Cahoon, J. G. | Search bioRxiv

bioRxiv2026

MEX3B is a positive pan-inflammasome regulator

Inflammasomes lead to activation of inflammatory caspases, which induce pyroptosis and an inflammatory immune response to control microbial infections. Inflammasomes are tightly regulated to avoid lethal sepsis and chronic autoimmune conditions. However, posttranslational regulation of inflammatory caspases remains poorly defined. We constructed 375 individual ubiquitin ligase knockout lines by CRISPR-Cas9, performed an unbiased screening, and identified Muscle Excess 3B (MEX3B), an RNA-binding protein and ubiquitin ligase, as a positive regulator of the caspase-4 inflammasome. Genetic depletion of MEX3B inhibited not only the caspase-4 but also NLRP3 and NLRC4 inflammasomes, regarding caspase activation, pyroptosis, and secretion of inflammasome-dependent cytokines, in human cells and murine primary macrophages. This MEX3B function required its RNA-binding, but not ubiquitin ligase activity. These results suggest that MEX3B is a pan-inflammasome regulator and a potential therapeutic target for inflammation.

immunology↗

bioRxiv2024

Unanchored ubiquitin chains promote the non-canonical inflammasome via UBXN1

Ubiquitination is a major posttranslational covalent modification that regulates numerous cellular processes including inflammasome signaling. Cells also contain unanchored ubiquitin chains (polyUb) that bind protein targets non-covalently, but their physiological functions in immunity have been appreciated only recently. Here, we report that ubiquitin regulatory x domain-containing protein 1 (UBXN1) activates the noncanonical inflammasome via unanchored Lysin (K) 48- or 63-linked polyUb. UBXN1 deficiency impairs the activation of caspase-4/11, secretion of inflammasome-dependent cytokines and pyroptosis in response to intracellular lipopolysaccharide (LPS). UBXN1-deficient mice are protected from LPS- and cecal-ligation-and-puncture-induced sepsis, evidenced by reduced mortality and systemic inflammation, compared to UBXN1-sufficient littermates. Mechanistically, UBXN1 together with unanchored K48/63-linked polyUb bind caspase-4/11, the intracellular sensors of LPS, and promote their assembly and activation. Depleting cellular unanchored polyUb with recombinant ubiquitin-specific proteinase 5 (USP5) reduces UBXN1 binding to caspase-4/11 and inflammasome signaling, while USP5 inhibitors enhance pyroptosis in an UBXN1-dependent manner. Thus, this study identifies a critical UBXN1-dependent posttranslational mechanism involved in noncanonical inflammasome activation and UBXN1 as a potential therapeutic target for sepsis and advances a fundamental understanding of unanchored polyUb biology.

immunology↗