The pyrin domain of IFI16 forms a distinct filament structure important for IFI16-mediated enhancement of STING activation

• Filament cryo-EM structure of the pyrin domain from IFI16, an innate immune DNA sensor • IFI16 PYD filament assembly is mediated by three types of PYD-PYD interfaces • IFI16 PYD filament differs from other known inflammasome-related PYD filaments • PYD filament formation is important for IFI16-mediated enhancement of STING activation Filament cryo-EM structure of the pyrin domain from IFI16, an innate immune DNA sensor IFI16 PYD filament assembly is mediated by three types of PYD-PYD interfaces IFI16 PYD filament differs from other known inflammasome-related PYD filaments PYD filament formation is important for IFI16-mediated enhancement of STING activation Interferon-gamma inducible protein 16 (IFI16) is a member of the HIN-200 family and has been proposed as a pathogenic DNA sensor with the dual function of activating both the inflammasome pathway and the stimulator of interferon genes (STING) pathway. Here, our cryo-EM analysis revealed that the pyrin domain (PYD) of IFI16 self-oligomerizes into a filamentous structure. The filament structure of IFI16 PYD is distinct from those of PYDs in other inflammasome-related proteins, including ASC, AIM2, and NLRP3. Specifically, the IFI16 PYD filament can be described as a three-start, right-handed helix assembly as for the known PYD filaments, but with a helical rise of 17.4 Å instead of ∼14 Å and a helical twist of 43.7° instead of ∼55° and lacking the C 3 symmetry, resulting in a smaller diameter. Moreover, IFI16 mutants with impaired filament formation exhibited a reduced ability to enhance STING activation compared with wildtype IFI16 in a cellular reporter assay.