Stalling by RNA polymerase II in the polyomavirus intergenic region is dependent on functional large T antigen

RNA polymerase II encounters an elongation block and stalls in vivo during transcription of the late strand of polyomavirus DNA. In this study, we performed transcriptional run-on assays and localized the stalling site to a 164-nucleotide region (nt 11-175) that contains specific binding sites for polyomavirus large T antigen. The effect of large T antigen on elongation by RNA polymerase II through this region was examined in cells infected with a mutant polyomavirus (AT3-ts25E) which encodes a thermolabile large T antigen. Removal of functional large T antigen by shifting to the nonpermissive temperature (39 degrees) eliminated stalling by RNA polymerase in this region, although RNA polymerases transcribing other regions of the viral genome were unaffected. RNA polymerase resumed stalling when functional large T antigen was again allowed to accumulate by shifting back to the permissive temperature (32 degrees). We conclude that stalling by RNA polymerase II in vivo is dependent on the presence of functional large T antigen.