DREB1A/CBF3 Is Repressed by Transgene-Induced DNA Methylation in the Arabidopsis ice1-1 Mutant

Satoshi Kidokoro,Kazuko Yamaguchi-Shinozaki,Tomona Ishikawa,Takamasa Suzuki,Kazuo Shinozaki,June-Sik Kim

doi:10.1105/tpc.19.00532

Abstract

DREB1/CBFs are key transcription factors involved in plant cold stress adaptation. The expression of DREB1/CBFs triggers a cold-responsive transcriptional cascade, after which many stress tolerance genes are expressed. Thus, elucidating the mechanisms of cold stress-inducible DREB1/CBF expression is important to understand the molecular mechanisms of plant cold stress responses and tolerance. We analyzed the roles of a transcription factor, INDUCER OF CBF EXPRESSION1 (ICE1), that is well known as an important transcriptional activator in the cold-inducible expression of DREB1A/CBF3 in Arabidopsis (Arabidopsis thaliana). ice1-1 is a widely accepted mutant allele known to abolish cold-inducible DREB1A expression, and this evidence has strongly supported ICE1-DREB1A regulation for many years. However, in ice1-1 outcross descendants, we unexpectedly discovered that ice1-1 DREB1A repression was genetically independent of the ice1-1 allele ICE1(R236H). Moreover, neither ICE1 overexpression nor double loss-of-function mutation of ICE1 and its homolog SCRM2 altered DREB1A expression. Instead, a transgene locus harboring a reporter gene in the ice1-1 genome was responsible for altering DREB1A expression. The DREB1A promoter was hypermethylated due to the transgene. We showed that DREB1A repression in ice1-1 results from transgene-induced silencing and not genetic regulation by ICE1. The ICE1(R236H) mutation has also been reported as scrm-D, which confers constitutive stomatal differentiation. The scrm-D phenotype and the expression of a stomatal differentiation marker gene were confirmed to be linked to the ICE1(R236H) mutation. We propose that the current ICE1-DREB1 regulatory model should be revalidated without the previous assumptions.

Highlights

Cold stress is an environmental condition that affects plant growth, development, and productivity
To identify the cis-acting elements involved in the cold-inducible expression of DREB1A, we generated transgenic Arabidopsis plants that express an emerald luciferase (ELUC) reporter gene driven by four tandem repeats of the DREB1A promoter fragment (2143 to 255 bp from the transcription start site), including two conserved sequences among the promoters of three DREB1s, and its minimal promoter (257 to 1118 bp) and named it 1AR:ELUC (Figure 1A)
Their genetic behaviors seemingly contrast: the INDUCER OF CBF EXPRESSION1 (ICE1)(R236H) mutation in ice1-1 showed a dominantnegative effect on DREB1A expression, while the same mutation in scrm-D showed a semidominant positive effect on the expression of the downstream genes involved in stomatal development

Summary

Introduction

Cold stress is an environmental condition that affects plant growth, development, and productivity. The expression of numerous genes that function in the stress response and in tolerance is induced in various plant species. The products of these genes function to enhance freezing stress tolerance and to regulate gene expression under cold stress conditions (Thomashow, 1999; Yamaguchi-Shinozaki and Shinozaki, 2006). DREB1A/CBF3, DREB1B/CBF1, and DREB1C/CBF2, bind to the DRE, activating the expression of many downstream cold-inducible genes. Overexpression of DREB1/CBFs improves stress tolerance to freezing, drought, Since all three DREB1 genes are rapidly and significantly induced by cold stress, their induction is considered to be the first switch in the cold-responsive expression of numerous genes (Yamaguchi-Shinozaki and Shinozaki, 1994). Some transcription factors have been identified to regulate the cold-inducible expression of DREB1s.

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Results

Conclusion