Figure 4

p30 homologue HTLV-2 p28 protein does not interact with Cyclin E and does not affect cell cycle progression. A) 293T cells were transfected with comparable expression amounts of plasmids expressing p30-HA and p28-HA proteins, WB with HA antibody or actin for loading control. B) Hela cells were transfected with p30HA or p28HA along with GFP expression vector and GFP expression was monitored to demonstrated equivalent transfection efficiencies. C) p28 does not interact with Cyclin E. 293T cells were transfected either with Cylin E-myc and p30-HA or cyclin E-myc and p28-HA. Cell extracts were prepared and the interaction in vivo between Cyclin E and both viral proteins were assessed by co-immunoprecipitation. D) p28 does not affect the cell cycle progression. Hela cells were transfected either with GFP-p30 or GFP-p28 expression vectors, synchronized with Hydroxyurea (2 mM) and then released for 0 h, 8 h. The cell cycle distribution was assessed by flow cytometry of GFP positive gated cells using the Dye cycle Violet. E) Histograms indicating the percentage of cells in each phase of the cell cycle distribution.