FIGURE 8.

Effect of LAMP-2C on cell cycle regulators. (A) Chk1 activation was examined by detecting cellular levels of Chk1 phosphorylation at Ser345. (B) Chk1 subcellular localization was determined by extracting cytoplasmic and nuclear proteins from DM331 pCMV and DM331 2C myc cells. Protein levels were detected by western blotting. (C) Protein and mRNA levels of cell cycle regulators in DM331 pCMV and DM331 2C myc, p53 and p21, were examined by western blotting and qPCR. (D) Lysates from DM331 zeo and DM331 2C were resolved by SDS-PAGE and probed to detect p21 levels. Protein expression was quantified by densitometry and levels were normalized to actin levels. (E) Protein levels of p21 in SLM2-Mel pCMV and SLM2-Mel 2C myc were detected by western blotting. Protein expression was quantified by densitometry and levels were normalized to actin levels. Measurements in (A–E) represent relative values calculated by setting the results obtained for cells transfected with an empty vector equal to one for comparison to cells with ectopic LAMP-2C. Data were analyzed by two-way ANOVA or by two-tailed, unpaired Student’s t-test. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001 (n = 3).