Arabidopsis thaliana CBF1 encodes an AP2 domain-containing transcriptional activator that binds to the C-repeat/DRE, a cis-acting DNA regulatory element that stimulates transcription in response to low temperature and water deficit

Abstract

Recent efforts have defined a cis-acting DNA regulatory element in plants, the C-repeat͞dehydration responsive element (DRE), that stimulates transcription in response to low temperature and water deficit. Here we report the isolation of an Arabidopsis thaliana cDNA that encodes a C-repeat͞DRE binding factor, CBF1 (C-repeat͞DRE Binding Factor 1). Analysis of the deduced CBF1 amino acid sequence indicates that the protein has a molecular mass of 24 kDa, a potential nuclear localization sequence, and a possible acidic activation domain. CBF1 also has an AP2 domain, which is a DNA-binding motif of about 60 aa present in the Arabidopsis proteins APETALA2, AINTEGUMENTA, and TINY; the tobacco ethylene response element binding proteins; and numerous other plant proteins of unknown function. The transcript levels for CBF1, which appears to be a single or low copy number gene, did not change appreciably in plants exposed to low temperature or in detached leaves subjected to water deficit. Binding of CBF1 to the C-repeat͞DRE was demonstrated in gel shift assays using recombinant CBF1 protein expressed in Escherichia coli. Moreover, expression of CBF1 in yeast was found to activate transcription of reporter genes containing the C-repeat͞DRE as an upstream activator sequence but not mutant versions of the DNA element. We conclude that CBF1 can function as a transcriptional activator that binds to the C-repeat͞DRE DNA regulatory element and, thus, is likely to have a role in cold-and dehydrationregulated gene expression in Arabidopsis.

Transcriptomic Analysis Reveals Calcium Regulation of Specific Promoter Motifs inArabidopsis

“…By analyzing the promoter sequences of this combined list of genes induced by our electrical impulses, we found four distinct classes of short DNA motif that appeared at a frequency statistically higher than in the genome as a whole (Table 2, Figure 2). These four motifs most closely resembled motifs already described in the plant molecular biology literature, namely the CRT/DRE coldand drought-responsive element (Yamaguchi-Shinozaki and Shinozaki, 1994;Stockinger et al, 1997;Liu et al, 1998), the ABRE abscisic acid response element (Hobo et al, 1999), the binding site of the CAMTA transcription factors (which we have named CAM box) (Yang and Poovaiah, 2002), and finally the Site II motif (Tré mousaygue et al, 2003). The ABRE (CACGTGT) and CAM box (ACGCGT) completely fit the consensus of the ABRE (CACGTG[T/C/G]) and ABRE coupling element ([C/A]ACGCG[T/ C/A]) identified by our previous microarray experiment (Kaplan et al, 2006).…”

“…Indeed, the CRT and CAM box were upregulated by cold treatment, whereas the ABRE and Site II were not (see Supplemental Figure 4 online), suggesting the possibility of differential responses of the four motifs to the [Ca 2+ ] c elevation elicited by this treatment. The CRT is already well-known to respond to low temperature treatment (Stockinger et al, 1997), and it is interesting that the CAM box is also upregulated by cold, because CAMTAs have been shown to be Arabidopsis seedlings expressing LUC+ under the control of either a minimal promoter only (control) or four repeats of the cis-element sequences CRT or Site II were pretreated with either 0.5 mM LaCl 3 (La 3+ ) or water for 15 min before the addition of either 0.3% DMSO control (mock) or W7 (to a final concentration of 600 mM). Tissue was harvested 3 h after treatment.…”

Cold Induction of Arabidopsis CBF Genes Involves Multiple ICE (Inducer of CBF Expression) Promoter Elements and a Cold-Regulatory Circuit That Is Desensitized by Low Temperature

“…Northern transfers were prepared and hybridized as described (Hajela et al, 1990) using normal high-stringency wash conditions (Stockinger et al, 1997).…”

“…Within 15 min of exposing plants to low temperatures, transcripts accumulate for a family of genes designated CBF1, CBF2, and CBF3 Jaglo-Ottosen et al, 1998; Medina et al, 1999), or DREB1b, DREB1c, and DREB1a (Liu et al, 1998), respectively, which encode transcriptional activators that are members of the AP2/EREBP family of DNA-binding proteins (Riechmann and Meyerowitz, 1998). These transcription factors bind the cold-and dehydration-responsive DNA regulatory element designated the CRT (C-repeat)/ DRE (dehydration response element); (Baker et al, 1994;Yamaguchi-Shinozaki and Shinozaki, 1994;Stockinger et al, 1997) that is present in the promoters of COR and many other cold-responsive genes and stimulate their transcription. Expression of the CBF regulon of target genes then leads to an increase in freezing tolerance Jaglo-Ottosen et al, 1998;Liu et al, 1998;Kasuga et al, 1999).…”

Large deletions in the CBF gene cluster at the Fr-B2 locus are associated with reduced frost tolerance in wheat

“…While Arabidopsis has three closely-related CBF genes tandemly arrayed in a cluster on chromosome four Medina et al 1999;Stockinger et al 1997), wheat and other temperate cereals have undergone an expansion in this family. In hexaploid wheat, for example, there are at least 15 CBF genes in each of the three wheat genomes, including eleven at each of the three Fr-2 loci (Badawi et al 2007).…”