Functional analysis of a TGA factor-binding site located in the promoter region controlling salicylic acid-induced NIMIN-1 expression in Arabidopsis.
TGA factors play a key role in plant defense by binding to the promoter region of defense genes, inducing expression. Salicylic acid (SA) induces the expression of the gene encoding NIMIN-1, which interacts with NPR1/NIM1, a key regulator of systemic acquired resistance. We investigated whether the TGA2-binding motif TGACG located upstream of the NIMIN-1 gene is necessary for SA induction of NIMIN-1 expression. A mutated version of the NIMIN-1 promoter was created by site-directed mutagenesis. We generated T-DNA constructs in which native NIMIN-1 and mutated promoters were fused to green fluorescent protein and beta-glucuronidase reporters. We produced transgenic Arabidopsis plants and observed NIMIN-1 promoter-driven green fluorescent protein expression in the roots, petiole and leaves. Constructs were agroinfiltrated into the leaves for transient quantitative assays of gene expression. Using quantitative real-time RT-PCR, we characterized the normal gene response to SA and compared it to the response of the mutant version of the NIMIN-1 promoter. Both the native NIMIN-1 construct and an endogenous copy of NIMIN-1 were induced by SA. However, the mutated promoter construct was much less sensitive to SA than the native NIMIN-1 promoter, indicating that this TGA2-binding motif is directly involved in the modulation of SA-induced NIMIN-1 expression in Arabidopsis.
Fonseca
J P
JP
Departamento de Genética, Evolução e Bioagentes, Universidade Estadual de Campinas, Campinas, SP, Brasil. zepedrof@gmail.com
Menossi
M
M
Thibaud-Nissen
F
F
Town
C D
CD
D017360:Arabidopsis / Q000187:drug effects* / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors / Q000378:metabolism*; D001665:Binding Sites; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D018506:Gene Expression Regulation, Plant / Q000187:drug effects*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism*; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects; D020156:Salicylic Acid / Q000494:pharmacology*; D014157:Transcription Factors
C489413:AHBP-1b protein, Arabidopsis; D029681:Arabidopsis Proteins; D050976:Basic-Leucine Zipper Transcription Factors; D002352:Carrier Proteins; C507154:NIMIN1 protein, Arabidopsis; D009687:Nuclear Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D020156:Salicylic Acid