Article Figures & Data
Figures
- Fig. 1.
Telomerase activity in stable HFF clones. Stable HFF clones obtained by transfection with pGRN145 expressing the catalytic subunit of telomerase (Tel) were analyzed for telomerase activity by the TRAP assay using a TRAPEZE kit (Intergen) according to the manufacturer's instructions. Two thousand cells were analyzed in all samples. TRAP products were separated by gel electrophoresis (11.25% polyacrylamide) and detected by autoradiography. As a negative control, cell lysates were incubated at 85°C for 10 min (ΔH) to inactivate telomerase. TSR8, positive control for the PCR; IC, internal control for all reactions.
- Fig. 2.
Replicative life span of telomerase-positive and -negative clones. Cells were counted and passaged every 5 to 7 days, and new population doublings were calculated. Cultures were considered senescent (*) if they had undergone less than one population doubling in 2 weeks.
- Fig. 3.
Comparison of immediate-early and late gene expression in HFF and telomerase-positive cells. HFF or telomerase 18 cells were infected with HCMV at an MOI of 5. Cell lysates were prepared at the indicated times postinfection (P.I.), and 50 μg of protein from each sample was analyzed by Western blotting using antibodies against either IE1 (Chemicon 810) or pp28 (9, 11).
Tables
- Table 1.
Comparison of wild-type HCMV growth on telomerase-positive and telomerase-negative cell linesa
Cell line Telomerase status No. of population doublings PFU/ml (107) HFF-R2 − 34.5 1.2 Telomerase 9 − 48.5 3.0 Telomerase 10 − 47 1.1 Telomerase 13 − 48 1.1 Telomerase 12 + 51 4.6 75.5 3.5 Telomerase 16 + 46.5 2.9 65 4.9 Telomerase 18 + 47 3.2 75 5.7 a Cells were infected at an MOI of 5 PFU/cell. Virus was harvested 5 days later, and yields were determined by plaque assay on HFF-R2 cells.
- Table 2.
Comparison of wild-type and ΔUL82 viral titers on complementing and noncomplementing cell linesa
Virus PFU/ml HFF-R2 WF28-71-HA Wild type 2.06 × 106 1.58 × 106 ΔUL82 NPD 3.25 × 105 a Cells were infected at an MOI of 0.01 PFU/cell. Virus was harvested 21 days later, and yields were determined by plaque assay on WF28-71-HA cells. NPD, no plaques detected in undiluted inoculum.
