Mutation of either G box or I box sequences profoundly affects expression from the Arabidopsis rbcS-1A promoter.

Open Access
Publisher Website
Google Scholar
Abstract
A deletion analysis of the Arabidopsis thaliana rbcS‐1A promoter defined a 196 bp region (‐320 to ‐125) sufficient to confer light‐regulated expression on a heterologous Arabidopsis alcohol dehydrogenase (Adh) reporter gene in transgenic Nicotiana tabacum (tobacco) leaves. This region, which contains DNA sequences I, G and GT boxes, with homology to other ribulose‐1,5‐bisphosphate carboxylase small subunit (RBCS) gene promoter sequences, directed expression independent of orientation and relative position in the Adh promoter. Site‐specific mutagenesis of these conserved sequences and subsequent expression analysis in transgenic tobacco showed that both G box and I box mutations in the context of the full (‐1700 to +21) rbcS‐1A promoter substantially reduced the expression of Adh and beta‐glucuronidase (GUS) reporter genes. The G box has previously been shown to specifically bind in vitro a factor isolated from nuclear extracts of tomato and Arabidopsis. This factor (GBF) is distinct from the factor GT‐1 which binds to adjacent GT boxes in the pea rbcS‐3A promoter. Multiple mutations in putative Arabidopsis rbcS‐1A promoter GT boxes had no pronounced affect on expression, possibly due to a redundancy of these sites. Experiments in which rbcS‐1A promoter fragments were fused to truncated 35S CaMV (cauliflower mosaic virus) promoter‐‐GUS reporter constructs showed that cis‐acting CaMV promoter elements could partially restore expression to G‐box‐mutated rbcS‐1A sequences.