Abstract:  【Aim】 Insect detoxification enzymes play important roles in insect adaptation to secondary plant substances. Studying the miRNAs, which regulate the expression of detoxification enzyme genes, will help to identify the target for pest control. This study aims to investigate the regulation of Sli-miR-34-5p on the detoxification enzyme glutathione S-transferase (GST) gene involved in the adaptation to secondary plant substances in Spodoptera litura. 【Methods】 The expression level of Sli-miR-34-5p in the midgut of the day-1 5th instar larvae of S. litura fed with Brassica juncea was determined by qPCR. After the overexpression of Sli-miR-34-5p mimic or inhibitor, the expression of the known or presumable target gene of Sli-miR-34-5p at the mRNA and protein levels in S. litura larvae was determined by qPCR and Western blot, respectively. The regulation of Sli-miR-34-5p on its target gene was verified by dual luciferase reporter system. The possible action mechanism of Sli-miR-34-5p on target genes was investigated by predicting the target genes of Sli-miR-34-5p with bioinformatics method.【Results】 The results showed that the expression of Sli-miR-34-5p was up-regulated in the midgut of the day-1 5th instar larvae of S. litura fed with B. juncea for 24 and 48 h. The injection of Sli-miR-34-5p mimic into the hemolymph of the day-1 5th instar larvae of S. litura resulted in the up-regulation of the mRNA level of SlGSTe1 in the midgut of B. juncea-fed larvae. Injection of Sli-miR-34-5p inhibitor into the hemolymph significantly downregulated the protein level of SlGSTE1. In the S. litura Spli-221 cell line treated with indole-3-carbinol, a secondary substance of B. juncea, the expression levels of both Sli-miR-34-5p pression of SlGSTe1 enhanced after Sli-miR-34-5p mimic was added to the cell line. However, the assay of dual luciferase activity showed that Sli-miR-34-5p did not directly bind to SlGSTe1Sli-miR-34-5p nformatics analysis showed that SlGSTe1 may be not the direct target gene of Sli-miR-34-5p. 【Conclusion】 The results suggest that Sli-miR-34-5p may be involved in the response of S. litura to secondary substances of host plants by enhancing the expression of the glutathione S-transferase gene SlGSTe1.

Key words: Spodoptera litura; miRNA, Sli-miR-34-5p, detoxification enzyme, expression regulation, secondary plant substances