In plants, low temperature and dehydration activate a set of genes containing C-repeat/dehydration-responsive elements in their promoter. It has been shown previously that the Arabidopsis CBF/DREB1 transcription activators are critical regulators of gene expression in the signal transduction of cold acclimation. Here, we report the isolation of an apparent homolog of the CBF/DREB1 proteins (CBF4) that plays the equivalent role during drought adaptation. In contrast to the three already identified CBF/DREB1 homologs, which are induced under cold stress, CBF4 gene expression is up-regulated by drought stress, but not by low temperature. Overexpression of CBF4 in transgenic Arabidopsis plants results in the activation of C-repeat/dehydration-responsive element containing downstream genes that are involved in cold acclimation and drought adaptation. As a result, the transgenic plants are more tolerant to freezing and drought stress. Because of the physiological similarity between freezing and drought stress, and the sequence and structural similarity of the CBF/DREB1 and the CBF4 proteins, we propose that the plant's response to cold and drought evolved from a common CBF-like transcription factor, first through gene duplication and then through promoter evolution.Many plants increase their tolerance to freezing after exposure to low nonfreezing temperatures-a phenomenon known as cold acclimation (Hughes and Dunn, 1996;Thomashow, 1998). The major component of this acquired freezing tolerance is the tolerance to dehydration stress caused by extracellular ice formation during the freezing process. The presence of ice lowers the water potential extracellularly and causes water to flow out of cells (Pearce, 1999). Thus, a major cause of freezing damage is the freeze-induced dehydration (Steponkus and Webb, 1992;Thomashow, 1998). Because a plant's ability to survive freezeinduced dehydration is related to its adaptation to drought, it is not surprising that plants respond to low temperature and drought very similarly at the molecular level . Many genes, such as RD (responsive to dehydration), ERD (early responsive to dehydration), COR (cold regulated), LTI (low-temperature induced), and KIN (cold inducible), are induced by both low temperature and drought stress (Ingram and Bartels, 1996;Pearce, 1999; Thomashow, 1999; Shinozaki and YamaguchiShinozaki, 2000). The similarity of cold and drought stresses is further demonstrated by experiments showing that mild drought stress can result in increased freezing tolerance in plants (Clavitier and Siminovitch, 1982;Siminovitch and Cloutier, 1983; Guy et al., 1992).Recently, a major transcriptional regulatory system that controls abscisic acid (ABA) independent gene expression in response to low temperature has been identified (Stockinger et al., 1997;Liu et al., 1998). The system is based on the C-repeat (CRT)/ dehydration-responsive element (DRE) cis-acting element and the trans-acting DNA-binding protein CBF/DREB1 (CRT-binding factor or DRE-binding protein). There are three CB...

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Many plants increase in freezing tolerance in response to low, nonfreezing temperatures, a phenomenon known as cold acclimation. Cold acclimation in Arabidopsis involves rapid cold-induced expression of the C-repeat/dehydrationresponsive element binding factor (CBF) transcriptional activators followed by expression of CBF-targeted genes that increase freezing tolerance. Here, we present evidence for a CBF cold-response pathway in Brassica napus. We show that B. napus encodes CBF-like genes and that transcripts for these genes accumulate rapidly in response to low temperature followed closely by expression of the cold-regulated Bn115 gene, an ortholog of the Arabidopsis CBF-targeted COR15a gene. Moreover, we show that constitutive overexpression of the Arabidopsis CBF genes in transgenic B. napus plants induces expression of orthologs of Arabidopsis CBF-targeted genes and increases the freezing tolerance of both nonacclimated and cold-acclimated plants. Transcripts encoding CBF-like proteins were also found to accumulate rapidly in response to low temperature in wheat (Triticum aestivum L. cv Norstar) and rye (Secale cereale L. cv Puma), which cold acclimate, as well as in tomato (Lycopersicon esculentum var. Bonny Best, Castle Mart, Micro-Tom, and D Huang), a freezing-sensitive plant that does not cold acclimate. An alignment of the CBF proteins from Arabidopsis, B. napus, wheat, rye, and tomato revealed the presence of conserved amino acid sequences, PKK/RPAGRxKFxETRHP and DSAWR, that bracket the AP2/EREBP DNA binding domains of the proteins and distinguish them from other members of the AP2/EREBP protein family. We conclude that components of the CBF cold-response pathway are highly conserved in flowering plants and not limited to those that cold acclimate.Plants vary greatly in their abilities to survive freezing temperatures (Sakai and Larcher, 1987). Whereas plants from tropical regions have essentially no capacity to withstand freezing, herbaceous plants from temperate regions can survive freezing at temperatures ranging from Ϫ5 to Ϫ30°C, depending on the species. It is significant that the maximum freezing tolerance of plants is not constitutive, but is induced in response to low temperatures (below approximately 10°C), a phenomenon known as "cold acclimation" (Hughes and Dunn, 1996;Thomashow, 1999). Nonacclimated wheat (Triticum aestivum L. cv Norstar) plants, for instance, are killed at freezing temperatures of about Ϫ5°C, but after cold acclimation, can survive temperatures down to about Ϫ20°C. Determining what accounts for the differences in freezing tolerance between plant species and the molecular basis of cold acclimation is of basic scientific interest and has the potential to provide new approaches to improve the freezing tolerance of plants, an important agronomic trait.A recent advance in understanding cold acclimation in Arabidopsis was the discovery of the C-repeat/dehydration-responsive element binding factor (CBF) cold-response pathway (see Thomashow, 2001). Arabidopsis encodes a small fami...

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