An essential and highly selective protein import pathway encoded by nucleus-forming phage

Abstract

Targeting proteins to specific subcellular destinations is essential in prokaryotes, eukaryotes, and the viruses that infect them. Chimalliviridae phages encapsulate their genomes in a nucleus-like replication compartment composed of the protein chimallin (ChmA) that excludes ribosomes and decouples transcription from translation. These phages selectively partition proteins between the phage nucleus and the bacterial cytoplasm. Currently, the genes and signals that govern selective protein import into the phage nucleus are unknown. Here, we identify two components of this protein import pathway: a species-specific surface-exposed region of a phage intranuclear protein required for nuclear entry and a conserved protein, PicA (Protein importer of chimalliviruses A), that facilitates cargo protein trafficking across the phage nuclear shell. We also identify a defective cargo protein that is targeted to PicA on the nuclear periphery but fails to enter the nucleus, providing insight into the mechanism of nuclear protein trafficking. Using CRISPRi-ART protein expression knockdown of PicA, we show that PicA is essential early in the chimallivirus replication cycle. Together, our results allow us to propose a multistep model for the Protein Import Chimallivirus pathway, where proteins are targeted to PicA by amino acids on their surface and then licensed by PicA for nuclear entry. The divergence in the selectivity of this pathway between closely related chimalliviruses implicates its role as a key player in the evolutionary arms race between competing phages and their hosts.

An intron endonuclease facilitates interference competition between coinfecting viruses

“…These two related proteins are only imported by their cognate phage during infections with either ΦKZ or ΦPA3 (“ΦPA3” and “ΦKZ,” Fig. 4, D and E) ( 25 ). However, during coinfections, 46.4% ( N = 140) of nuclei formed only imported one protein, whereas the other 53.6% imported both proteins (“ΦKZ + ΦPA3,” Fig.…”

“…These phages form a proteinaceous phage nucleus ( 19 – 21 ) that shields their DNA genomes from host defenses, ( 22 – 24 ) and, similar to the eukaryotic nucleus, these phages can strictly separate transcription and DNA replication from translation and metabolic processes. Certain proteins are selectively imported into the phage nucleus, whereas others are excluded, and the selectivity of this process is determined by the conserved chimallivirus protein PicA ( 25 , 26 ). The ΦPA3-encoded endonuclease gp210 is naturally imported into the phage nucleus of ΦPA3 but is excluded from that of ΦKZ ( 17 ).…”