FIG. 6.

Ubc9 and Smt3 are required for the induction of antimicrobial peptides. (A) Conjugation of Smt3 to Dorsal correlates with increased LPS induction of CecA1. 529SU cells were induced with Cu²⁺ for 24 h (lanes 1 and 2) and/or challenged with LPS for 6 h (lanes 2 and 4). Total RNA was then isolated and subjected to quantitative RT-PCR with primers specific to the CecA1 mRNA (top panel). Whole-cell lysates from these cells were fractionated by SDS-PAGE (15 or 5% acrylamide) and analyzed by immunoblotting to confirm Ubc9 and Smt3 expression (middle panel, lanes 3 and 4) and Smt3-Dorsal production (bottom panel, lanes 3 and 4). (B) Smt3 and Ubc9 are required for the induction of CecA1 and Drs in S2 cells. mRNA corresponding to the genes indicated at the top were disrupted using dsRNAi (36 h). Total RNA was isolated following 6 h of LPS challenge. RT-PCR was then performed using primers specific to the genes indicated to the right of the gels.