title: Modification of Cry4Aa toward Improved Toxin Processing in the Gut of the Pea Aphid, Acyrthosiphon pisum.
abstract: Aphids are sap-sucking insects (order: Hemiptera) that cause extensive damage to a wide range of agricultural crops. Our goal was to optimize a naturally occurring insecticidal crystalline (Cry) toxins produced by the soil-dwelling bacterium Bacillus thuringiensis for use against the pea aphid, Acyrthosiphon pisum. On the basis that activation of the Cry4Aa toxin is a rate-limiting factor contributing to the relatively low aphicidal activity of this toxin, we introduced cathepsin L and cathepsin B cleavage sites into Cry4Aa for rapid activation in the aphid gut environment. Incubation of modified Cry4Aa and aphid proteases in vitro demonstrated enhanced processing of the toxin into the active form for some of the modified constructs relative to non-modified Cry4Aa. Aphids fed artificial diet with toxin at a final concentration of 125 μg/ml showed enhanced mortality after two days for one of the four modified constructs. Although only modest toxin improvement was achieved by use of this strategy, such specific toxin modifications designed to overcome factors that limit aphid toxicity could be applied toward managing aphid populations via transgenic plant resistance.
authors.affiliation: Department of Entomology, Iowa State University, Ames, Iowa, United States of America.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Rausch
authors.affiliation: Department of Entomology, Iowa State University, Ames, Iowa, United States of America.
authors.forename: Nanasaheb P
authors.initials: NP
authors.lastname: Chougule
authors.affiliation: Department of Entomology, Iowa State University, Ames, Iowa, United States of America.
authors.forename: Benjamin R
authors.initials: BR
authors.lastname: Deist
authors.affiliation: Department of Entomology, Iowa State University, Ames, Iowa, United States of America.
authors.forename: Bryony C
authors.initials: BC
authors.lastname: Bonning
mesh_terms: D000818:Animals; D001042:Aphids / Q000187:drug effects / Q000378:metabolism*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000633:toxicity*; D002401:Cathepsin B / Q000378:metabolism; D056668:Cathepsin L / Q000378:metabolism; D004731:Endotoxins / Q000633:toxicity*; D041981:Gastrointestinal Tract / Q000187:drug effects / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000633:toxicity*; D018532:Pisum sativum / Q000469:parasitology*; D015202:Protein Engineering; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D018675:Toxicity Tests; D014357:Trypsin / Q000378:metabolism
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D014357:Trypsin; D002401:Cathepsin B; D056668:Cathepsin L