The role of E3 ubiquitin-protein ligase DTX3L in the innate immune response

Abstract

Post translational modifications (PTMs) control many mechanisms of cellular homeostasis including the immune response. Ubiquitylation is a major PTM in eukaryotic cells which denotes complex information transfer through a variety of possible chain types mediated by >600 E3 ubiquitin-protein ligases. I studied ubiquitylation in the macrophage innate immune response to identify E3 ligases responding to pro-inflammatory ligands. Amongst 45 E3 ligases identified by LC-MS/MS based proteomics, Deltex 3-like (DTX3L) displayed the greatest upregulation. Bioinformatic interrogation of DTX3L revealed its involvement in the hostresponse to viral infection and association with protein mono-ADP-polymerase PARP9 (PARP9) and -14 (PARP14). Proteomics analysis of Dtx3l gene knockout models suggested DTX3L functions as a negative regulator of the immune response and interferon stimulated genes (ISG). Co-immunoprecipitation proteomics of HA-DTX3L revealed interactions with PARP9 as well as endo-lysosomal marker Rab7a. DTX3L was required for normal mRNA and protein expression as well as interferon-gamma (IFN-γ) induced induction of PARP9 and -14. PARP9 and -14 are ISGs implicated in controlling macrophage phenotype through influence over signal transducer and activator of transcription 1 (STAT1) and -6 (STAT6), regulators of pro-inflammatory and alternatively activated macrophages, respectively. Consequently, Dtx3l KO increased STAT1 and decreased STAT6 levels, as well as reducing interferon regulatory factor 1 induction, whilst maintaining normal arginase-1 induction. Bacterial infections of Dtx3l KO BMDMs revealed increased killing of S. Typhimurium and increased uptake and killing of S. aureus, while Murine Respirovirus (SeV) infection showed an increased IFN-γ mediated immune response and a propensity for antigen presentation. Together I found that DTX3L acts as a negative regulator of the IFN-γ-STAT1 mediated innate immune response by controlling the resting condition of macrophages as well as dampening the immune response to virus and bacteria