The content of JH-esterase was assayed by radial immunodiffusion in Drosophila virilis pupae under normal conditions and under the effects of extreme factors. It was found that JH-esterase content is the same (not different from the control) in pupae showing a high activity of the enzyme and in those not showing it. These data are evidence for a gene controlling JH-esterase activity. It was also shown that a regulatory factor converts inactive into active JH-esterase when homogenates of pupae, with active and inactive forms, were mixed and incubated together. It was demonstrated that the source of the activating factor is the larval brain. Sublines 147-R and 147-I were produced by introducing the second chromosome pair of stocks 103 and 101, which are heat resistant, into the genome of individuals of stock 147, which is heat sensitive. Sublines 160-III, 160-IV, 160-V, and 160-VI were produced by introducing the third, fourth, fifth, and sixth chromosome pairs of stock 147 into the genome of stock 160S, which is heat-resistant. The results of analysis of JH-esterase activity and the viability of individuals of these sublines at high temperatures indicated that the gene regulating the activity of JH-esterase is located in the sixth chromosome of D. virilis.