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Rice tungro bacilliform virus (RTBV) is a double-stranded DNA virus with a single, tissue-specific promoter that is expressed primarily in phloem tissues. Rice transcription factors RF2a and RF2b bind to Box II, a cis element adjacent to the TATA box, and control gene expression from the promoter. Mutations were made in the promoter to delete or mutate Box II and the mutated promoters were fused to a reporter gene; the chimeric genes were expressed in transient BY-2 protoplast assays and in transgenic Arabidopsis plants. The results of these studies showed that Box II is essential to the activity of the RTBV promoter. A chimeric beta-glucuronidase (GUS) reporter gene containing the Box II sequence and a minimal promoter derived from the Cauliflower mosaic virus 35S promoter were co-transfected into protoplasts with gene constructs that encoded RF2a or RF2b. The reporter gene produced threefold higher GUS activity when co-transfected with RF2a, and 11-fold higher activity when co-transfected with RF2b, than in the absence of added transcription factors. Moreover, chimeric reporter genes were activated by approximately threefold following induction of expression of the RF2a gene in transgenic Arabidopsis plants. The work presented here and earlier findings show that Box II and its interactions with cognate rice transcription factors, including RF2a and RF2b, are essential to the activity of the RTBV promoter and are probably involved in expression of the RTBV genome during virus replication.

Essential role of the Box II cis element and cognate host factors in regulating the promoter of Rice tungro bacilliform virus