Characterization of three rice basic/leucine zipper factors, including two inhibitors of EmBP-1 DNA binding activity

The promoter of the wheat Em gene contains elements with a CACGTG core sequence (G-boxes), which are recognized by EmBP-1, a wheat basic/leucine zipper (bZIP) protein. G-boxes are required for Em expression in response to the phytohormone abscisic acid and for transactivation by the Viviparous-1 protein (VP1) using transient expression systems. In order to identify other factors that are part of the transcriptional complex that associates with G-boxes, we have screened a rice (Oryza sativa) cDNA library with biotinylated EmBP-1. We have isolated osZIP-1a, a homolog of EmBP-1 and other plant G-box-binding factors. We show that EmBP-1 and osZIP-1a will preferentially heterodimerize in vitro. Overexpression of osZIP-1a in rice protoplasts can enhance expression from the Em promoter only in the presence of abscisic acid. Two other clones have been identified by screening with EmBP-1: osZIP-2a and osZIP-2b. These osZIP-2 factors represent a novel class of bZIP proteins with an unusual DNA-binding domain that does not recognize G-boxes. The osZIP-2 factors can heterodimerize with EmBP-1 and prevent it from binding to the Em promoter. Interestingly, osZIP-1a does not heterodimerize with the osZIP-2 factors and its DNA binding activity is unaffected by their presence. Thus, osZIP-2 factors may be involved in sequestering a particular group of G-box-binding factors into inactive heterodimers.