title: Depletion of juvenile hormone esterase extends larval growth in Bombyx mori.
abstract: Two major hormones, juvenile hormone (JH) and 20-hydroxyecdysone (20E), regulate insect growth and development according to their precisely coordinated titres, which are controlled by both biosynthesis and degradation pathways. Juvenile hormone esterase (JHE) is the primary JH-specific degradation enzyme that plays a key role in regulating JH titers, along with JH epoxide hydrolase (JHEH) and JH diol kinase (JHDK). In the current study, a loss-of-function analysis of JHE in the silkworm, Bombyx mori, was performed by targeted gene disruption using the transgenic CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/RNA-guided Cas9 nucleases) system. Depletion of B. mori JHE (BmJHE) resulted in the extension of larval stages, especially the penultimate and ultimate larval stages, without deleterious effects to silkworm physiology. The expression of JHEH and JHDK was upregulated in mutant animals, indicating the existence of complementary routes in the JH metabolism pathway in which inactivation of one enzyme will activate other enzymes. RNA-Seq analysis of mutant animals revealed that genes involved in protein processing in the endoplasmic reticulum and in amino acid metabolism were affected by BmJHE depletion. Depletion of JHE and subsequent delayed JH metabolism activated genes in the TOR pathway, which are ultimately responsible for extending larval growth. The transgenic Cas9 system used in the current study provides a promising approach for analysing the actions of JH, especially in nondrosophilid insects. Furthermore, prolonging larval stages produced larger larvae and cocoons, which is greatly beneficial to silk production.
authors.affiliation: School of Life Science, East China Normal University, Shanghai 200062, China; Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Zhongjie
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Liu
authors.affiliation: Insect Growth Regulation Research Unit, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Shiotsuki
authors.affiliation: College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China.
authors.forename: Zhisheng
authors.initials: Z
authors.lastname: Wang
authors.affiliation: School of Life Science, East China Normal University, Shanghai 200062, China.
authors.forename: Xia
authors.initials: X
authors.lastname: Xu
authors.affiliation: Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Yongping
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, Jiangsu, China.
authors.forename: Muwang
authors.initials: M
authors.lastname: Li
authors.affiliation: School of Life Science, East China Normal University, Shanghai 200062, China. Electronic address: kaili@bio.ecnu.edu.cn.
authors.forename: Kai
authors.initials: K
authors.lastname: Li
authors.affiliation: Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China. Electronic address: ajtan01@sibs.ac.cn.
authors.forename: Anjiang
authors.initials: A
authors.lastname: Tan
mesh_terms: D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000201:enzymology / Q000235:genetics / Q000254:growth & development*; D064113:CRISPR-Cas Systems; D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D005260:Female; D007328:Insulin / Q000378:metabolism*; D007814:Larva / Q000254:growth & development; D008297:Male; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D060888:Real-Time Polymerase Chain Reaction; D017423:Sequence Analysis, RNA; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: D007328:Insulin; D058570:TOR Serine-Threonine Kinases; D002265:Carboxylic Ester Hydrolases; C038003:juvenile hormone esterase
keywords: Bombyx mori; CRISPR/Cas9; Juvenile hormone esterase