Treatment of Drosophila salivary glands with a mild detergent, digitonin, activates puffing at 35 chromosome loci. These digitonin-activated puffs include all of the nine heat-shock puffs known in D. melanogaster. Here we show that the activation of heat-shock genes, but not of other digitoninstimulated puffs, is repressed in salivary glands which have been subjected to and have recovered from heat shock before being treated with digitonin. The findings indicate that, (a) the activation of heat-shock genes by digitonin, as that by temperature elevation, is self-regulated by the heat-shock proteins (HSPs). (b) the gene repressive activity of HSPs is heat-shock-gene specific, and (c) the repression mechanism of heat-shock genes by HSPs is resistant to digitonin, in contrast to that the suppression of heat-shock genes is prevented by the detergent in non-heat-shocked salivary glands. The selective repression of heat-shock genes in preheated salivary glands suggests that the heat-shock genes and other digitonin-activated genes may be controlled by a different mechanism(s).