FIG 2.

Y102E abrogates E2 transcriptional activation. (A) C33a cells were transfected in triplicate with 50 ng/well of the pCG-E2 wild type or mutants (Y102F and Y102E) or mRFP-GFP (−) as well as 75 ng/well pGL2-E2BS-Luc. Cells were lysed on-plate for luminescence detection using the PHERAstar system. RLU, relative light units; **, P < 0.01; NS, not significant. Values are expressed as means ± SEM. (B) One microgram each of the WT and mutant constructs (YF and YE) or mRFP-GFP (−) was expressed in C33a cells and probed for BPV-1 E2 with B201 antibodies. E2, full-length BPV-1 E2; *, nonspecific band. (C) CV-1 cells were transfected with 10 ng/well E2, pCG-E2R (E2R [aa 162 to 410]), or mRFP-GFP plus wells with 10 times the amount of Y102E (100 ng/well) (10×YE). **, P < 0.01. Values are expressed as means ± SEM. (D) CV-1 cells were transfected and prepared for Western blotting with the inclusion of samples with 5 times the amount of Y102E (5×YE), 10 times the amount of Y102E (10×YE), or E2R. The blot was probed for BPV-1 E2 with B201. E2, full-length BPV-1 E2; *, nonspecific band.