The Phage Nucleus and PhuZ Spindle: Defining Features of the Subcellular Organization and Speciation of Nucleus-Forming Jumbo Phages

Abstract

Bacteriophages and their bacterial hosts are ancient organisms that have been co-evolving for billions of years. Some jumbo phages, those with a genome size larger than 200 kilobases, have recently been discovered to establish complex subcellular organization during replication. Here, we review our current understanding of jumbo phages that form a nucleus-like structure, or “Phage Nucleus,” during replication. The phage nucleus is made of a proteinaceous shell that surrounds replicating phage DNA and imparts a unique subcellular organization that is temporally and spatially controlled within bacterial host cells by a phage-encoded tubulin (PhuZ)-based spindle. This subcellular architecture serves as a replication factory for jumbo Pseudomonas phages and provides a selective advantage when these replicate in some host strains. Throughout the lytic cycle, the phage nucleus compartmentalizes proteins according to function and protects the phage genome from host defense mechanisms. Early during infection, the PhuZ spindle positions the newly formed phage nucleus at midcell and, later in the infection cycle, the spindle rotates the nucleus while delivering capsids and distributing them uniformly on the nuclear surface, where they dock for DNA packaging. During the co-infection of two different nucleus-forming jumbo phages in a bacterial cell, the phage nucleus establishes Subcellular Genetic Isolation that limits the potential for viral genetic exchange by physically separating co-infection genomes, and the PhuZ spindle causes Virogenesis Incompatibility, whereby interacting components from two diverging phages negatively affect phage reproduction. Thus, the phage nucleus and PhuZ spindle are defining cell biological structures that serve roles in both the life cycle of nucleus-forming jumbo phages and phage speciation.

Subcellular organization of viral particles during maturation of nucleus-forming jumbo phage

“…Our updated model for jumbo phage replication, shown in Fig. 6 together with our proposed model ( 40 ), begins with the attachment of phage to the host cell and injection of viral DNA. The expression of the nuclear shell protein occurs immediately after injection to form an enclosure around phage DNA that provides a safe compartment for DNA replication to occur.…”

“…Viral capsid proteins assemble on the membrane midway through the infection cycle and are transported by treadmilling PhuZ filaments to the phage nucleus, where they dock to initiate DNA packaging. Treadmilling filaments rotate the phage nucleus to allow capsids to be evenly distributed on its surface ( 40 ). Once capsids are full of viral DNA, they coassemble with tails and form bouquets containing what appear to be mature particles.…”

Phage-resistant Pseudomonas aeruginosa against a novel lytic phage JJ01 exhibits hypersensitivity to colistin and reduces biofilm production

“…It is possible that different Pseudomonas phages isolated at different periods and from distant geographical areas contain high genome sequence similarities. For example, distinct giant phages that were classified in different families (siphophage SD1-M and nucleus-forming myophage phiKZ) shared a very high genome identity up to 99% with only two DNA segments different between their genomes ( Kwan et al, 2006 ; Krylov et al, 2007 ; Chaikeeratisak et al, 2021 ). Specifically, in the Pbunavirus genus, PB1-like phages, which are the very common virulent P. aeruginosa phages, shared a remarkable genome similarity to one another (over 95%).…”

A cytoskeletal vortex drives phage nucleus rotation during jumbo phage replication in E. coli

“…Subcellular organization is essential for all domains of life, including viruses ( Chaikeeratisak et al, 2021a ; Charman and Weitzman, 2020 ). We recently identified a nucleus-like structure ( Chaikeeratisak et al, 2017b ) formed by Pseudomonas chlororaphis jumbo phage 201φ2–1 ( Thomas et al, 2008 , 2010 , 2012 ), which was subsequently shown to be conserved ( Chaikeeratisak et al, 2017a ; Danilova et al, 2020 ) in related phages ΦKZ ( Krylov et al, 2021 ; Lecoutere et al, 2009 ; Mesyanzhinov et al, 2002 ; Weintraub et al, 2019 ), and ΦPA3 ( Monson et al, 2011 ).…”