Heat sensitivity of bleomycin-sensitive CHO derivatives is not due to improper initialization of heat shock response

We have investigated whether differences in the heat shock response exist between CHO and three bleomycin-sensitive, heat-sensitive CHO derivatives. The binding of heat shock factor (HSF) in response to heat and varying concentrations of bleomycin in the four cell lines was examined using a gel shift assay and a synthetic heat shock element (HSE). Heat (45 degrees C, 10 min) and exposure to 1 micrograms/ml bleomycin for 1 h at 37 degrees C induced similar levels of HSF binding in all four cell lines. We also examined if bleomycin dose and the length of recovery from bleomycin treatment affected the induction of HSF binding. The level of activated HSF binding to HSE was higher in cells treated with low doses (1 ng/ml) of bleomycin than in cells treated with 1 or 25 micrograms/ml bleomycin. The amount of activated HSF was directly proportional to the time elapsed since bleomycin treatment. Our results therefore indicate no difference between CHO and its bleomycin-sensitive derivatives in the ability to initiate the heat shock response as determined by the production of activated HSF in response to either heat or bleomycin. We conclude that the intrinsic thermosensitivity of these cell lines is not related to the early response to heat shock, but either occurs later in the pathway or is unrelated to events after heating.