Fig. 2.

Stress-induced changes in mtDNA copy number requires ongoing mtDNA replication. U20S cells were transfected with nonsilencing (A) or Rad51-specific (B) siRNAs (10 nm). 48 h following transfection, cells were treated with 5 milliunits/ml GO or treated with GO following a 2 h pre-treatment with 20 μM ddC or 3 μM aphidicolin. DNA was isolated at 0, 0.5, and 1 h after GO addition. MtDNA copy number was determined by qPCR, with changes calculated relative to unstressed controls and normalized to the 18S rRNA gene. Error bars indicate S.E.M. (n = 4). (C) Western blot analysis of Rad51 in total cell lysates following 48 h of nonsilencing (N.S.) or Rad51-specific siRNA treatment. GAPDH serves as the loading control. Blots are representative of 4 independent experiments.