38th Annual Meeting of the Society for Invertebrate Pathology

Genetic knockout of the Heliothis virescens cadherin-like protein (HevCaLP) has been linked to high levels of resistance to Cry1Ac toxin (Gahan et al. 2001, Science 293:857-60). We demonstrated a lack of Cry1Aa binding to brush border membrane vesicles from larvae lacking HevCaLP (Jurat-Fuentes et al. 2004, Biochemistry 43:14299-305). Recently, HevCaLP peptide fragments expressed in Escherichia coli were reported to bind Cry1Ac in ligand blots (Xie et al. 2004, J. Biol. Chem. 280:8416-25).
The goal of this work was to transiently express full length HevCadLP on the surface of insect cells to test its putative role as a Cry1Ac toxin receptor. We cloned the full-length cDNA encoding HevCaLP into the pIZT vector. Immunocytochemistry was used to detect HevCaLP expression on the surface of transfected Drosophila melanogaster S2 and Trichoplusia ni High Five cells. Cry1Ac toxin binding was tested using fluorescence microscopy, dot blotting and cell binding assays. The receptor role of HevCaLP was studied using fluorescence assisted cell sorting (FACS). In these assays, Cry1Ac killed less than 20% of cells expressing HevCaLP. Therefore, while HevCaLP can be considered a functional receptor for Cry1Ac, the low level of cytotoxicity may suggest the participation of additional molecules in Cry1Ac intoxication in vivo.