Transkingdom mechanism of MAMP generation by chitotriosidase (CHIT1) feeds oligomeric chitin from fungal pathogens and allergens into TLR2-mediated innate immune sensing

Abstract

Chitin is a highly abundant polysaccharide in nature and linked to immune recognition of fungal infections and asthma in humans. Ubiquitous in fungi and insects, chitin is absent in mammals and plants and, thus, represents a microbe-associated molecular pattern (MAMP). However, the highly polymeric chitin is insoluble, which potentially hampers recognition by host immune sensors. In plants, secreted chitinases degrade polymeric chitin into diffusible oligomers, which are "fed to" innate immune receptors and co-receptors. In human and murine immune cells, a similar enzymatic activity was shown for human chitotriosidase (CHIT1) and oligomeric chitin is sensed via an innate immune receptor, Toll-like receptor (TLR) 2. However, a complete system of generating MAMPs from chitin and feeding them into a specific receptor/co-receptor-aided sensing mechanism has remained unknown in mammals. Here, we show that the secreted chitinolytic host enzyme, CHIT1, converts inert polymeric chitin into diffusible oligomers that can be sensed by TLR1-TLR2 co-receptor/receptor heterodimers, a process promoted by the lipopolysaccharide binding protein (LBP) and CD14. Furthermore, we observed that Chit1 is induced via the beta-glucan receptor Dectin-1 upon direct contact of immortalized human macrophages to the fungal pathogen Candida albicans, whereas the defined fungal secreted aspartyl proteases, Sap2 and Sap6, from C. albicans were able to degrade CHIT1 in vitro. Our study shows the existence of an inducible system of MAMP generation in the human host that enables contact-independent immune activation by diffusible MAMP ligands with striking similarity to the plant kingdom. Moreover, this study highlights CHIT1 as a potential therapeutic target for TLR2-mediated inflammatory processes that are fueled by oligomeric chitin.

Chitin oligomers directly promote lymphoid innate and adaptive immune cell activation

“…Compared to earlier results, chitin oligomers thus not only possess a strong capacity to fully signal danger to epithelial 11 and myeloid innate cell types (e.g. macrophages, PMNs) 10, 15 but also to lymphoid innate immune cells such as NK cells. They can also directly act on B lymphocytes in an antigen-independent manner.…”

“…By activating antigen-presenting cells such as MoDCs, chitin oligomers are also able to indirectly promote T cell mediated adaptive immunity and display adjuvant like properties. It could be envisaged that in house dust mite allergies, chitin oligomers, once released from Dermatophagoides house dust mites by the host chitinase, CHIT1 10 , thus perform a "self-adjuvanting" role for associated T cell antigens 17 . From a translational perspective, our study raises the possibilities that blocking chitin-TLR2 interactions might prevent immune activation more broadly than previously thought or that chitin oligomers could be a useful addition to the short list of adjuvants suitable for use in humans 18 .…”

“…oligomer length, rather than crude chitin, and identified six-subunit-long chitin oligomers as the smallest immunologically active motif. Additionally, we identified the innate immune receptor Toll-like receptor (TLR2) as a primary fungal chitin sensor on human and murine myeloid cells {Fuchs, 2018 #9694} and the host chitinase CHIT1 as an enzyme involved in oligomer generation 10 . Illustrating the validity of using pure chitin oligomers, He and colleagues have also shown that chitin oligomers can be recognized by the Lysin motif (LysM)-domain containing receptor LYSMD3 on the surface of human airway epithelial cells and elicit production of inflammatory cytokines 11 .…”

Release of the pre-assembled naRNA-LL37 composite DAMP re-defines neutrophil extracellular traps (NETs) as intentional DAMP webs

“…Preparation of fungal RNA from C. albicans C. albicans SC5314 (kindly cultured and prepared by Tzu-Hsuan Chang, Tübingen) was plated in a slant tube containing YPD agar and grown overnight at 30 °C as described in (Chang, Gloria et al 2022). One colony was picked from the slant tube and resuspended in 500 μL YPD medium, centrifuged at 10,000 rpm for 1 min and washed with phosphate-buffered saline (PBS).…”

“…The copyright holder for this preprint (which this version posted July 27, 2022. ; https://doi.org/10. 1101/2022 We speculated that RNA contained in NETs, so-called NET-associated RNA or naRNA, might lead to a feed-forward loop of rapidly expanding NET release amongst neutrophils which might also engulf additional tissue or immune cells able to respond to exRNA. We show here that naRNA drives such a self-amplifying loop not only engaging PMN but also macrophages, and keratinocytes.…”