Identification of 34 Novel Proinflammatory Proteins in a Genome-Wide Macrophage Functional Screen

Abstract

Signal transduction pathways activated by Toll-like Receptors and the IL-1 family of cytokines are fundamental to mounting an innate immune response and thus to clearing pathogens and promoting wound healing. Whilst mechanistic understanding of the regulation of innate signalling pathways has advanced considerably in recent years, there are still a number of critical controllers to be discovered. In order to characterise novel regulators of macrophage inflammation, we have carried out an extensive, cDNA-based forward genetic screen and identified 34 novel activators, based on their ability to induce the expression of cxcl2. Many are physiologically expressed in macrophages, although the majority of genes uncovered in our screen have not previously been linked to innate immunity. We show that expression of particular activators has profound but distinct impacts on LPS-induced inflammatory gene expression, including switch-type, amplifier and sensitiser behaviours. Furthermore, the novel genes identified here interact with the canonical inflammatory signalling network via specific mechanisms, as demonstrated by the use of dominant negative forms of IL1/TLR signalling mediators.

TMEM203 is a binding partner and regulator of STING-mediated inflammatory signaling in macrophages

“…We have previously identified multiple regulators of inflammatory signaling in macrophages by genome-wide expression screening for genes which drove expression of the inflammatory chemokine Cxcl2 when transfected into RAW 264.7 cells (21); TMEM203 was one such protein. Multiple alignment of TMEM203 orthologs from a wide range of species demonstrated that TMEM203 is an evolutionally highly conserved gene ( SI Appendix , Fig.…”

“…In previous functional screens discovering novel regulators of inflammation (21), we reported the identification of TMEM203 as a previously unknown proinflammatory gene in mouse macrophages (21). Here, we demonstrate that TMEM203, a protein that was recently shown to be endosomal and interacts with the pleiotropic inositol phosphate signaling pathway protein IP3R (22), is associated with SLE disease activity, forms a functional and ligand-dependent complex with STING, and promotes its translocation to endo-lysosomes.…”

Lipid Mediators-Role in Resolution of Inflammation

“…11,12 Defects in these clearance mechanisms appear to be associated with persistent tissue inflammation and autoimmunity to cellular contents. 13,14 Tissue damage resulting from uncontrolled acute inflammatory responses causes discomfort and severely compromises normal tissue function. 15 In both acute and chronic models of inflammation, endogenous resolvins have been shown to accelerate resolution of inflammation.…”

An In vitro Cell-Based LDL Uptake Model for Screening PCSK9 Modulators

“…High-throughput screening was carried out in high throughput robotics facility in the university of Sheffield [11]. Human liver HepG2 cells were seeded in 96 well tissue plates at concentration 2×10 5 cells/ml.…”