FIG 6.

macroH2A1 is not recruited to the replication in HPV16ΔE8^E2 cells. (A) HFK (strain 20) cell lines containing either the HPV16 wild-type or HPV16ΔE8^E2 genome were immunostained with antibodies against macroH2A1.2 (green) and RPA (red). Nuclei are indicated with dotted lines. A white arrow indicates core staining of macroH2A1.2. (B) Quantification of panel A. The mean fluorescent intensity of macroH2A1.2 was calculated using ImageJ (values of >1.0 indicate enrichment). A total of 136 cells were scored with the parental HFK20 strain as a negative control (no RPA foci were detected). In HPV16ΔE8^E2 cells, 58 RPA-positive foci were counted from 41 cells (n = 41) in two independent experiments. A total of 94.8% of foci (55/58) showed no enrichment of macroH2A1. In wild-type HPV16 genome-containing cells (n = 66), 68 RPA-positive foci were scored and 89.1% (61/68) showed enrichment of macroH2A1 (mostly single foci are present in wild-type HPV16 cells). (C) Fluorescence intensity light scans obtained by drawing a line through a nucleus shown in panel A. A gray bar above the scan delineates the position of the replication focus.