Figure 6. PTEN expression in U87MG cells increases the formation of autophagosomes under various proteolysis conditions.

U87MG cells expressing WT-PTEN, G129E-PTEN or C124S-PTEN (A) and mock-treated U87MG cells or expressing WT-PTEN or C124S-PTEN (B) were incubated under high (KH), intermediate (Ins or EAA)(A and B) and low (Ins/EAA)(only in A) proteolysis conditions for 2 h as in Figure 5C in the presence of lysosomal inhibitors (100 µM leupeptin and 20 mM NH₄Cl). Extracts (75 µg protein) were analyzed by SDS-PAGE and immunoblot with antibodies that recognize LC3 and, as a loading control, actin. The position of LC3-I and LC3-II bands are indicated on the left and molecular weight markers are indicated on the right. The Western blots on the left show representative experiments and the histograms on the right show the means ± SD of the densitometric analysis of the LC3-II/ratios from five different experiments. Stars indicate statistically significant differences from the corresponding (high, intermediate or low proteolysis conditions) values in the cells expressing the C124S PTEN mutant (A) and mock-treated cells (B) at **p<0.01 and ***p<0.005.