FIGURE 4. CHIP orchestrates both stress response and recovery processes.

a, Mouse fibroblasts were heat shocked (HS) at 42 °C for 10 min, and aliquots were collected every 4 h during recovery at 37 °C. Hsp70 levels were determined by immunoblotting. Values are means s.d. b, CHIP^-/- cells were infected with AdV expressing GFP (squares), CHIP (diamonds) or CHIP(K30A) (triangles). Infected cells were heat shocked at 42 °C for 10 min, and aliquots were collected every 2 h during recovery at 37 °C. Hsp70 levels were determined by immunoblotting and quantified by densitometry. c, The same AdV-infected CHIP^-/- cells as in b were heat shocked at 42 °C for 10 min, and radiolabelled with [³⁵S]methionine for 30 min at 3 h after HS and chased with unlabelled methionine for up to 6 h. Immunoprecipitation was performed with a Hsp70-specific antibody and quantified with a PhosphorImager. Symbols are as in b. Values are means s.d. d, HEK-293 cells were transfected with plasmids as indicated with 0, 0.1 and 0.5 g per well in a six-well plate. Hsp70 levels were determined by immunoblotting. e, Transfected HEK-293 cells were heat shocked 42 °C for 5 min, and aliquots were collected every 2 h during recovery at 37 °C. Hsp70 levels were determined by immunoblotting.