Specific Interaction Between a Nicotiana tabacum Nuclear Protein and the Agrobacterium rhizogenes rolB Promoter

Nuclear protein extracts from Nicotiana tabacum plants and cells culture were utilized in gel retardation assays and in vitro and in viva footprinting experiments in order to identify nuclear proteins, that would bind to the 5' non-coding region of the plant oncogene rolB from the Ri plasmid of Agrobacterium rhizogenes. A factor, RBF1 (Rol Binding Factor) was identified that specifically binds to a sequence located between positions -553 and -530 from the translational start codon. This DNA segment is located within a domain controlling the level of expression of rolB and it's inducibility in the non meristematic cells in the root apex (in particular protoderm and root cap). The binding sequence for RBF1 has been identified. The factor is present in extracts prepared from leaves and cells culture of untransformed N. tabacum as well as in transgenic plants, expressing rolA, rolB, and rolC (Spena et al., 1987), in apparently the same amount. Comparing the in vitro footprinting data with an in vivo high-resolution footprinting, obtained by means of a linear amplification procedure utilizing the polymerase chain reaction, we find substantially the same DNA core sequence recognized by RBF1 protein. Our results point to RBF1 as a plant endogenous factor that could be involved in controlling the level and/or tissue specificity expression of the rolB gene.