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Abstract

Phages exhibit remarkable genetic modularity, which allows their genomes to evolve independently and combine, resulting in astounding diversity [1]. While genome mosaicism in phage populations has been studied, less is known about protein modularity and its impact on viral evolution. To fill this knowledge gap, here we quantified such modularity by detecting instances of protein mosaicism, defined as a homologous fragment between two otherwise unrelated proteins. We used highly sensitive homology detection [2] to quantify protein mosaicism between pairs of 133,574 representative phage proteins and to understand its relationship with functional diversity in phage genomes. We found that diverse functional classes often shared homologous domains which was often linked to extensive protein mosaicism, particularly in receptor-binding proteins, endolysins, and DNA polymerases. We also identified multiple instances of recent diversification via exchange and gain/loss of domains in receptor-binding proteins, neck passage structures, endolysins and some members of the core replication machinery.
To further investigate protein mosaicism in phages, we analyzed the coverage of various protein functions in domain databases such as ECOD and PFAM. We found underrepresentation of some functional groups, particularly structural proteins like tail fibers and tape measure proteins. To address this issue, we created a database of evolutionary conserved fragments (ECFs) of phage proteins. This database contains fragments not covered by known domain databases such as PFAM or ECOD, enabling better examination of the structural architecture and mosaicism of diverse phage proteins, including receptor-binding proteins, and enhancing understanding of rare accessory proteins and underrepresented proteins due to their diversity.
Our study reveals ongoing diversification via shuffling of protein domains, indicative of co-evolutionary arms races and diversifying selection towards circumventing bacterial resistance. Given the extent of protein modularity within phage proteins, we propose representing the phage pangenome as a set of ECFs, which recombine in various ways to create new proteins and genomes. This integrated understanding of protein modularity will provide novel insights into phage evolution and complex interactions with bacteria.