Wheat transcription factors HBP-1a and HBP-1b bind to the hexamer motif, ACGTCA, of wheat histone gene promoters. HBP-1b also binds to the hexamer motif in the promoter of the 35S RNA gene of cauliflower mosaic virus, whereas HBP-1a does not. A cDNA clone encoding HBP-1b was isolated on the basis of its binding specificity to the hexamer motif. The deduced amino acid sequence indicates that HBP-1b, like HBP-1a, belongs to a leucine zipper class of transcription factors. Mutational analyses of the HBP-1a and -1b encoded cDNAs revealed that truncated polypeptides containing the leucine zipper and basic regions are sufficient for DNA binding. HBP-1a and -1b form homodimers, as expected from earlier studies on this class of transcription factors, but did not form heterodimers. Although the hexamer motif or its homologs exist in several plant genes, HBP-1a and -1b exhibited the highest binding affinity to the hexamer motif in the histone promoters, suggesting that both DNA binding proteins are involved in transcriptional regulation of wheat histone genes.