Sequence-specific interactions of wound-inducible nuclear factors with mannopine synthase 2' promoter...

A 318 bp mannopine synthase 2' (mas2') promoter element from the T-DNA of Agrobacterium tumefacians can direct wound-inducible and root-preferential expression of a linked uidA gene in transgenic tobacco plants. Wound inducibility is further enhanced by sucrose in the medium. Promoter deletion analysis indicated that the sucrose enhancement is conferred by a region extending from -318 to -213. DNase I footprinting indicated that an A/T-rich DNA sequence in this region is protected by tobacco nuclear factors. Regions extending from -103 to +66 and from -213 to -138 directed wound-inducibile expression of a linked uidA gene when placed downstream of a CaMV 35S enhancer or upstream of a truncated (-209) CaMV 35S promoter, respectively. DNase I footprinting analyses indicated that proteins from wounded tobacco leaves specifically bound to three contiguous motifs downstream of the mas2' TATA box. In addition to a common retarded band formed by the upstream wound-responsive element complexed with proteins from either wounded or unwounded tobacco leaves, two unique retarded bands were observed when this element was incubated with protein from wounded leaves. Methylation interference analysis additionally identified an unique motif composed of promoter elements and nuclear factors derived specifically from wounded tobacco leaves. We propose a model to describe the involvement of nuclear factors with mas2' promoter elements in wound-inducible gene expression.