Fig. 3. L1 co-expression increases E2-dependent transcription activation. (a) C33a cells were co-transfected with an E2-responsive firefly luciferase reporter (6E2tk), E2 expression plasmid, and increasing amounts of L1 expression plasmid. E2 and L1 protein levels were determined by Western blotting. Molecular mass standards are indicated on the left. (b) Luciferase activity in cell lysates was measured and normalized to activity in cells transfected with 6E2-tk-Luc alone. (c) C33a cells were co-transfected as described in (a) and (d) transcription activity of full-length E2 was compared with a truncated E2 protein containing aa 200–365 (E2-HC). (d) Luciferase activity was determined as described in (b). (e) C33a cells were co-transfected with HPV16 Ori plasmid, E2 and E1 expression plasmids and increasing amounts of L1 expression plasmid. Protein expression was detected by Western blotting. Molecular mass standards are indicated on the left. (f) DpnI-digested DNA quantified by qPCR. The data are expressed as fold increase in replication over DpnI-digested pOri16M alone. All data shown represent the mean ± se of three independent experiments. Significance was tested using Student's t-test; *P < 0.05, **P < 0.01, ***P < 0.001.