Bruton´s tyrosine kinase-mediated NLRP3 phosphorylation promotes inflammasome activation

In dieser Studie haben wir angestrebt, die Mechanismen, über die Bruton´s Tyrosinkinase (BTK) NLRP3 reguliert, aufzuklären. Vorarbeiten haben gezeigt, dass die Präsenz einer intakten BTK die IL-1β Ausschüttung erhöht. Inhibierung von BTK mit dem FDA-genehmigten Kinaseinhibitor Ibrutinib blockierte die IL-1β Produktion. Wir stellten die Hypothese auf, dass die Kinaseaktivität von BTK in dem Regulationsmechanismus eine Rolle spielt. Wir haben festgestellt, dass NLRP3 und BTK interagierten und tyrosinphosphoryliert wurden, wenn man das NLRP3 Inflammasom in primären PBMCs oder BMDMs stimulierte. Daraufhin haben wir gezeigt, dass BTK NLRP3 phosphoryliert und haben die modifizierten Tyrosinreste identifiziert: Y136/Y140/Y143/Y168. Wir haben bemerkt, dass Y136/Y140/Y143 in der kürzlich beschriebenen polybasischen Region liegen, die für die Phospholipid-Bindung von NLRP3 am „trans-Golgi Network“ sorgt. Wir haben vermutet, dass Phosphorylierung dieser Region durch die zusätzliche negative Ladung zur Lösung von NLRP3 vom Golgi führt. Tatsächlich haben „phosphomimetische“ NLRP3 Konstrukte (Y>E) weniger an Phospholipid-beschichteten Agarosebeads gebunden, als WT Konstrukte. Zusätzlich haben Btk KO Zellen und ibrutinib-behandelte Immunzellen weniger NLRP3 Oligomere im Zytosol gebildet. Außerdem haben mit NLRP3 Y>F rekonstituierte Makrophagen weniger IL-1β produziert, als mit WT NLRP3 rekonstituierten Zellen, was einen klaren Hinweis auf die funktionelle Rolle der Phosphostellen darstellt

Pyroptosis and Its Role in SARS-CoV-2 Infection

The pore-forming inflammatory cell death pathway, pyroptosis, was first described in the early 1990s and its role in health and disease has been intensively studied since. The effector molecule GSDMD is cleaved by activated caspases, mainly Caspase 1 or 11 (Caspase 4/5 in humans), downstream of inflammasome formation. In this review, we describe the molecular events related to GSDMD-mediated pore formation. Furthermore, we summarize the so far elucidated ways of SARS-CoV-2 induced NLRP3 inflammasome formation leading to pyroptosis, which strongly contributes to COVID-19 pathology. We also explore the potential of NLRP3 and GSDMD inhibitors as therapeutics to counter excessive inflammation

Bruton’s Tyrosine Kinase: An Emerging Key Player in Innate Immunity

Bruton’s tyrosine kinase (BTK) was initially discovered as a critical mediator of B cell receptor signaling in the development and functioning of adaptive immunity. Growing evidence also suggests multiple roles for BTK in mononuclear cells of the innate immune system, especially in dendritic cells and macrophages. For example, BTK has been shown to function in Toll-like receptor-mediated recognition of infectious agents, cellular maturation and recruitment processes, and Fc receptor signaling. Most recently, BTK was additionally identified as a direct regulator of a key innate inflammatory machinery, the NLRP3 inflammasome. BTK has thus attracted interest not only for gaining a more thorough basic understanding of the human innate immune system but also as a target to therapeutically modulate innate immunity. We here review the latest developments on the role of BTK in mononuclear innate immune cells in mouse versus man, with specific emphasis on the sensing of infectious agents and the induction of inflammation. Therapeutic implications for modulating innate immunity and critical open questions are also discussed

Platelets Aggregate With Neutrophils and Promote Skin Pathology in Psoriasis

Psoriasis is a frequent systemic inflammatory autoimmune disease characterized primarily by skin lesions with massive infiltration of leukocytes, but frequently also presents with cardiovascular comorbidities. Especially polymorphonuclear neutrophils (PMNs) abundantly infiltrate psoriatic skin but the cues that prompt PMNs to home to the skin are not well-defined. To identify PMN surface receptors that may explain PMN skin homing in psoriasis patients, we screened 332 surface antigens on primary human blood PMNs from healthy donors and psoriasis patients. We identified platelet surface antigens as a defining feature of psoriasis PMNs, due to a significantly increased aggregation of neutrophils and platelets in the blood of psoriasis patients. Similarly, in the imiquimod-induced experimental in vivo mouse model of psoriasis, disease induction promoted PMN-platelet aggregate formation. In psoriasis patients, disease incidence directly correlated with blood platelet counts and platelets were detected in direct contact with PMNs in psoriatic but not healthy skin. Importantly, depletion of circulating platelets in mice in vivo ameliorated disease severity significantly, indicating that both PMNs and platelets may be relevant for psoriasis pathology and disease severity

The fungal ligand chitin directly binds TLR2 and triggers inflammation dependent on oligomer size

Chitin is the second most abundant polysaccharide in nature and linked to fungal infection and asthma. However, bona fide immune receptors directly binding chitin and signaling immune activation and inflammation have not been clearly identified because polymeric crude chitin with unknown purity and molecular composition has been used. By using defined chitin (N-acetyl-glucosamine) oligomers, we here identify six-subunit-long chitin chains as the smallest immunologically active motif and the innate immune receptor Toll-like receptor (TLR2) as a primary fungal chitin sensor on human and murine immune cells. Chitin oligomers directly bind TLR2 with nanomolar affinity, and this fungal TLR2 ligand shows overlapping and distinct signaling outcomes compared to known mycobacterial TLR2 ligands. Unexpectedly, chitin oligomers composed of five or less subunits are inactive, hinting to a size-dependent system of immuno-modulation that appears conserved in plants and humans. Since blocking of the chitin-TLR2 interaction effectively prevents chitin-mediated inflammation in vitro and in vivo, our study highlights the chitin-TLR2 interaction as a potential target for developing novel therapies in chitin-related pathologies and fungal disease

The fungal ligand chitin directly binds TLR2 and triggers inflammation dependent on oligomer size.

Chitin is the second most abundant polysaccharide in nature and linked to fungal infection and asthma. However, bona fide immune receptors directly binding chitin and signaling immune activation and inflammation have not been clearly identified because polymeric crude chitin with unknown purity and molecular composition has been used. By using defined chitin (N-acetyl-glucosamine) oligomers, we here identify six-subunit-long chitin chains as the smallest immunologically active motif and the innate immune receptor Toll-like receptor (TLR2) as a primary fungal chitin sensor on human and murine immune cells. Chitin oligomers directly bind TLR2 with nanomolar affinity, and this fungal TLR2 ligand shows overlapping and distinct signaling outcomes compared to known mycobacterial TLR2 ligands. Unexpectedly, chitin oligomers composed of five or less subunits are inactive, hinting to a size-dependent system of immuno-modulation that appears conserved in plants and humans. Since blocking of the chitin-TLR2 interaction effectively prevents chitin-mediated inflammation in vitro and in vivo, our study highlights the chitin-TLR2 interaction as a potential target for developing novel therapies in chitin-related pathologies and fungal disease