Conserved jumbo phage factors required for protein import into a phage nucleus

Kokontis, Claire; Klein, Timothy A; Silas, Sukrit; Bondy-Denomy, Joseph

doi:10.1101/2024.03.27.586873

Cited by 4 publications

(3 citation statements)

References 24 publications

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“…The fact that knockdown of PicA caused the same transcriptional phenotype as knockdown of ChmA and nvRNAP suggests that the nuclear import of nvRNAP subunits might be dependent on the recently discovered PicA protein importer (Morgan et al 2024, Kokontis et al 2024).…”

Section: Rna-seq After Aso Knockdown Reveals Molecular Phenotypes Of ...mentioning

confidence: 97%

“…Silencing of PicA (ΦKZ069), which is required for protein import into the phage nucleus (Morgan et al 2024, Kokontis et al 2024) abrogated plaque formation as well. We also observed strong effects upon knockdown of uncharacterised genes.…”

Section: Systematic Aso Screen For Factors Important For Phage Replic...mentioning

confidence: 99%

“…Several conserved phage factors enable formation and organisation of the phage nucleus, e.g., the shell protein chimallin (ChmA) (Chaikeeratisak et al 2017a, Laughlin et al 2022, Nieweglowska et al 2023, PicA, which mediates cargo trafficking across the phage nucleus (Morgan et al 2024, Kokontis et al 2024, and the tubulinlike protein PhuZ, which centres the phage nucleus in the middle of the cell and is involved in intracellular trafficking of newly assembled capsids (Erb et al 2014, Chaikeeratisak et al 2017a, Chaikeeratisak et al 2017b, Chaikeeratisak et al 2019.…”

Section: Introductionmentioning

confidence: 99%

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Non-genetic messenger RNA silencing reveals essential genes in phage-host interplay

Gerovac,

Ðurica-Mitić,

Buhlmann

et al. 2024

Preprint

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Bacteriophages are the most abundant entities on earth and exhibit vast genetic and phenotypic diversity. Exploitation of this largely unexplored molecular space requires identification and functional characterisation of genes that act at the phage-host interface. To date, this is restricted to few model phage-host systems that are amenable to genetic manipulation. To overcome this limitation, we introduce a non-genetic mRNA targeting approach using exogenous delivery of programmable antisense oligomers (ASOs) to silence genes of both DNA and RNA phages. A systematic knockdown screen of core and accessory genes of the nucleus-forming jumbo phage ΦKZ, coupled to RNA-sequencing and microscopy analyses, reveals previously unrecognised proteins that are essential for phage replication and that, upon silencing, elicit distinct phenotypes at the level of the phage and host response. One of these factors is a ΦKZ-encoded nuclease that acts at a major decision point during infection, linking the formation of the protective phage nucleus to phage genome amplification. The non-genetic ASO-based gene silencing used here promises to be a versatile tool for molecular discovery in phage biology, will help elucidate defence and anti-defence mechanisms in non-model phage-host pairs, and offers potential for optimising phage therapy and biotechnological procedures.

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Section: Rna-seq After Aso Knockdown Reveals Molecular Phenotypes Of ...mentioning

confidence: 97%

Section: Systematic Aso Screen For Factors Important For Phage Replic...mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Non-genetic messenger RNA silencing reveals essential genes in phage-host interplay

Gerovac,

Ðurica-Mitić,

Buhlmann

et al. 2024

Preprint

View full text Add to dashboard Cite

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Genome-wide identification of bacterial genes contributing to nucleus-forming jumbo phage infection

Harding,

Malone,

Kyte

et al. 2024

Nucleic Acids Research

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The Chimalliviridae family of bacteriophages (phages) form a proteinaceous nucleus-like structure during infection of their bacterial hosts. This phage ‘nucleus’ compartmentalises phage DNA replication and transcription, and shields the phage genome from DNA-targeting defence systems such as CRISPR-Cas and restriction-modification. Their insensitivity to DNA-targeting defences makes nucleus-forming jumbo phages attractive for phage therapy. However, little is known about the bacterial gene requirements during the infectious cycle of nucleus-forming phages or how phage resistance may emerge. To address this, we used the Serratia nucleus-forming jumbo phage PCH45 and exploited a combination of high-throughput transposon mutagenesis and deep sequencing (Tn-seq), and CRISPR interference (CRISPRi). We identified over 90 host genes involved in nucleus-forming phage infection, the majority of which were either involved in the biosynthesis of the primary receptor, flagella, or influenced swimming motility. In addition, the bacterial outer membrane lipopolysaccharide contributed to PCH45 adsorption. Other unrelated Serratia-flagellotropic phages used similar host genes as the nucleus-forming phage, indicating that phage resistance can lead to cross-resistance against diverse phages. Our findings demonstrate that resistance to nucleus-forming jumbo phages can readily emerge via bacterial surface receptor mutation and this should be a major factor when designing strategies for their use in phage therapy.

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An intron endonuclease facilitates interference competition between coinfecting viruses

Birkholz,

Morgan,

Laughlin

et al. 2024

Science

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Introns containing homing endonucleases are widespread in nature and have long been assumed to be selfish elements that provide no benefit to the host organism. These genetic elements are common in viruses, but whether they confer a selective advantage is unclear. In this work, we studied intron-encoded homing endonuclease gp210 in bacteriophage ΦPA3 and found that it contributes to viral competition by interfering with the replication of a coinfecting phage, ΦKZ. We show that gp210 targets a specific sequence in ΦKZ, which prevents the assembly of progeny viruses. This work demonstrates how a homing endonuclease can be deployed in interference competition among viruses and provide a relative fitness advantage. Given the ubiquity of homing endonucleases, this selective advantage likely has widespread evolutionary implications in diverse plasmid and viral competition as well as virus-host interactions.

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Conserved jumbo phage factors required for protein import into a phage nucleus

Cited by 4 publications

References 24 publications

Non-genetic messenger RNA silencing reveals essential genes in phage-host interplay

Non-genetic messenger RNA silencing reveals essential genes in phage-host interplay

Genome-wide identification of bacterial genes contributing to nucleus-forming jumbo phage infection

An intron endonuclease facilitates interference competition between coinfecting viruses

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