Metagenomic mining reveals novel viral histones in dsDNA viruses

The compaction of genomic DNA into nucleosomes with the help of histones has long been considered a fundamental feature exclusive to eukaryotic cells. However, it was recently shown that archaea and bacteria also encode histones. A complex picture has emerged with more recent discoveries of eukaryotic-like histones within one phylum of double-stranded DNA (dsDNA) viruses. Nevertheless, the extent to which other dsDNA viruses encode histones remains largely unexplored. Here, we conducted a metagenomic survey of viral histones that were further clustered based on sequence and predicted structural similarities. We identified over 1500 viral histones and histone-fold proteins, including previously undescribed proteins found in the viral class Caudoviricetes. Structural predictions and in vitro assays demonstrated that histone triplets (where three histone folds are fused) and singlets co-occurring in the same viral genome are capable of forming nucleosome-like particles. Beyond nucleosomal histone functions, our analysis revealed six types of structurally and functionally diverse viral histone-fold proteins, some of which do not have known structural or functional homologs. Altogether, our findings reveal a previously unrecognized diversity of viral histones in dsDNA viruses, expanding the known repertoire, structural diversity, and functional versatility of viral histones beyond nucleocytoplasmic large DNA viruses. • Histone triplets and singlets co-occurring in the same double-stranded DNA viral genomes assemble into nucleosome-like particles. • Six novel types of structurally and functionally distinct viral histone-fold proteins are identified. • The presence of viral histones is significantly positively associated with proteins containing sucrose nonfermenting 2 domain in viral genomes.