Influence of proteasome and redox state on heat shock-induced activation of stress kinases, AP-1 and HSF

AbstractWe studied the pattern of activation of stress kinases and of transcription factors activator protein-1 (AP-1) and heat shock factor (HSF) in FAO cells by combining two treatments, i.e. heating (42°C for 1 h) and proteasome inhibition, each known to cause cellular heat shock response. The co-treatment heat shock (HS) and proteasome inhibitor (a peptidyl aldehyde or lactacystin) showed cumulative effects on the intensity and duration of activation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) at the end of the HS period and during recovery. Similarly, the thiol-reducing agents N-(2-mercaptoethyl)-1,3-diaminopropane and dithiothreitol strongly activated both JNK and p38 MAPK in cells undergoing HS. AP-1 DNA binding activity in response to proteasome inhibitors was so strong that it shadowed the stimulatory effect of HS in the combined treatment, but lactacystin, which is the most potent and specific proteasome inhibitor, decreased the binding late during recovery from HS. Thiol-reducing agents prevented AP-1 DNA binding induced by HS. The combined HS/proteasome inhibitors or HS/thiol-reducing agents treatments cooperatively activated HSF DNA binding. Expression of collagenase I and hsp 70 mRNAs reflects the different behavior of AP-1 and HSF transcription factors in cells exposed to HS and proteasome inhibition. The data seem to indicate that JNK and p38 MAPK activations are not necessarily coupled to DNA binding of AP-1, which can be either increased or inhibited when these kinases are activated. AP-1 and HSF show opposite patterns of response to HS in the presence of proteasome inhibitors or reducing agents