FIG. 3.

Effect of HCMV infection on cell cycle regulatory proteins. Immunoblots of cellular lysates corresponding to mock- and HCMV-infected HELF are shown. Cells were growth arrested by serum starvation and subsequently infected with HCMV (MOI, 3) or mock infected with conditioned medium containing 10% FCS as described in Materials and Methods. At different time points after infection, cell lysates were prepared, and equal amounts (30 μg) were separated by gel electrophoresis and transferred to Immobilon membranes. The proteins were then probed with anti-IE1/IE2, anti-pRb, anti-ppRb (phosphorylated form of pRb), anti-p53, anti-p16, and anti-p21 Abs. The membrane was then incubated with the appropriate secondary Ab conjugated with horseradish peroxidase and visualized with an ECL kit (Amersham). Actin immunodetection was performed as an internal control.