TAK1-dependent Ubiquitin Chain Editing Regulates IMD Signaling. Li Chen, Uday Aggarwal, Boae Choi, Neal Silverman. Med/Div Infectious Dis, Univ Massachusetts Med Sch, Worcester, MA.

   The humoral immune response in Drosophila is characterized by the robust and rapid induction of a battery of antimicrobial peptides (AMPs) immediately upon infection. Through the IMD pathway, AMP gene expression is induced upon sensing DAP-type peptidoglycan (PGN), common to the cell wall of Gram-negative and certain Gram-positive bacteria. The IMD pathway drives AMP expression through the activation the transcription factor Relish, an NF-B precursor protein. Signal transduction leading from PGN recognition to Relish activation requires both proteolytic cleavage and K63-polyubiquitination of IMD. However the molecular mechanisms regulating polyubiquitination in the IMD pathway remain unclear. Here, we demonstrate that upon stimulation by PGN, the cleaved IMD protein is rapidly K63-polyubiquitinated at lysine residues 137 and 153, by the E3 ligase DIAP2 and two E2 (ubiquitin conjugating) enzymes: Effete (a Drosophila Ubc5 homolog), and a complex of Uev1a and Bendless (the Drosophila Ubc13 homolog). Furthermore, ubiquitination of IMD leads to activation of the kinase TAK1, which, in turn, is required for the phosphorylation of IMD at T162 and S164. TAK1 is not only required for IMD phosphorylation, but also for the removal of K63-polyubiquitin and subsequent conjugation with K48-polyubiquitin. These data suggest that the TAK1-dependent phosphorylation of IMD protein is crucial for the ubiquitin editing of IMD. Upon stimulation, TAK1 is activated and, in a feedback loop, triggers the phosphorylation and subsequent transition from K63- to K48-polyubiquitination of IMD. Once conjugated with K48-chains, IMD is degraded by the proteasome, a critical event to down-modulate the immune response.