1995 Volume 118 Issue 5 Pages 1021-1029
We investigated the regulatory expression of the HSP70-1 gene in rat liver after whole body hyperthermia (rectal temperature of 42°C for 30min). After heat treatment, HSP70-1 mRNA showed a peak at 1 h, and its protein product began to increase at 3h. Under these conditions, we identified cis-acting elements involved in the regulatory expression of the HSP70-1 gene by in vivo genomic footprinting. We observed that the second heat shock element (HSE-2; -201 to -178) was clearly protected from methylation by dimethyl sulfate in parallel with the increase in HSP70-1 mRNA. On the contrary, the most proximal HSE-1 (-120 to -101) was protected only slightly. Furthermore, the flanking regions of HSE-2, that is, an NF-xB-like element on the upstream side and a GC box on the downstream side were also protected simultaneously in accordance with the induced expression of the HSP70-1 gene. Gel mobility shift assays with nuclear extracts from rat liver at 30min after heat treatment showed maximal DNA binding activity equally with oligonucleotides containing HSE-1 and HSE-2, respectively, and also demonstrated the participation of heat shock factor-1 (HSF1) in both cases. These results suggest that, in addition to HSF1, other regulatory factors might be involved in the dominant protection of HSE-2 in the promoter of the HSP70-1 gene in rat liver after whole body hyperthermia.
