(A) Comparison of the promoter regions of K-Rta and R-Rta, the
replication and transactivator genes for KSHV and RRV, respectively. Nucleotides
71170 to 71697 of KSHV (NC_009333.1) and 66858 to 67364 of RRV (NC_003401.1)
genomic sequence were aligned. Nucleotide numbering indicates the position
relative to the Rta translational start site. Identical nucleotides are
highlighted in gray. Known transcription factor binding sites for C/EBP, Octl,
YY1, Apl, Spl, and XBP-1 in the KSHV Rta promoter (9, 39, 56, 67, 68, 70, 74,
75) are indicated. Of these sites, only the Sp1 binding site was conserved in
the RRV Rta promoter. Transcriptional start sites for KSHV Rta and RRV Rta (16,
42) are identified by solid arrows. Translational start sites for Rta and ORF48
are shown with block arrows with initiation codons shown in black. TATA-like
elements upstream of ORF48 and Rta are shown. (B) 5’ serial truncations
of the RRV Rta promoter region were constructed in the pGL2 firefly luciferase
backbone. (C) Luciferase activity was measured in extracts from permissive Vero
cells transfected with pGL2 R-Rta promoter deletion clones and normalized using
pRL-CMV. Error bars represent the standard deviation of results from triplicate
wells. The region from −136 to −102 containing the conserved Sp1
binding site was critical for Rta promoter activity.