Abstract
Under nutrient limitation or stress, ribosome hibernation factors inactivate and protect ribosomes. Although ribosome hibernation plays an important role in microbes, we lack a complete understanding of this process in archaea. Here, we identify a family of hibernation factors, which we designate as single ribosomal subunit inhibitors (SriA-SriD), from the methanogenic archaeon Methanosarcina acetivorans. All four sri genes are encoded in an operon and each Sri protein inhibits protein synthesis in vitro. Deletion of sri genes in M. acetivorans impaired growth recovery after prolonged stationary phase and also led to depletion of the small ribosomal subunit. Cryo-EM structures show that Sri proteins bind to the ribosome in tandem and form conserved protein-protein interfaces. Sri is broadly distributed across archaeal phyla and sri genes frequently co-occur. Together, these findings establish Sri proteins as a distinct group of hibernation factors that protect ribosomes during dormancy and expand our understanding of ribosome hibernation in archaea.