title: Transgenic Clustered Regularly Interspaced Short Palindromic Repeat/Cas9-Mediated Viral Gene Targeting for Antiviral Therapy of Bombyx mori Nucleopolyhedrovirus.
abstract: We developed a novel antiviral strategy by combining transposon-based transgenesis and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system for the direct cleavage of Bombyx mori nucleopolyhedrovirus (BmNPV) genome DNA to promote virus clearance in silkworms. We demonstrate that transgenic silkworms constitutively expressing Cas9 and guide RNAs targeting the BmNPVimmediate early-1(ie-1) andme53genes effectively induce target-specific cleavage and subsequent mutagenesis, especially large (∼7-kbp) segment deletions in BmNPV genomes, and thus exhibit robust suppression of BmNPV proliferation. Transgenic animals exhibited higher and inheritable resistance to BmNPV infection than wild-type animals. Our approach will not only contribute to modern sericulture but also shed light on future antiviral therapy.IMPORTANCEPathogen genome targeting has shown its potential in antiviral research. However, transgenic CRISPR/Cas9 system-mediated viral genome targeting has not been reported as an antiviral strategy in a natural animal host of a virus. Our data provide an effective approach against BmNPV infection in a real-world biological system and demonstrate the potential of transgenic CRISPR/Cas9 systems in antiviral research in other species.
authors.affiliation: Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.|University of Chinese Academy of Sciences, Beijing, China.
authors.forename: Shuqing
authors.initials: S
authors.lastname: Chen
authors.affiliation: Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang, Jiangsu, China.
authors.forename: Chengxiang
authors.initials: C
authors.lastname: Hou
authors.affiliation: Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Honglun
authors.initials: H
authors.lastname: Bi
authors.affiliation: Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Yueqiang
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.|University of Chinese Academy of Sciences, Beijing, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Xu
authors.affiliation: Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang, Jiangsu, China.
authors.forename: Muwang
authors.initials: M
authors.lastname: Li
authors.affiliation: Departments of Microbiology & Molecular Genetics and Molecular Biology & Biochemistry, University of California, Irvine, Irvine, California, USA.
authors.forename: Anthony A
authors.initials: AA
authors.lastname: James
authors.affiliation: Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China yphuang@sibs.ac.cn ajtan01@sibs.ac.cn.
authors.forename: Yongping
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China yphuang@sibs.ac.cn ajtan01@sibs.ac.cn.
authors.forename: Anjiang
authors.initials: A
authors.lastname: Tan
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D000998:Antiviral Agents / Q000378:metabolism*; D016367:Baculoviridae / Q000235:genetics* / Q000254:growth & development*; D012831:Bombyx / Q000276:immunology* / Q000502:physiology / Q000821:virology; D064113:CRISPR-Cas Systems; D004279:DNA, Viral / Q000378:metabolism*; D060467:Disease Resistance; D050296:Microbial Viability; D016296:Mutagenesis; D017384:Sequence Deletion
chemical_list: D000998:Antiviral Agents; D004279:DNA, Viral
keywords: Bombyx mori nucleopolyhedrovirus; CRISPR/Cas9; antiviral therapy; transgenic silkworm