Real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to characterize the effects of juvenile hormone (JH) on Epac (Exchange Protein directly Activated by Cyclic AMP; NM_001103732), a guanine nucleotide exchange factor for Rap1 in Drosophila S2 cells. JH treatment led to a rapid, dose-dependent increase in Epac relative expression ratio (RER) when compared to treatment with methyl linoleate (MLA) that lacks biological activity. The minimal level of hormone needed to elicit a response was 100 ng/ml. Time-course studies indicated a significant rise in the RER 1h after treatment. S2 cells were challenged with 20-hydroxyecdysone and a series of compounds similar in structure to JH to determine the specificity of the response. Methoprene and JH III displayed the greatest increases in RER. Late third instar (96 h) Drosophila were exposed to diet containing methoprene (500 ng/g diet); significantly higher RERs for Epac were observed 12h after exposure. JH had no effect on Epac RERs in the human cell line HEK-293.
