SPECIFIC INTERACTION BETWEEN A NICOTIANA-TABACUM NUCLEAR-PROTEIN AND THE AGROBACTERIUM-RHIZOGENES ROLB PROMOTER

Nuclear protein extracts from Nicotiana tubacum plants and cells cultu re were utilized in gel retardation assays and in vitro and in vivo fo otprinting experiments in order to identify nuclear proteins, that wou ld bind to the 5' non-coding region of the plant oncogene rolB from th e Ri plasmid of Agrobacterium rhizogenes. A factor, RBF1 (Rol Binding Factor) was identified that specifically binds to a sequence located b etween positions -553 and -530 from the translational start codon. Thi s DNA segment is located within a domain controlling the level of expr ession of rolB and it's inducibility in the non meristematic cells in the root apex (in particular protoderm and root cap). The binding sequ ence for RBF1 has been identified. The factor is present in extracts p repared from leaves and cells culture of untransformed N. tabacum as w ell as in transgenic plants, expressing rolA, rolB, and rolC (Spena et al., 1987), in apparently the same amount. Comparing the in vitro foo tprinting 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 recog nized. by RBF1 protein. Our results point to RBF1 as a plant endogenou s factor that could be involved in controlling the level and/or tissue specificity expression of the rolB gene.