Fig 5.

ORC1 maintains the repressive histone modifications enriched on the HIV-1 promoter. (A) J-Lat 10.6 cells were lysed and immunoprecipitated with anti-HP1α antibody. The endogenous ORC1 enriched by HP1α was immunoblotted with anti-ORC1 antibody. (B) HEK293T cells transfected with GFP-tagged ORC1 were treated with AF568-tagged antibodies against HP1α, SUV39H1, and H3K9me3, and then imaged using super-resolution structured illumination microscopy (SR-SIM). (C) J-Lat 10.6 cells were lysed and immunoprecipitated with anti-SUV39H1 antibody. The endogenous ORC1 enriched by SUV39H1 was immunoblotted with anti-ORC1 antibody. (D and E) ChIP assay with antibodies against HP1α and H3K9me3 was performed in shLuc- and shORC1-treated J-Lat 10.6 cells. The knockdown efficiency was validated by Western blot, as shown in (E). (F) Flag-tagged ORC1 was transfected in J-Lat 10.6 cells. The endogenous EZH2 enriched by ORC1 was immunoblotted with anti-EZH2 antibody. (G) HEK293T cells transfected with GFP-tagged ORC1 were treated with AF568-tagged antibodies against EZH2 and H3K27me3, and then imaged using SR-SIM. (H and I) ChIP assay with antibody against H3K27me3 was performed in shLuc- and shORC1-treated J-Lat 10.6 cells. The knockdown efficiency was validated by Western blot, as shown in (I).