Functional Analysis of DNA Sequences Responsible for Ethylene Regulation of a Bean Chitinase Gene in Transgenic Tobacco

Broglie, Karen; Biddle, Phyllis; Cressman, Robert; Broglie, Richard

doi:10.2307/3868947

Cited by 37 publications

(62 citation statements)

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Section: Discussionmentioning

confidence: 99%

“…Studies of this type with a bean chitinase gene are already in progress [4] and it will be of interest to compare the 5' regulatory regions of the co-induced 1,3-fl-D-glucanase and chitinase genes. The work on bean chitinase genes has been performed in cultivar Saxa [4], which we have now shown to contain a single 1,3-fl-D-glucanase gene. Furthermore, the apparent up-regulation of elicitorinduced glucanase transcript appearance by cinnamic acid warrants further study, as recent results from our laboratory have demonstrated the presence of specific cis-acfing elements conditioning the responsiveness of a bean chalcone synthase gene promoter to this phenylpropanoid pathway intermediate (A.D. Choudhary, G. Loake, M.J. Harrison, M. Mavandad, C.J.…”

Section: Discussionmentioning

confidence: 99%

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cDNA cloning and characterization of a putative 1,3-?-D-glucanase transcript induced by fungal elicitor in bean cell suspension cultures

1991

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Synthetic oligonucleotides based on similarity between tobacco 1,3-beta-D-glucanase and barley 1,3-1,4-beta-D-glucanase were used to prime the synthesis and amplification of a 162 bp bean (Phaseolus vulgaris L.) beta-glucanase cDNA by the polymerase chain reaction (PCR). The PCR product was used to isolate a near full-length beta-glucanase cDNA corresponding to an approximately 1400 bp full-length transcript, from a library containing cDNA sequences complementary to mRNA from fungal elicitor-treated bean cells. At the amino acid level, the bean beta-glucanase cDNA was 59% similar to tobacco 1,3-beta-D-glucanase, 46% similar to barley 1,3-beta-D-glucanase and 46% similar to barley 1,3-1,4-beta-D-glucanase. At the nucleotide level, the similarities were 65, 50 and 53% respectively. The beta-glucanase appeared to be encoded by a single gene with similar genomic organization in bean cultivars Canadian Wonder, Imuna and Saxa. On the basis of predicted Mr, isoelectric point, sequence similarity, and comparisons of rate of transcript appearance with induced enzyme activity, it was concluded that the cDNA encodes the basic bean endo-1,3-beta-D-glucanase. Glucanase transcripts were induced, from very low basal levels, with similar kinetics to chitinase transcripts in elicitor-treated bean cell suspension cultures.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

cDNA cloning and characterization of a putative 1,3-?-D-glucanase transcript induced by fungal elicitor in bean cell suspension cultures

1991

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“…The effect of ethylene on fruit ripening and other developmental processes is due, at least in part, to changes in the transcription ofspecific genes (3)(4)(5). To understand how gene expression is regulated by ethylene during fruit ripening, a number of ethylene-responsive genes have been cloned and studied in this laboratory (6).…”

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confidence: 99%

Identification of an ethylene-responsive region in the promoter of a fruit ripening gene.

Montgomery

Goldman

Deikman

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

209

155

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Transcription of the E4 gene is controlled by an increase in ethylene concentration during tomato fruit ripening. To investigate the molecular basis for ethylene regulation, we have examined the E4 promoter to identify cis elements and trans-acting factors that are involved in E4 gene expression. In transgenic tomato plants a chimeric gene construct containing a 1.4-kilobase E4 promoter fused to a f-glucuronidase reporter gene Is rapidly induced by ethylene in ripening fruit. Deletion of E4 promoter sequences to 193 base pairs reduces the level of GUS activity but does not affect ethylene induction. Transient expression of E4 promoterluciferase chimeric gene constructs containing various deletions, introduced into tomato fruit pericarp by particle bombardment, indicates that a positive ethylene-responsive region is localized between nucleotides -161 and -85 relative to the ascription start site. DNase I footprint analysis shows that a nuclear factor in unripe fruit interacts specifically with sequences in this element, from -142 to -110, which are required for the ethylene response. The DNase I footprint of this factor is reduced in ethylene-treated unripe fruit and undetectable in ripe fruit. Based on the correlation of a nuclear factor binding site with promoter sequences required for ethylene induction, we propose that this in vitro DNA-binding activity may represent a factor that is involved in ethyleneregulated E4 gene expression.The plant hormone ethylene influences numerous aspects of plant growth and development (1), including the ripening of climacteric fruits such as tomato. The role of ethylene in tomato fruit ripening has been demonstrated unequivocally by inhibiting endogenous ethylene production through the expression of antisense RNAs that block the accumulation of 1-aminocyclopropane-l-carboxylate (ACC) synthase, which catalyzes the rate-limiting step in ethylene biosynthesis (2). Fruit from plants transformed with the ACC synthase antisense gene evolve only very low levels of ethylene and do not ripen.The effect of ethylene on fruit ripening and other developmental processes is due, at least in part, to changes in the transcription ofspecific genes (3-5). To understand how gene

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“…Interestingly, the sequence between -7 1 4 and -5 6 7 is conserved in the corresponding region of the other tobacco basic chitinase gene CHN17. This region contains 12 bp direct repeat sequences including a 6-basepair GC-rich sequence (GCCGCC), which is also present in the 5' flanking regions of the bean [6] and the tomato [9] chitinase genes. In the bean chitinase gene the region containing this GC-rich sequence is responsible for ethylene-regulated expression.…”

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confidence: 99%

Gene structure and expression of a tobacco endochitinase gene in suspension-cultured tobacco cells

1991

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We have isolated and characterized the genomic clone 2CHN50 corresponding to tobacco basic endochitinase (E.C.3.2.1.14). DNA sequence and blotting analysis reveal that the coding sequence of the gene present on 2CHN50 is identical to that of the cDNA clone pCHN50 and, moreover, the CHN50 gene has its origin in the progenitor of tobacco, Nicotiana sylvestris. Tobacco basic chitinases are encoded by a small gene family that consists of at least two members, the CHN50 gene and a closely related CHN 17 gene which was characterized previously. By northern blot analysis, it is shown that the CHN50 gene is highly expressed in suspension-cultured tobacco cells and the mRNA accumulates at late logarithmic growth phase. To identify cis-DNA elements involved in the expression of the CHN50 gene in suspensioncultured cells, the chimeric gene consisting of 1.1 kb CHN50 5' upstream region fused to the coding sequence of fl-glucuronidase (GUS) was introduced by electroporation into protoplasts isolated from suspension-cultured tobacco cells. Transient GUS activity was found to be dependent on the growth phase of the cultured cells, from which protoplasts had been prepared. Functional analysis of 5' deletions suggests that the distal region between -788 and -345 contains sequences that potentiate the high-level expression in tobacco protoplasts and the region ( -68 to -4 7 ) proximal to the TATA box functions as a putative silencer.

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Functional Analysis of DNA Sequences Responsible for Ethylene Regulation of a Bean Chitinase Gene in Transgenic Tobacco

Cited by 37 publications

References 0 publications

cDNA cloning and characterization of a putative 1,3-?-D-glucanase transcript induced by fungal elicitor in bean cell suspension cultures

cDNA cloning and characterization of a putative 1,3-?-D-glucanase transcript induced by fungal elicitor in bean cell suspension cultures

Identification of an ethylene-responsive region in the promoter of a fruit ripening gene.

Gene structure and expression of a tobacco endochitinase gene in suspension-cultured tobacco cells

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