Identification and Characterization of MicroRNAs in Gonads of Helicoverpa armigera (Lepidoptera: Noctuidae)

SIMPLE SUMMARY: For most insects, the development of the testis and ovary directly determines their reproductive ability. The cotton bollworm, Helicoverpa armigera (Hübner), is a polyphagous crop pest of the Lepidoptera Noctuidae. Owing to its broad range of host plants and strong fertility, H. armigera causes huge economic losses to agricultural production. Acting as a type of post-transcriptional regulatory factor, miRNAs participate in the gonadal development and reproductive regulation of H. arimgera. Our study uses H. armigera as a research object to identify and characterize the miRNAs and study their potential functions in the testis and ovary of this destructive crop pest. A total of 7,592,150 and 8,815,237 clean reads were obtained by constructing small RNA libraries of the testis and ovary, respectively. Length distribution analysis showed that the main types of small RNAs in the testis and ovary were different. Among the 74 known miRNAs, 60 miRNAs existed in the ovary, and 72 existed in the testis. Gene Ontology (GO) and KEGG pathway analyses indicated that the 8 gonad-biased differentially expressed miRNAs (miR-989a, miR-263-5p, miR-34, miR-2763, miR-998, miR-2c, miR-2765, and miR-252a-5p) had many target transcripts involved in the reproduction process. ABSTRACT: The high fecundity of the most destructive pest Helicoverpa armigera and its great resistance risk to insecticides and Bt crops make the reproductive-destruction-based control of this pest extremely appealing. To find suitable targets for disruption of its reproduction, we observed the testis and ovary development of H. armigera and conducted deep sequencing of the ovary and testis small RNAs of H. armigera and quantitative RT-PCR (RT-qPCR) validation to identify reproduction-related micro RNAs (miRNAs). A total of 7,592,150 and 8,815,237 clean reads were obtained from the testis and ovary tissue, respectively. After further analysis, we obtained 173 novel and 74 known miRNAs from the two libraries. Among the 74 known miRNAs, 60 miRNAs existed in the ovary and 72 existed in the testis. Further RT-qPCR validation of 5 miRNAs from the ovary and 6 miRNAs from the testis confirmed 8 of them were indeed ovary- (miR-989a, miR-263-5p, miR-34) or testis-biased (miR-2763, miR-998, miR-2c, miR-2765, miR-252a-5p). The 8 ovary- or testis-biased miRNAs had a total of 30,172 putative non-redundant target transcripts, as predicted by miRanda and RNAhybrid. Many of these target transcripts are assigned to reproduction-related GO terms (e.g., oocyte maturation, vitellogenesis, spermatogenesis) and are members of multiple reproduction-related KEGG pathways, such as the JAK-STAT signaling pathway, oocyte meiosis, the insulin signaling pathway, and insect hormone biosynthesis. These results suggest that the 8 gonad-biased miRNAs play important roles in reproduction and may be used as the targets for the development of reproductive-destruction-based control of H. armigera and, possibly, other lepidopteran pests.