FIG. 9.

Reduced cell energy charge and slowed cell growth with PNPase reduction. (A) Growth medium color change for continuously growing HEK293 cells (and HeLa and Jurkat T cells [data not shown]) with PNPase knockdown (RNAi). (B) PNPase knockdown cells have increased lactate accumulation over time. Cells were incubated in medium with 10% serum, and lactate was measured each day for 6 days. Day 0 equals day 8 following PNPase RNAi knockdown. (C) PNPase knockdown cells have impaired ATP production. ATP levels were measured in both control and PNPase knockdown cells. Three independent experiments showed similar results. (D) Increase in AMPK phosphorylation with PNPase knockdown as assessed by immunoblotting. (E) Cells with reduced PNPase have slowed growth. Control and PNPase RNAi cells were assessed in triplicate each day for 3 days by MTT assay (left panel) or for up to 6 days by trypan blue exclusion assay (right panel) for changes in cell growth. Data are representative of three independent experiments.