title: RNAi induced knockdown of a cadherin-like protein (EF531715) does not affect toxicity of Cry34/35Ab1 or Cry3Aa to Diabrotica virgifera virgifera larvae (Coleoptera: Chrysomelidae).
abstract: The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, is an important maize pest throughout most of the U.S. Corn Belt. Bacillus thuringiensis (Bt) insecticidal proteins including modified Cry3Aa and Cry34/35Ab1 have been expressed in transgenic maize to protect against WCR feeding damage. To date, there is limited information regarding the WCR midgut target sites for these proteins. In this study, we examined whether a cadherin-like gene from Diabrotica virgifera virgifera (DvvCad; GenBank accession # EF531715) associated with WCR larval midgut tissue is necessary for Cry3Aa or Cry34/35Ab1 toxicity. Experiments were designed to examine the sensitivity of WCR to trypsin activated Cry3Aa and Cry34/35Ab1 after oral feeding of the DvvCad dsRNA to knockdown gene expression. Quantitative real-time PCR confirmed that DvvCad mRNA transcript levels were reduced in larvae treated with cadherin dsRNA. Relative cadherin expression by immunoblot analysis and nano-liquid chromatography - mass spectrometry (nanoLC-MS) of WCR neonate brush border membrane vesicle (BBMV) preparations exposed to DvvCad dsRNA confirmed reduced cadherin expression when compared to BBMV from untreated larvae. However, the larval mortality and growth inhibition of WCR neonates exposed to cadherin dsRNA for two days followed by feeding exposure to either Cry3Aa or Cry34/35Ab1 for four days was not significantly different to that observed in insects exposed to either Cry3Aa or Cry34/35Ab1 alone. In combination, these results suggest that cadherin is unlikely to be involved in the toxicity of Cry3Aa or Cry34/35Ab1 to WCR.
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: Sek Yee
authors.initials: SY
authors.lastname: Tan
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: Murugesan
authors.initials: M
authors.lastname: Rangasamy
authors.affiliation: University of Nebraska-Lincoln, 103 Entomology Hall, Lincoln, NE 68583, United States.
authors.forename: Haichuan
authors.initials: H
authors.lastname: Wang
authors.affiliation: University of Nebraska-Lincoln, 103 Entomology Hall, Lincoln, NE 68583, United States.
authors.forename: Ana María
authors.initials: AM
authors.lastname: Vélez
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: James
authors.initials: J
authors.lastname: Hasler
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: David
authors.initials: D
authors.lastname: McCaskill
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: Tao
authors.initials: T
authors.lastname: Xu
authors.affiliation: University of Nebraska-Lincoln, 103 Entomology Hall, Lincoln, NE 68583, United States.
authors.forename: Hong
authors.initials: H
authors.lastname: Chen
authors.affiliation: University of Nebraska-Lincoln, 103 Entomology Hall, Lincoln, NE 68583, United States.
authors.forename: Jessica
authors.initials: J
authors.lastname: Jurzenski
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: Matthew
authors.initials: M
authors.lastname: Kelker
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Xu
authors.affiliation: Dow AgroSciences, 9330 Zionsville Rd, Indianapolis, IN 46268, United States.
authors.forename: Kenneth
authors.initials: K
authors.lastname: Narva
authors.affiliation: University of Florida, Entomology and Nematology Department, Charles Steinmetz Hall, PO Box 110620, Gainesville, FL 32611-0620, United States. Electronic address: bsiegfried1@ufl.edu.
authors.forename: Blair D
authors.initials: BD
authors.lastname: Siegfried
mesh_terms: D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000494:pharmacology*; D015820:Cadherins / Q000235:genetics* / Q000378:metabolism; D001517:Coleoptera / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D004731:Endotoxins / Q000494:pharmacology*; D006460:Hemolysin Proteins / Q000494:pharmacology*; D019476:Insect Proteins / Q000235:genetics* / Q000378:metabolism; D007305:Insecticide Resistance; D007306:Insecticides / Q000494:pharmacology*; D007814:Larva / Q000187:drug effects / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000737:chemistry; D034622:RNA Interference; D060888:Real-Time Polymerase Chain Reaction; D003313:Zea mays / Q000737:chemistry
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D015820:Cadherins; D004731:Endotoxins; D006460:Hemolysin Proteins; D019476:Insect Proteins; D007306:Insecticides; C030956:insecticidal crystal protein, Bacillus Thuringiensis
keywords: Cadherin; Cry34/35Ab1; Cry34Ab1; Cry35Ab1; Cry3Aa; Diabrotica virgifera virgifera; RNAi