Li, H.; Vallese, F.; Clarke, O. B.

Vaults are some of the largest ribonucleoprotein complexes known, and are highly conserved across eukaryotes, but both their function and key details of their architecture remain unclear. While high-resolution structures of the vault shell are available, the architecture and symmetry of the cap at either end of the vault has remained unresolved. Here we present a 2.25 A cryo-EM structure of the vault cap, revealing an unexpected 13-fold symmetric arrangement that contrasts with the 39-fold symmetry of the vault body, with each repeating module of the cap formed by an asymmetric homotrimer of adjacent subunits, in which two C-termini remain in the vault interior, and one projects into the cytosol. The center of the cap features an unusual architecture reminiscent of the Stomatin, Flotillin and HflK/C (SPFH) superfamily, consisting of two concentric beta barrels surrounded by an interwoven two-layer stack of alpha-helices, with the innermost barrel forming a 15 A aperture connecting the interior of the vault with the cytosol. The vault cap features a positively charged exterior and a negatively charged interior surface, with implications for binding partner recruitment and putative binding of the vault particle to microtubules and lipid rafts. These findings uncover a new facet of vault particle architecture, and have implications for engineering and design of modified vault particles for therapeutic delivery, as well as providing new opportunities for interrogating the functional roles of the vault particle in biological systems.