斑翅聪聪虫（Plutella xylostella）中，游戏体特异性因子1对雄性生育能力和转座子稳态至关重要。

Abstract

Simple SummaryThe Gametocyte-specific factor 1 (Gtsf1) gene is known to modulate gonadal development and sex determination via the piRNA pathway in limited insect species, while its biological function in the destructive cruciferous pestPlutella xylostellaremains unknown. Here, we functionally characterizedPxGtsf1in diamondback moths using CRISPR/Cas9-mediated gene knockout.PxGtsf1was specifically enriched in male pupae and testes, and its deletion caused defective male testicular morphology and sperm structure, further reducing mating fecundity and egg hatching efficiency. No obvious abnormalities were observed in the female reproductive system and sex determination process afterPxGtsf1knockout. Molecular verification confirmed that PxGtsf1 interacts with PxPiwi to maintain testicular transposon homeostasis, without influencing piRNA biosynthesis. This study clarifies a novel sex-specific function of Gtsf1 in regulating lepidopteran male fertility, which updates the understanding of insect Gtsf1 functional diversity.Gametocyte-specific factor 1 (Gtsf1) plays a pivotal role in the gonadal development by transposon silencing via the piRNA pathway inDrosophila melanogasterandBombyx mori, and is implicated in the female sex determination inB. mori. However, its function remains elusive in other insect species, includingPlutella xylostella, a highly destructive pest in cruciferous crops. In this study, we characterized PxGtsf1, encoding a 27.84 kDa protein containing two conserved U11-48K-like CHHC zinc-finger domains.PxGtsf1was predominantly expressed in male pupae, males and testes. Using CRISPR/Cas9 technique, we successfully generated a homozygousPxGtsf1-knockout strain (△PxGtsf1) with 11 bp deletion. △PxGtsf1 females exhibited normal morphology in terms of external genitalia, body weight, ovariole morphology, ovariole length, oocytes number, and internal ovarian structure, and maintained the expression levels of genes related to female sex determination and fertil