Fig 1. The TFIIB recognition element is required for antisense transcriptional activity of the LTR promoter.

(A) Schematic representation of sense and antisense transcription initiation of the BLV provirus. Sense transcription requires the binding of the TATA-binding protein (TBP), a subunit of the transcription factor IID (TFIID), to the TATA box located in U3. TFIID further interacts with other general transcription factors (GTFs), resulting in recruitment of RNA polymerase type-II (RNAPII). Initiation of antisense transcription is predicted to be initiated by the binding of TFIIB to the TFIIB recognition element (BRE) located in U5 (B) Nucleotide sequence of mutations (A, A1 and A2 in the BRE and B1 in the motif ten element/downstream promoter element MTE/DPE) introduced in the LTR and tested in reported assays. (C) Schematic representation of Firefly-Renilla dual-luciferase reporter plasmids containing either the full-length LTR or the R-U5 region. (D-E) Luciferase activities promoted by the WT and mutated minimal antisense promoter (R-U5). HEK293FT cells were co-transfected with the reporter plasmids (WT, A, A1, A2, B1) and a vector expressing either mock (pSG5) or the viral transactivator Tax (pSGTax). (F-G) Same experiment as described in panels D-E except that a dual reporter containing a full-length LTR was used. Twenty-four hours after transfection, Firefly and Renilla luciferase activities were measured and normalized to the corresponding mean luminescence generated by the full-length LTR reporter and pSGTax, arbitrarily set to 100. RLU: relative luminescence units. Data results from at least 3 independent experiments. p-values were calculated according to Mann-Whitney tests.