The rice bZIP transcriptional activator RITA-1 is highly expressed during seed development.
Systematic protein-DNA binding studies have shown that plant basic leucine zipper (bZIP) proteins exhibit a differential binding specificity for ACGT motifs. Here, we show that the rice transcription activator-1 (RITA-1) displays a broad binding specificity for palindromic ACGT elements, being able to bind A-, C-, and G-box but not T-box elements. By using gel mobility shift assays with probes differing in sequences flanking the hexameric core, we identified high-affinity A-, C-, and G-box binding sites. Quantitative and competition DNA binding studies confirmed RITA-1 specificity for these sites. Using rice protoplasts as a transient expression system, we demonstrated that RITA-1 can transactivate reporter genes possessing high-affinity but not low-affinity RITA-1 binding sites. Our results established a direct relationship between in vivo transactivation and in vitro binding activity. Transient expression assays that demonstrated the ability of RITA-1 to transactivate a construct containing rita-1 5' flanking sequences suggest that the factor may be autoregulated. Histochemical analysis of transgenic rice plants showed that a rita-1-beta-glucuronidase transgene is expressed in aleurone and endosperm cells of developing rice seeds. We propose that RITA-1 plays a role in the regulation of rice genes expressed in developing rice seeds.
Izawa
T
T
Laboratory of Plant Molecular Biology, Rockefeller University, New York, New York 10021-6399.
Foster
R
R
Nakajima
M
M
Shimamoto
K
K
Chua
N H
NH
D000595:Amino Acid Sequence; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D018506:Gene Expression Regulation, Plant / Q000235:genetics; D016350:Leucine Zippers / Q000235:genetics; D008969:Molecular Sequence Data; D012275:Oryza / Q000235:genetics*; D012639:Seeds / Q000235:genetics / Q000254:growth & development*; D015533:Transcriptional Activation / Q000235:genetics*
D018076:DNA, Complementary