FIGURE 5.
HSP70 regulates NOD2's stability in the cell. A, HEK293T-NOD2Myc/Tet-op cells were incubated with 100 μm GGA or 100 μm KNK437 or DMSO for 2 h, and lysates were collected after cycloheximide treatment during the indicated time intervals. Equal amounts of lysates were subjected to Western blot and probed using rabbit anti-Myc antibody (1:1000) or rabbit anti-HSP70 antibody (1:1000). β-Actin (1:1000) was used as a loading control. Relative amount of NOD2Myc to β-actin was plotted as the means ± S.D. from three independent experiments. B, cell extracts from HEK293T-NOD2Myc/Tet-op cells, transfected with HSP70 or control vector, and treated with cycloheximide. Cell lysates were collected during the indicated time intervals and subjected to Western blot and probed using rabbit anti-Myc antibody (1:1000) or rabbit anti-HSP70 antibody (1:1000). β-Actin (1:1000) was used as a loading control. Relative amount of NOD2Myc to β-actin was plotted as the means ± S.D. from three independent experiments. Western blots were performed on separate cell lysates at least three times. Using the replicates, the bands were quantified using ImageLabTM.