A proximity-dependent biotinylation (BioID) approach flags the p62/sequestosome-1 protein as a caspase-1 substrate

Area of Science:

• Immunology
• Molecular Biology
• Cellular Biology

Background:

• The inflammasome is crucial for innate immunity, activating caspase-1 to promote inflammation via IL-1β maturation and release.
• Inflammasome regulation is vital to prevent excessive inflammation, but its insoluble nature hinders protein interaction studies.
• Caspase-1 is a key protease in inflammatory pathways, tightly controlled by cellular mechanisms.

Purpose of the Study:

• To identify proteins interacting with caspase-1 during inflammasome activation.
• To elucidate the functional consequences of caspase-1-mediated cleavage of interacting proteins.
• To understand the role of p62 protein in regulating inflammasome activity and IL-1β release.

Main Methods:

• Proximity-dependent biotinylation assay (BioID) in a cell-free system to identify caspase-1 interacting proteins.
• Co-immunoprecipitation to confirm interactions between caspase-1 and p62.
• Mechanistic and functional analyses of p62 cleavage products and their effect on IL-1β release.

Main Results:

• BioID identified 111 candidate proteins interacting with caspase-1, including p62/sequestosome-1 (p62).
• Caspase-1 directly cleaves p62 at Asp-329, disrupting its interaction with LC3B.
• Cleaved p62 fragments differentially regulate IL-1β release: N-terminal fragment decreases release, C-terminal fragment enhances it by affecting pro-IL-1β levels.

Conclusions:

• Caspase-1-mediated cleavage of p62 is a critical regulatory step in inflammasome signaling.
• p62 acts as a complex modulator of inflammation, with its fragments influencing IL-1β release through distinct mechanisms.
• Understanding p62 cleavage provides insights into balancing caspase-1-driven inflammation and potential therapeutic targets.