Functional analysis of the 5′ untranslated region of the sucrose phosphate synthase rice gene ( sps1)

Abstract

The sps1 gene, encoding the sucrose phosphate synthase (SPS) in rice has an unusually long 5′ untranslated region (leader) of 368 nt with a complex predicted secondary structure. Elimination of the leader from the native sps1 promoter diminishes the in vivo expression of the GUS reporter gene by 10-fold. Insertion of the leader downstream, but not upstream, of the −46(35S) CaMV promoter in the correct orientation produces an expression enhancement equivalent to that conferred in the sps1 native promoter. This expression enhancement is observed using both in vivo transient expressions in tobacco leaves and in stably transformed Arabidopsis plants. The sps1 leader also enhances the level of reporter gene expression in the strong 35S CaMV and Cassava Vein Mosaic Virus (CVMV) promoters, albeit at a lower level than in weaker promoters. In vitro transcription and translation studies using transcripts of the uidA gene, with and without the sps1 leader, show that the sps1 5′ untranslated region enhances translation over three-fold. Deletion analysis of the sps1 leader showed that in vivo enhancement is totally conserved in the 5′Δ85 and diminished by half in the 5′Δ148; the deletion 3′Δ190 conserves two thirds of the enhancement, suggesting redundancy in sequence elements.