title: Determining the involvement of two aminopeptidase Ns in the resistance of Plutella xylostella to the Bt toxin Cry1Ac: cloning and study of in vitro function.
abstract: The cloning, expression in vitro, and characterization of two aminopeptidase Ns (APN5s and APN2s) isolated from the midgut of Cry1Ac-resistant (R) and susceptible (S) strains of Plutella xylostella larvae are presented in this paper. The deduced amino acid sequences of APN5s included C-terminal GPI-modification sites, the gluzincin aminopeptidase motif GATEN, and three N-glycosylated sites; those of APN2s had no GPI-modification sites, had gluzincin aminopeptidase motif GAMEN, and had four N-glycosylated sites. O-glycosylated sites were not predicted for either APN. Because APN2R and APN2S cDNAs contained the same nucleotides, only full-length cDNAs encoding APN5R and APN5S were expressed in Trichoplusia ni cells. Far-Western blotting showed that the expressed receptor APN5 bound to the Cry1Ac toxin. An enzyme-specific activity experiment also showed that APN5 genes were expressed in T. ni cells. ELISA revealed no differences in the binding of expression proteins from the resistant and susceptible strain with Cry1Ac.
authors.affiliation: Department of Plant Protection, Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, China.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Chang
authors.forename: Qingjun
authors.initials: Q
authors.lastname: Wu
authors.forename: Shaoli
authors.initials: S
authors.lastname: Wang
authors.forename: Ran
authors.initials: R
authors.lastname: Wang
authors.forename: Zhongxia
authors.initials: Z
authors.lastname: Yang
authors.forename: Defeng
authors.initials: D
authors.lastname: Chen
authors.forename: Xiaoguo
authors.initials: X
authors.lastname: Jiao
authors.forename: Zhenchuan
authors.initials: Z
authors.lastname: Mao
authors.forename: Youjun
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry*; D001483:Base Sequence; D018826:CD13 Antigens / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004731:Endotoxins / Q000737:chemistry*; D006460:Hemolysin Proteins / Q000737:chemistry*; D019476:Insect Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D007305:Insecticide Resistance; D007306:Insecticides / Q000737:chemistry*; D007700:Kinetics; D007814:Larva / Q000201:enzymology*; D008969:Molecular Sequence Data; D009036:Moths / Q000201:enzymology*; D010572:Pest Control, Biological; D010802:Phylogeny; D030821:Plants, Genetically Modified; D011485:Protein Binding; D011994:Recombinant Proteins; D017422:Sequence Analysis, DNA
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D019476:Insect Proteins; D007306:Insecticides; D011994:Recombinant Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D018826:CD13 Antigens