The Chaperone UNC93B1 Regulates Toll-like Receptor Stability Independently of Endosomal TLR Transport.

Unc-93 homolog B1 (UNC93B1) is a key regulator of nucleic acid (NA)-sensing Toll-like receptors (TLRs). Loss of NA-sensing TLR responses in UNC93B1-deficient patients facilitates Herpes simplex virus type 1 (HSV-1) encephalitis. UNC93B1 is thought to guide NA-sensing TLRs from the endoplasmic reticulum (ER) to their respective endosomal signaling compartments and to guide the flagellin receptor TLR5 to the cell surface, raising the question of how UNC93B1 mediates differential TLR trafficking. Here, we report that UNC93B1 regulates a step upstream of the differential TLR trafficking process. We discovered that UNC93B1 deficiency resulted in near-complete loss of TLR3 and TLR7 proteins in primary splenic mouse dendritic cells and macrophages, showing that UNC93B1 is critical for maintaining TLR expression. Notably, expression of an ER-retained UNC93B1 version was sufficient to stabilize TLRs and largely restore endosomal TLR trafficking and activity. These data are critical for an understanding of how UNC93B1 can regulate the function of a broad subset of TLRs.

HMGB1, IL-1α, IL-33 and S100 proteins: dual-function alarmins.

Our immune system is based on the close collaboration of the innate and adaptive immune systems for the rapid detection of any threats to the host. Recognition of pathogen-derived molecules is entrusted to specific germline-encoded signaling receptors. The same receptors have now also emerged as efficient detectors of misplaced or altered self-molecules that signal tissue damage and cell death following, for example, disruption of the blood supply and subsequent hypoxia. Many types of endogenous molecules have been shown to provoke such sterile inflammatory states when released from dying cells. However, a group of proteins referred to as alarmins have both intracellular and extracellular functions which have been the subject of intense research. Indeed, alarmins can either exert beneficial cell housekeeping functions, leading to tissue repair, or provoke deleterious uncontrolled inflammation. This group of proteins includes the high-mobility group box 1 protein (HMGB1), interleukin (IL)-1α, IL-33 and the Ca2+-binding S100 proteins. These dual-function proteins share conserved regulatory mechanisms, such as secretory routes, post-translational modifications and enzymatic processing, that govern their extracellular functions in time and space. Release of alarmins from mesenchymal cells is a highly relevant mechanism by which immune cells can be alerted of tissue damage, and alarmins play a key role in the development of acute or chronic inflammatory diseases and in cancer development.

Transcriptome assessment reveals a dominant role for TLR4 in the activation of human monocytes by the alarmin MRP8.

The alarmins myeloid-related protein (MRP)8 and MRP14 are the most prevalent cytoplasmic proteins in phagocytes. When released from activated or necrotic phagocytes, extracellular MRP8/MRP14 promote inflammation in many diseases, including infections, allergies, autoimmune diseases, rheumatoid arthritis, and inflammatory bowel disease. The involvement of TLR4 and the multiligand receptor for advanced glycation end products as receptors during MRP8-mediated effects on inflammation remains controversial. By comparative bioinformatic analysis of genome-wide response patterns of human monocytes to MRP8, endotoxins, and various cytokines, we have developed a model in which TLR4 is the dominant receptor for MRP8-mediated phagocyte activation. The relevance of the TLR4 signaling pathway was experimentally validated using human and murine models of TLR4- and receptor for advanced glycation end products-dependent signaling. Furthermore, our systems biology approach has uncovered an antiapoptotic role for MRP8 in monocytes, which was corroborated by independent functional experiments. Our data confirm the primary importance of the TLR4/MRP8 axis in the activation of human monocytes, representing a novel and attractive target for modulation of the overwhelming innate immune response.

Innate immune receptors for nucleic acids.

The innate immune system has evolved to detect microbes and sterile tissue damage with the help of a series of signaling receptors. One key strategy is to detect infectious microbes or host cell damage by recognizing nucleic acids that are modified or appear in compartment normally devoid of nucleic acids. Here, we describe two methods that allow studying the molecular interaction between various nucleic acid recognizing signaling receptors with their ligands. A ligand pull-down assay can be used to show a known interaction between a ligand and its receptor or the method can be utilized as a discovery approach to identify an unknown receptor to a given ligand. An AlphaScreen experiment can be set up to assess the ligand binding affinity to a given receptor.