1. Downregulation of Rab7 expression with RNAi caused accumulation of late autophagic vacuoles.
Downregulation of Rab7 expression with RNAi caused accumulation of late autophagic vacuoles. HeLa cells were transfected with empty pSUPER or pSUPER Rab7 RNAi and grown for 3 days. Three parallel dished were kept in full culture medium (FCS), another three dishes were starved of amino acids for 2 hours (EBS), and another three dishes were first starved of amino acids for 2 hours and then shifted back to full medium for 2 hours (EBS - FCS). (A) Expression levels of Rab7 and LC3 were controlled by western blotting. Equal amounts of cell extract (40 μg) were loaded in each lane, and Ponceau staining was used to confirm equal transfer of proteins to each lane on the PVDF membrane (not shown). (B) Accumulation of early (Avi) and late (Avd) autophagic vacuoles was estimated by quantitative electron microscopy. The P values indicate statistical significance of the difference between the Rab7 RNAi and the pSUPER sample (t-test). Error bars indicate s.e.m.
Stefanie Jäger et al. J Cell Sci 2004;117:
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