Figure 4.

SIRT1-mediated HDAC1 deacetylation decreases during heat shock. (a) HCT116 cells were subjected to heat shock at 42 °C for 1 h and then recovered at 37 °C for an indicated time. SIRT1 protein level was tested by western blot. (b) HCT116 cells were treated with or without 10 μM MG132 for 2 h and then subjected to heat shock. Cells were recovered at 37 °C for 2 h. Upper panel shows representative western blots and lower panel shows quantification of SIRT1 levels normalized to β-actin controls. (c) HCT116 cells were treated with or without 10 μM CHX for 2 h and then subjected to heat shock. Cells were rescued at 37 °C for 2 h. Upper panel shows representative western blots and lower panel shows quantification of SIRT1 levels normalized to β-actin controls. (d) Flag-SIRT1 was overexpressed in HCT116 cells and then the cells were subjected to heat shock at 42 °C for 1 h and recovered for indicated times. The cell extracts were harvested and subjected to western blot with indicated antibodies. Upper panel shows representative western blots and lower panel shows quantification of ac-HDAC1 levels normalized to β-actin controls. (e) SIRT1 was knocked down with shRNA in HCT116 cells, subjected to heat shock at 42 °C for 1 h, and recovered at 37 °C for 2 h. Upper panel shows representative Western blots and lower panel shows quantification of ac-HDAC1 levels normalized to β-actin controls