Figure 4.

Activation of antimicrobial peptide promoters by Spaetzle and LPS in S2 cells. (A) S2 cells were cotransfected with 1 μg of either an empty expression vector or a vector expressing the carboxyl-terminal 106 aa of Spaetzle (Spz) (4, 30) and 0.1 μg of reporter constructs expressing luciferase under the control of the drosomycin (Drom) or attacin (Att) promoters. Cells transfected with the empty expression vector were either left untreated or exposed to LPS (10 μg/ml) for 16 h before harvesting and determination of luciferase activity in cell extracts. (B) S2 cells were cotransfected with 0.1 μg of drosomycin-luciferase reporter construct and 0.5 μg of expression vector encoding the C106 processed form of Spaetzle (Spz) together with 0.5 μg of expression vector empty or encoding a truncated Toll version deleted of its intracytoplasmic domain (TollΔIC) or a mutant Pelle version deleted of its kinase domain (PelleDN). (C) S2 cells were cotransfected with 0.1 μg of attacin-luciferase reporter construct and 1 μg of expression vector empty or encoding TollΔIC or PelleDN. Twenty-four hours after transfection, cells were either left untreated or exposed to LPS (10 μg/ml) for 16 h before harvesting and determination of luciferase activity in cell extracts. (D) S2 cells were cotransfected with 0.1 μg of drosomycin-luciferase reporter plasmid and 0.5 μg of expression vector encoding the Toll^ΔLRR-Toll5 chimeric protein together with 0.5 μg of expression vector either empty or encoding PelleDN. All transfections were done in triplicate, and results represent means ± SD.