A Toll-ML-LPS pathway in insects

Toll/MD2/LPS pathway In mammal, Toll-like receptor 4 (TLR-4) is the receptor that recognize lipopolysaccharide (LPS). MD2 is required for the recognition and binding between TLR-4 and LPS in mammal. There are two different pathways in invertebrate immunity system including Imd and Toll pathway. However, in invertebrates, receptors that recognize LPS have not been determined. Our aim is to investigate whether invertebrate exist the similar pathway to mammal. We hypothesize that the recognition and response of LPS in invertebrate is via Toll pathway which requires MD-2-related lipid-recognition protein (ML -1). Plasmid DNAs for proteins with ecto-MsToll, Chimeric Toll (recombinant Toll with extracellular Manduca sexta Toll, transmembrane and intracellular region from Drosophila), MsML-1 will be constructed and stable cell lines will be established in Drosophila S2 cells. We apply Co-immunoprecipitation (CoIP) to discover that ecto-MsToll can directly interact with MsML-1. LPS can increase this interaction. In dual-luciferase reporter assay, we also confirm that LPS can stimulate Toll pathway in vertebrates. MsML-1 is required for the Toll pathway induced by LPS.