关键词: 葱蝇; FOXO1, 滞育, 超氧化物歧化酶, 蛹期, RNAi

Abstract: 【Aim】 The aim of this study is to investigate the role of DaFOXO1 in the regulation of superoxide dismutase (SOD) gene expression and the developmental duration of summer diapause pupae of the onion fly, Delia antiqua. 【Methods】 Genes encoding copper-zinc superoxide dismutase (DaCu/Zn SOD) and manganese superoxide dismutase (DaMn SOD), downstream genes of DaFOXO1, were identified from the transcriptome data of D. antiqua. The features of amino acid sequences, subcellular localization and phylogeny of DaCu/Zn SOD and DaMn SOD of D. antiqua were analyzed by using bioinformatic tools. The expression patterns of DaFOXO1, DaCu/Zn SOD, and DaMn SOD at different developmental stages of summer diapause pupae of D. antiqua were determined by qRT-PCR. The effect of knocking down DaFOXO1 by RNAi on the expressions of DaCu/Zn SOD and DaMn SOD, the activity variations of DaCu/Zn SOD and DaMn SOD and the developmental duration of summer diapause pupae of D. antiqua were further analyzed. 【Results】 The open reading frame (ORF) of DaCu/Zn SOD (GenBank accession no. KR072551) of D. antiqua is 459 bp in length and encodes 153 amino acids with a predicted molecular weight (MW) of 22.4 kD and an ioselectric point (pI) of 6.44, which belongs to a cytoplasmic Cu/Zn SOD. The ORF of DaMn SOD (GenBank accession no. KR072549) is 648 bp in length and encodes 216 amino acids with a predicted MW of 24.4 kD and a pI of 8.85, which belongs to mitochondrial Mn SOD. Amino acid sequence alignment revealed that DaCu/Zn SOD and DaMn SOD share 75%-94% identity with their homologues from other 10 species of Diptera, and contain typical SOD family domains. Phylogenetic analysis showed that they formed a robust phylogenetic branch with their homologues in Lucilia cuprina. qRT-PCR analysis revealed that the expression levels of DaFOXO1 were higher at the pre-diapause and post-diapause stages, but lower at the diapause stage. Higher expression of DaCu/Zn occurred at the diapause stage and post-diapause stage. However, the highest expression level of DaMn SOD was detected at the prediapause stage and diapause stage, and followed by at post-diapause stage. Knockdown of DaFOXO1 by RNAi significantly decreased the expression levels of DaCu/Zn SOD and DaMn SOD, and the activities of their corresponding enzymes, leading to a significantly extended duration of summer diapause pupae. 【Conclusion】 The results suggest that DaCu/Zn SOD and DaMn SOD are important members of FOXO1 signaling network, and DaFOXO1 plays an important role in the regulation of the duration of summer diapause pupae of D. antiqua.

Key words: Delia antiqua, FOXO1, diapause, superoxide dismutase, pupal duration, RNAi