OPAQUE2 (O2) is a regulatory gene that predominantly affects the expression of the 22-kD class of zein storage protein genes at the level of transcription. The O2 gene encodes a polypeptide belonging to the basic domain/leucine zipper (bZIP) class of transcriptional regulatory proteins. Our prior analyses have demonstrated that the O2 protein binds 22-kD zein gene promoters as a homodimer in vitro and have also suggested that O2 may bind as a heterodimer in vivo. To identify cDNAs encoding other bZIP motifs that might interact with O2, a portion encoding the bZIP motif from an O2 cDNA was used to screen an endosperm cDNA library. Sequence analysis of one isolated recombinant phage indicated the presence of a bZIP motif similar to O2. The protein product of this partial cDNA, designated OHP1, can bind the O2 target site both as a homodimer and in a heterodimeric complex with O2. Whole genome DNA gel blot analysis of maize recombinant inbreds revealed two strongly hybridizing restriction fragments, neither of which mapped close to any locus known to affect zein expression. RNA gel blot analysis revealed an approximately 1.7-kb transcript that is expressed in all organs examined except the female flower and is also expressed in endosperms homozygous for o2 and other mutations that affect zein expression (opaque7, floury2, and Defective endosperm b-30). Based on these results and previously reported data, we propose models to accommodate OHP1 in the regulation of zein gene expression by O2.
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