Synergistic Activation of Seed Storage Protein Gene Expression in Arabidopsis by ABI3 and Two bZIPs Related to OPAQUE2*

The expression of many seed storage protein genes in cereals relies on transcription factors of the bZIP class, belonging to the maize OPAQUE2 family. Here, we describe a survey of such factors in the genome of Arabidopsis thaliana, and the characterization of two of them, AtbZIP10 and AtbZIP25. Expression analysis by in situ hybridization shows that the occurrence of their mRNAs in the seed starts from early stages of development, peaks at maturation, and declines later in seed development, matching temporally and spatially those of the seed storage protein genes encoding 2S albumins and cruciferins. Gel mobility shift assays showed that AtbZIP10 and AtbZIP25 bind the ACGT boxes present in At2S and CRU3 promoters. Moreover, using the yeast two-hybrid system we show that AtbZIP10 and AtbZIP25 can interact in vivo with ABI3, an important regulator of gene expression in the seed of Arabidopsis. Transient expression analyses of a reporter gene under the control of the At2S1 promoter in transgenic plants overexpressing ectopically AtbZIP10, AtbZIP25, and ABI3 reveal that none of these factors could activate significantly the reporter gene when expressed individually. However, co-expression of AtbZIP10/25 with ABI3 resulted in a remarkable increase in the activation capacity over the At2S1 promoter, suggesting that they are part of a regulatory complex involved in seed-specific expression. This study shows a common mechanism of ABI3 in regulating different seed-specific genes through combinatorial interactions with particular bZIP proteins and a conserved role of O2-like bZIPs in monocot and dicot species.