A proteomic approach to study Cry1Ac binding proteins and their alterations in resistant Heliothis virescens larvae.
Binding of the Bacillus thuringiensis Cry1Ac toxin to specific receptors in the midgut brush border membrane is required for toxicity. Alteration of these receptors is the most reported mechanism of resistance. We used a proteomic approach to identify Cry1Ac binding proteins from intestinal brush border membrane (BBM) prepared from Heliothis virescens larvae. Cry1Ac binding BBM proteins were detected in 2D blots and identified using peptide mass fingerprinting (PMF) or de novo sequencing. Among other proteins, the membrane bound alkaline phosphatase (HvALP), and a novel phosphatase, were identified as Cry1Ac binding proteins. Reduction of HvALP expression levels correlated directly with resistance to Cry1Ac in the YHD2-B strain of H. virescens. To study additional proteomic alterations in resistant H. virescens larvae, we used two-dimensional differential in-gel electrophoresis (2D-DIGE) to compare three independent resistant strains with a susceptible strain. Our results validate the use of proteomic approaches to identify toxin binding proteins and proteome alterations in resistant insects.
Jurat-Fuentes
Juan L
JL
Department of Entomology and Plant Pathology, University of Tennessee, Knoxville, TN 37996-4560, USA. jurat@utk.edu
Adang
Michael J
MJ
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001413:Bacillus thuringiensis / Q000502:physiology; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001427:Bacterial Toxins / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000201:enzymology; D015180:Electrophoresis, Gel, Two-Dimensional; D004731:Endotoxins / Q000737:chemistry / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry / Q000378:metabolism*; D007305:Insecticide Resistance / Q000187:drug effects* / Q000235:genetics; D007306:Insecticides / Q000737:chemistry / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000737:chemistry / Q000201:enzymology; D007814:Larva / Q000378:metabolism* / Q000382:microbiology; D007915:Lepidoptera / Q000145:classification / Q000235:genetics / Q000502:physiology*; D010449:Peptide Mapping; D010572:Pest Control, Biological / Q000379:methods; D040901:Proteomics; D013045:Species Specificity
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; D007306:Insecticides; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D000469:Alkaline Phosphatase