Downregulation of lipopolysaccharide response in drosophila by negative crosstalk between the AP1 and NF-jB signaling modules Taeil Kim 1 , Joonsun Yoon 1 , Hwansung Cho 1 , Wook-bin Lee 1 , Joon Kim 1 , Young-Hwa Song 2 , Se Nyun Kim 2 , Jeong Ho Yoon 2 , Jeongsil Kim-Ha 3 & Young-Joon Kim 1 IjB kinase (IKK) and Jun N-terminal kinase (Jnk) signaling modules are important in the synthesis of immune effector molecules during innate immune responses against lipopolysaccharide and peptidoglycan. However, the regulatory mechanisms required for specificity and termination of these immune responses are unclear. We show here that crosstalk occurred between the drosophila Jnk and IKK pathways, which led to downregulation of each other’s activity. The inhibitory action of Jnk was mediated by binding of drosophila activator protein 1 (AP1) to promoters activated by the transcription factor NF-jB. This binding led to recruitment of the histone deacetylase dHDAC1 to the promoter of the gene encoding the antibacterial protein Attacin-A and to local modification of histone acetylation content. Thus, AP1 acts as a repressor by recruiting the deacetylase complex to terminate activation of a group of NF-jB target genes. Innate immunity is the first-line defense system of multicellular organisms in response to various microbial invaders 1–3 . Recognition of pathogen-associated molecular patterns (PAMPs) by germline- encoded receptors initiates the signaling cascades leading to the activation of various genes encoding antimicrobial peptides, cytokines, inflammatory mediators and regulators of phagocytosis 4–6 . In droso- phila, two distinctive signaling pathways, the Toll and immune deficiency (Imd) pathways, have specific functions in inducing PAMP-dependent innate immune responses 2,3,7–10 . The Toll signaling pathway was initially identified as controlling embryonic dorsoventral patterning in drosophila 11,12 but was later found to control the innate immune response against fungal and Gram-positive bacterial infections 12–14 . Spa ¨tzle, a ligand for Toll, is activated by proteolytic cleavage by a serine protease after infec- tion 15,16 , and binding of Spa ¨tzle to the Toll receptor activates a signaling cascade through dMyD88 (drosophila myeloid differentia- tion factor 88 homolog, an adaptor in the Toll pathway), the IRAK-like serine–threonine kinase Pelle, and Tube (adaptor protein contain- ing death domain). This activation results in the degradation of IkB-like Cactus, followed by nuclear translocation of the transcription factor NF-kB homologs Dorsal and Dorsal-related immunity factor to activate distinctive antifungal and anti–Gram-positive bacterial immune reactions 17–20 . In contrast, the immune response to Gram- negative bacterial infection is controlled by the Imd pathway, which culminates in the expression of antimicrobial peptides such as Attacin, Cecropin and Diptericin 2,6 . Lipopolysaccharide (LPS) was initially thought to be the triggering ligand for the Imd pathway, but peptidoglycan (PGN) contaminating the purified LPS (LPS-PGN) has been identified as the actual stimulant for the Imd pathway 21,22 . Flies mutant for Imd have severe defects in resistance to Gram- negative bacterial infection but are normal in their response to fungal and Gram-positive bacterial infection 23–26 . Recognition of Gram- negative bacterial PGN by a PGN recognition protein 22,27,28 recruits and activates Imd, which is a homolog of mammalian RIP (tumor necrosis factor receptor–interacting protein), dTAK1 (transforming growth factor-b–activated kinase 1) and the IkB kinase (IKK) com- plex 8,29 . Phosphorylation of Relish by IKK 30–32 and its subsequent nuclear translocation activates expression of the Imd pathway–specific antimicrobial peptide genes 33 . In addition to the activation of IKK, which leads to the synthesis of large amounts of antimicrobial peptides, the activated dTAK1 also stimulates the Jun N-terminal kinase (Jnk) signaling module 8,34 . Jnk activation has been linked to the control of diverse biological pro- cesses, including morphogenesis, immunity and apoptosis 35 . In parti- cular, LPS-PGN-induced Jnk activation of the Imd pathway is involved in the synthesis of various cytokines and the cytoskeletal remodeling needed for phagocytotic responses 7,34–36 . Mammalian NF-kB negatively regulates the Jnk pathway in tumor necrosis factor– induced apoptosis 37,38 . This type of negative crosstalk has also been found in drosophila challenged with LPS and involves proteasomal degradation of dTAK1 by Relish-dependent genes 39 . Inhibition of the Jnk module by NF-kB results in transient Jnk activation, thus © 2004 Nature Publishing Group http://www.nature.com/natureimmunology Published online 9 January 2005; doi:10.1038/ni1159 1 Department of Biochemistry, Yonsei University, Seoul 120-749, South Korea. 2 Digital Genomics, Seoul 120-749, South Korea. 3 Department of Molecular Biology, Sejong University, Seoul 143-747, South Korea. Correspondence should be addressed to Y.-J.K. (yjkim@yonsei.ac.kr). NATURE IMMUNOLOGY ADVANCE ONLINE PUBLICATION 1 ARTICLES