Aphis craccivora (Hemiptera: Aphididae) synthesizes juvenile hormone III via a pathway involving epoxidation followed by esterification, potentially providing an epoxidation active site for the synthesis of juvenile hormone SB3.

Juvenile hormones (JHs) play a crucial role in regulating development and reproduction in insects. Most insects predominantly synthesize JH III, which typically involves esterification followed by epoxidation, lepidopteran insects use a pathway of epoxidation followed by esterification. Although hemipteran insects have JH III and JH skipped bisepoxide III (JH SB3), the synthesis pathway and key epoxidases remain unclear. This study was conducted on Aphis craccivora, and demonstrated that corpora allata, microsomes, Ac-CYP15C1, and Ac-JHAMT catalyze JH III production in vitro, establishing the pathway of epoxidation followed by esterification. These findings were further confirmed through RNA interference and molecular docking. The presence of JH III and JH SB3 in A. craccivora was identified, and their synthesis pathway was elucidated as follows: Ac-CYP15C1 oxidizes farnesic acid to JH A, followed by methylation to JH III by Ac-JHAMT, possibly providing an epoxidation site on the second carbon for JH SB3. This alteration may significantly contribute to the differentiation and functional diversification of JH types in insects.