A CONSTITUTIVE HEAT-SHOCK ELEMENT-BINDING FACTOR IS IMMUNOLOGICALLY IDENTICAL TO THE KU AUTOANTIGEN

Analysis of the heat shock element (HSE)-binding proteins in extracts of rodent cells, during heat shock and their post-heat shock recovery, indicates that the regulation of heat shock response involves a constitutive HSE-binding factor (CHEF), in addition to the heat-inducible heat shock factor HSF1. We purified the CHEF to apparent homogeneity from HeLa cells using column chromatographic techniques including an HSE oligonucleotide affinity column. The purified CHEF consists of two polypeptides with apparent molecular masses of 70 and 86 kDa. Immunoblot and gel mobility shift analysis verify that CHEF is identical or closely related to the Ku autoantigen. The DNA binding characteristics of CHEF to double-stranded or single-stranded DNA are similar to that of Ku autoantigen. In gel mobility shift analysis using purified CHEF and recombinant human HSF1, CHEF competes with HSF1 for the binding of DNA sequences containing HSEs in vitro. Furthermore, when Rat-1 cells were co-transfected with human Ku expression vectors and the hsp70-promoter-driven luciferase reporter gene, thermal induction of luciferase is significantly suppressed relative to cells transfected with only the hsp70-luciferase construct, These data suggest a role of CHEF (or Ku protein) in the regulation of heat response in vivo.