Poster Presentation The 3rd Prato Conference on Pore Forming Proteins 2015

Bacillus thuringiensis Cry3Aa toxin functional receptors in Colorado potato beetle larval midgut (#40)

Victor Manuel Ruiz-Arroyo 1 , Inmaculada Garcia-Robles 1 , Camila Ochoa-Campuzano 1 , Adrian Goig 1 , Sara Oltra 1 , Amparo C. Martinez-Ramirez 1 , Carolina Rausell 1 , M. Dolores Real 1
  1. Department of Genetics, University of Valencia, Burjassot, Valencia, Spain

The Gram-positive bacterium B. thuringiensis (Bt) produces during sporulation a-PFTs insecticidal Cry proteins that bind specific receptor molecules on the brush border membrane of the insect midgut cells. Bt toxin-receptor interaction is widely accepted as a crucial step for toxicity and determines insect specificity. Bt Cry3Aa toxin is highly active against larvae of Colorado potato beetle (CPB), the most devastating insect pest of potatoes worldwide, but the receptor molecules involved in the Cry3Aa toxic action have not been identified yet. In this work, we searched in the CPB transcriptome (Kumar et al., 2014) and in the CPB genome that is currently being annotated (i5k-pilot-project), the homologous gene sequences encoding Bt toxin functional receptors in the coleopteran model insect Tribolium castaneum, cadherin-like and sodium solute symporter proteins. We also searched for the CPB ADAM10 metalloprotease gene sequence that we previously proposed encodes a Cry3Aa toxin receptor in CPB larvae (Ochoa-Campuzano et al., 2007). Next, we assessed the functional role of these three molecules as Cry3Aa toxin receptors in CPB larvae by RNAi gene silencing. Results demonstrated that CPB-ADAM10 protein is a Cry3Aa functional receptor in CPB. And finally, we investigated the relevance in Cry3Aa toxin mode of action of the proteolytic processing by digestive midgut juice proteases and by ADAM10 membrane associated metalloprotease in the context of the general mechanism of PFTs pathogenesis.

  1. Kumar A., Congiu L., Lindström L., Piiroinen S., Vidotto M., and Grapputo A. (2014). Sequencing, de novo assembly and annotation of the Colorado potato beetle, Leptinotarsa decemlineata, transcriptome. PLoS ONE, 9, e86012
  2. Ochoa-Campuzano C., Real M.D., Martínez-Ramírez A.C., Bravo A., and Rausell C. (2007). An ADAM metalloprotease is a Cry3Aa Bacillus thuringiensis toxin receptor, Biochem Biophys Res Commun 362: 437-442.
  3. i5k-pilot-project: https://www.hgsc.bcm.edu/arthropods/i5k-pilot-project-summary