The 5'-upstream region of the rolC gene of the Ri plasmid is expressed specifically in phloem cells of transgenic higher plants. In this study, we demonstrated that the rolC promoter is activated by sucrose in phloem cells of transgenic tobacco seedlings bearing rolC promoter-uidA chimeric fusion gene. Since the rolC promoter is not activated by sorbitol, sucrose metabolism rather than osmotic pressure exerted by the disaccharide may be responsible for induction. Thus, experiments using 5'-upstream deletion mutants, internal deletion mutants, and chimeric constructs with a heterologous promoter (-90 region of the cauliflower mosaic virus 35S promoter) were conducted to define the region of the rolC promoter involved in sucrose activation. The results indicated that a cis-acting sucrose responsive region of the rolC promoter is located between -135 and -94 bp with respect to the transcription initiation site. In phloem cells, high concentrations of sucrose are encountered owing to ongoing translocation of photosynthates from source to sink tissues. Therefore, sucrose as a signal molecule may regulate the phloem-specific expression of the rolC promoter.