The DNA‐binding activity of an AP2 transcriptional activator HvCBF2 involved in regulation of low‐temperature responsive genes in barley is modulated by temperature

Abstract

Summary: An HvCBF2 cDNA was isolated from barley leaves. It encoded a protein containing an AP2 DNA‐binding domain homologous to C‐repeat (CRT)/dehydration‐responsive element (DRE) binding factors (CBF/DREB1). In contrast to the previously reported cold‐inducible CBF/DREB1 genes, HvCBF2 was expressed in barley leaves under non‐stress conditions. Only a transient increase in the HvCBF2 transcript level was observed during cold treatment. Transactivation analysis showed that HvCBF2 was a transcriptional activator, capable of activating expression of a reporter gene driven by a low‐temperature and drought‐responsive HVA1s promoter in barley leaves. The activity of HvCBF2 as a transcriptional activator was upregulated by low temperature. DNA‐binding analysis revealed that HvCBF2 did not bind to the CRT/DRE motif at 30°C. A low, but detectable, binding activity was observed at 25°C and the binding activity gradually increased as the temperature decreased. The binding activity at 0°C was the highest and more than 10 times higher than that at 25°C. The activation and inactivation of HvCBF2 activity were reversible and were achieved in a cell‐free system simply by temperature change. Analysis of the binding sequence showed that HvCBF2 bound to a (G/a)(T/c)CGAC core motif, where the lower‐case letters are less efficient bases. These data suggest that HvCBF2 is a transcription factor interacting with the core CRT/DRE motif containing a preferred sequence of GTCGAC and its DNA‐binding activity is regulated by temperature. This represents a new type of activation mechanism for transcriptional activators.