Acta Entomologica Sinica ›› 2022, Vol. 65 ›› Issue (5): 548-557.doi: 10.16380/j.kcxb.2022.05.002

• RESEARCH PAPERS • Previous Articles     Next Articles

Expression and functional analysis of lncRNA63 and its co-expression gene Hr3 in Bombyx mori

WANG Bo, LI Miao-Miao, WANG Jing-Ya, HU Xiao-Yue, KAN Yun-Chao, QIAO Hui-Li*   

  1.  (Henan Key Laboratory of Insect Biology in Funiu Mountain, College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang, Henan 473061, China)
  • Online:2022-05-20 Published:2022-05-08

Abstract: 【Aim】 This study aims to analyze the expression and function of a new long non-coding RNA (lncRNA) and its co-expression gene in the domesticated silkworm, Bombyx mori, so as to further clarify the molecular mechanism of lncRNA in regulating the metamorphosis and development of B. mori. 【Methods】 Based on the transcriptome sequencing data of the fat body of B. mori, we found that the expression of lncRNA63 was significantly up-regulated in the fat body of the early 5th instar larvae treated with 20-hydroxyecdysone (20E)(2 μg/μL) and the hormone receptor gene Hr3 of B. mori was co-expressed with lncRNA63. qRT-PCR was used to analyze the expression profiles of lncRNA63 and Hr3 in different developmental stages (4th-5th instar larva, pupa and adult), different tissues (head, integument, hemolymph, midgut, Malpighian tubules, fat body, silk gland, testis and ovary) of the 5th instar larvae and the fat body of the 5th instar larvae of B. mori treated with 20E (2 μg/μL) for 2, 6, 12 and 24 h. The localization of lncRNA63 in BmN cells of B. mori was detected by nucleocytoplasmic separation and in situ hybridization. The expression of lncRNA63 in the 5th instar larvae of B. mori was interfered by dsRNA, and the effects of RNA interference of lncRNA63 on the expression of Hr3 in the head, silk gland, fat body and integument were detected using qRT-PCR. 【Results】 The qRT-PCR results showed that there was an expression peak of lncRNA63 and Hr3 at the end of pupal stage before eclosion, and both are highly expressed in the head, integument and silk gland of the 5th instar larvae of B. mori. The expression of lncRNA63 and Hr3 in the fat body of the 5th instar larvae of B. mori treated with 2 μg/μL 20E for 2, 6 and 12 h was significantly up-regulated as compared to the control group (ethanol treatment group) and decreased to the level of the control group at 24 h after treatment, with their expression trend being completely consistent. LncRNA63 was mainly localized in the nucleus of BmN cells, but 20E induced the nucleocytoplasmic translocation of lncRNA63. After interference of the expression of lncRNA63 with dsRNA, the expression of Hr3 was significantly down-regulated. 【Conclusion】 20E can not only regulate the expression of lncRNA63, but also induce the nucleocytoplasmic translocation of lncRNA63. LncRNA63 may participate in the metamorphosis and development of B. mori by regulating the expression of its co-expressed gene Hr3.

Key words: Bombyx mori, long non-coding RNA, fat body, expression analysis, metamorphosis and development