The DNA-binding activity of an AP2 transcriptional activator HvCBF2 involved in regulation of low-temperature responsive genes in barley is modulated by temperature.
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 degrees C. A low, but detectable, binding activity was observed at 25 degrees C and the binding activity gradually increased as the temperature decreased. The binding activity at 0 degrees C was the highest and more than 10 times higher than that at 25 degrees 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.
Xue
Gang-Ping
GP
CSIRO Plant Industry, 120 Meiers Road, Indooroopilly, Queensland 4068, Australia. Gang.Ping.Xue@pi.csiro.au
D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins; D001483:Base Sequence; D001665:Binding Sites; D003080:Cold Temperature; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics*; D001467:Hordeum / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D020218:Response Elements / Q000235:genetics; D016415:Sequence Alignment; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D015854:Up-Regulation
D029681:Arabidopsis Proteins; C118228:CBF2 protein, Arabidopsis; D015534:Trans-Activators; D004247:DNA