Endosomal Localization of TLR8 Confers Distinctive Proteolytic Processing on Human Myeloid Cells

Abstract

Nucleic acid–sensing TLRs are involved in both antimicrobial immune responses and autoimmune inflammation. TLR8 is phylogenetically and structurally related to TLR7 and TLR9, which undergo proteolytic processing in the endolysosomes to generate functional receptors. Recent structural analyses of human TLR8 ectodomain and its liganded form demonstrated that TLR8 is also cleaved, and both the N- and C-terminal halves contribute to ligand binding. However, the structures and ssRNA recognition mode of endogenous TLR8 in human primary cells are largely unknown. In this study, we show that proteolytic processing of TLR8 occurs in human monocytes and macrophages in a different manner compared with TLR7/9 cleavage. The insertion loop between leucine-rich repeats 14 and 15 in TLR8 is indispensable for the cleavage and stepwise processing that occurs in the N-terminal fragment. Both furin-like proprotein convertase and cathepsins contribute to TLR8 cleavage in the early/late endosomes. TLR8 recognizes viral ssRNA and endogenous RNA, such as microRNAs, resulting in the production of proinflammatory cytokines. Hence, localization sites of the receptors are crucial for the nucleic acid–sensing mode and downstream signaling.

TLR8 Senses Staphylococcus aureus RNA in Human Primary Monocytes and Macrophages and Induces IFN-β Production via a TAK1–IKKβ–IRF5 Signaling Pathway

“…However, two TLR8 bands of ∼100 kDa were not completely eliminated even several days after mRNA knockdown. These bands probably correspond to N-terminal fragments of TLR8 after cleavage in endosomes and can be a functional PRR as recently shown (32). MDMs showed similar responses as monocytes, as costimulation with synthetic TLR2 ligand or S. aureus lipoproteins strongly antagonized IFN-b induction by S. aureus and synthetic TLR8 ligand (not shown).…”

LRRC59 Regulates Trafficking of Nucleic Acid–Sensing TLRs from the Endoplasmic Reticulum via Association with UNC93B1

“…in the endosomes/lysosomes, whereas nonimmune cells such as MEFs and epithelial cells have low levels of UNC93B1, resulting in the predominance of the uncleaved form of these TLRs in the ER (5,11,31,32). In the steady-state, UNC93B1 controls trafficking of NA-sensing TLRs from the ER through binding to their transmembrane domains and the juxtamembrane region at the ER (12)(13)(14)33), and coexists with translocated TLRs.…”

“…TLR3, 7, 8, and 9 sense endocytosed nucleic acids (NAs) in endosomal compartments (3). Endosomal localization and intracellular trafficking of these TLRs are strictly regulated to evade recognition of self-RNA/DNA (4), which is also necessary for generation of functional cleaved/associated form of receptors (5)(6)(7)(8)(9)(10)(11). After synthesis, NA-sensing TLRs are retained in the endoplasmic reticulum (ER) and translocate to the endosomal compartment.…”

Lipoproteins of Gram-Positive Bacteria: Key Players in the Immune Response and Virulence

“…One explanation could be that the TLR2 stimulus competes with TLR8 for the use of MyD88. Another explanation refers to the different localizations of TLR8 and TLR2 within the cell: TLR2 is localized mainly in the cell membrane, while TLR8 is localized mainly in the endosome (100,101). This means that free Lpp can immediately activate TLR2 at the host cell surface, while TLR8 activation usually requires bacterial uptake and release of RNA in the host cell.…”