Binding of an endosperm-specific nuclear protein to a maize beta-zein gene correlates with zein transcriptional...

Promoter regions of alpha- and beta-zein genes were analyzed for binding of nuclear proteins from developing endosperm and seedling tissue of maize. Using a band-shift assay, we identified two distinct protein factors, alpha-1 and beta-1, that interacted specifically with alpha- and beta-zein gene promoter regions, respectively. Alpha-1 was present in nuclei from both endosperm and seedling tissue, whereas beta-1 was found only in nuclei from developing endosperm tissue. Mixing of nuclear extracts demonstrated that seedling tissue contained undetectable amounts of beta-1, rather than having an inhibitor for formation of the beta-1/DNA complex. Chemical footprinting analysis localized the beta-1 recognition site to a 22 bp sequence flanked by CCAT and TATA boxes. The apparent molecular mass of beta-1 was determined to be 29 kDa by southwestern blotting. Based on in vitro binding assays, the greatest concentration of the beta-1 in endosperm nuclei is at 16 days after pollination, which coincides with the time of highest transcriptional activity of the beta-zein gene. These results suggest that beta-1 may act as a tissue-specific, trans-acting regulator of the expression of the beta-zein gene in developing maize endosperm.