedly between R. solani strains and was not observed with aggressive strains that caused death of the seedlings. The three strains tested in detail progressed through typical stages of infection, but ZG1-1 induced the GSTF8 promoter in most seedlings, ZG3 induced it in approximately 25% of seedlings, and ZG5 caused little response. Induction of specific root segments occurred early in the infection process in root regions with very limited mycelium visible. In root segments with substantial mycelium, GSTF8 promoter activity no longer was observed. Induction by ZG1-1 also was observed in plants harboring a tetramer of the ocs element from the GSTF8 promoter, suggesting that this element helps mediate the response. Crossing GSTF8:luciferase plants with plants harboring an Nah-G construct that degrades salicylic acid did not abolish the response, indicating that the GSTF8 promoter response to R. solani may be mediated by signals other than salicylic acid.