Diversification and functional expansion of archaeal T4F machineries.

Archaea have several cell surface structures belonging to the type 4 filament (T4F) superfamily. All share a minimal core assembly system consisting of an ATPase, a platform protein, a filament-forming protein, and a class III signal peptidase. Here, we generated novel MacSyFinder2 models to identify and classify archaeal T4F systems. Our analysis revealed a vast diversity of archaeal T4F, with many organisms harboring one or more assembly machineries. Structure-based phylogenetic analysis showed that the variable N-terminal domain of T4F-related ATPases mirrors the subsystem clustering, indicating that the core machinery has diversified within the archaeal secretion ATPase family through structural innovation. Genome-wide screening for signal peptidase III cleavage site (SPIII) containing proteins revealed the widespread presence of substrate-binding proteins carrying a class III signal peptide. We propose that these binding proteins, with canonical SPIII cleavage sites, functionalize T4F machineries for efficient substrate scavenging, expanding the functional repertoire of archaeal T4F systems beyond their currently characterized roles.