Arabidopsis Ethylene-Responsive Element Binding Factors Act as Transcriptional Activators or Repressors of GCC Box–Mediated Gene Expression

TLDR

Ethylene‑responsive element binding factors (ERFs) are plant‑specific transcription factors characterized by a conserved ERF DNA‑binding domain. The study aimed to characterize the Arabidopsis ERF family by isolating cDNAs for five members (AtERF1–5) and examining their structure, DNA‑binding specificity, transactivation capacity, and expression patterns. Researchers cloned the five AtERF cDNAs, performed sequence analysis to define domain conservation, assessed GCC box binding via in vitro assays, and measured transactivation in Arabidopsis leaf protoplasts and reporter gene expression. AtERFs were grouped into three classes based on ERF domain identity, all binding the GCC box; AtERF1,2,5 acted as transcriptional activators, whereas AtERF3,4 served as repressors, and their expression was differentially regulated by ethylene, abiotic stresses, and cycloheximide through EIN2‑dependent or independent pathways, demonstrating that AtERFs modulate GCC box‑mediated gene expression positively or negatively.

Abstract

Ethylene-responsive element binding factors (ERFs) are members of a novel family of transcription factors that are specific to plants. A highly conserved DNA binding domain known as the ERF domain is the unique feature of this protein family. To characterize in detail this family of transcription factors, we isolated Arabidopsis cDNAs encoding five different ERF proteins (AtERF1 to AtERF5) and analyzed their structure, DNA binding preference, transactivation ability, and mRNA expression profiles. The isolated AtERFs were placed into three classes based on amino acid identity within the ERF domain, although all five displayed GCC box–specific binding activity. AtERF1, AtERF2, and AtERF5 functioned as activators of GCC box–dependent transcription in Arabidopsis leaves. By contrast, AtERF3 and AtERF4 acted as repressors that downregulated not only basal transcription levels of a reporter gene but also the transactivation activity of other transcription factors. The AtERF genes were differentially regulated by ethylene and by abiotic stress conditions, such as wounding, cold, high salinity, or drought, via ETHYLENE-INSENSITIVE2 (EIN2)–dependent or –independent pathways. Cycloheximide, a protein synthesis inhibitor, also induced marked accumulation of AtERF mRNAs. Thus, we conclude that AtERFs are factors that respond to extracellular signals to modulate GCC box–mediated gene expression positively or negatively.