Genome Modularization Reveals Overlapped Gene Topology Is Necessary for Efficient Viral Reproduction

Wright, Bradley W.; Ruan, Juanfang; Molloy, Mark P.; Jaschke, Paul R.

doi:10.1021/acssynbio.0c00323

Cited by 17 publications

(14 citation statements)

References 111 publications

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“…Bacteriophage φX174 is a member of the family Microviridae , which make up a group of small icosahedral viruses encoded by a single-stranded DNA genome. The small 5,386-nucleotide genome of φX174 containing genes encoding 11 proteins makes it a tractable target for proteomic and transcriptomic studies ( 12 , 13 ). Microviridae have been used as model systems for DNA sequencing, genome engineering ( 14 , 15 ), evolution ( 16 ), icosahedral virus packaging ( 17 , 18 ), and novel virus infection mechanisms ( 19 , 20 ).…”

Section: Introductionmentioning

confidence: 99%

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Proteomic and Transcriptomic Analysis of Microviridae φX174 Infection Reveals Broad Upregulation of Host Escherichia coli Membrane Damage and Heat Shock Responses

et al. 2021

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Measuring host-bacteriophage dynamics is an important approach to understanding bacterial survival functions and responses to infection. The model Microviridae bacteriophage φX174 is endemic to the human gut and has been studied for over 70 years, but the host response to infection has never been investigated in detail. To address this gap in our understanding of this important interaction within our microbiome, we have measured host Escherichia coli C proteomic and transcriptomic response to φX174 infection. We used mass spectrometry and RNA sequencing (RNA-seq) to identify and quantify all 11 φX174 proteins and over 1,700 E. coli proteins, enabling us to comprehensively map host pathways involved in φX174 infection. Most notably, we see significant host responses centered on membrane damage and remodeling, cellular chaperone and translocon activity, and lipoprotein processing, which we speculate is due to the peptidoglycan-disruptive effects of the φX174 lysis protein E on MraY activity. We also observe the massive upregulation of small heat shock proteins IbpA/B, along with other heat shock pathway chaperones, and speculate on how the specific characteristics of holdase protein activity may be beneficial for viral infections. Together, this study enables us to begin to understand the proteomic and transcriptomic host responses of E. coli to Microviridae infections and contributes insights to the activities of this important model host-phage interaction. IMPORTANCE A major part of the healthy human gut microbiome is the Microviridae bacteriophage, exemplified by the model φX174 phage, and their E. coli hosts. Although much has been learned from studying φX174 over the last half-century, until this work, the E. coli host response to infection has never been investigated in detail. We reveal the proteomic and transcriptomic pathways differentially regulated during the φX174 infection cycle and uncover the details of a coordinated cellular response to membrane damage that results in increased lipoprotein processing and membrane trafficking, likely due to the phage antibiotic-like lysis protein. We also reveal that small heat shock proteins IbpA/B are massively upregulated during infection and that these holdase chaperones are highly conserved across the domains of life, indicating that reliance on them is likely widespread across viruses.

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Section: Introductionmentioning

confidence: 99%

“…Bacteriophage w X174 is a member of the family Microviridae, which make up a group of small icosahedral viruses encoded by a single-stranded DNA genome. The small 5,386-nucleotide genome of w X174 containing genes encoding 11 proteins makes it a tractable target for proteomic and transcriptomic studies (12,13).…”

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confidence: 99%

Proteomic and Transcriptomic Analysis of Microviridae φX174 Infection Reveals Broad Upregulation of Host Escherichia coli Membrane Damage and Heat Shock Responses

et al. 2021

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“…Phage genomes are typically small and highly gene dense, which is likely to limit the overall complexity of their native gene expression programs [33, 34]. The phage PhiX, for instance, encodes only 11 essential genes on a genome that spans 5,000 base-pairs [35, 36].…”

Section: Discussionmentioning

confidence: 99%

Generating complex patterns of gene expression without regulatory circuits

Hill

et al. 2020

Preprint

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Synthetic biology has successfully advanced our ability to design complex, time-varying genetic circuits executing precisely specified gene expression patterns. However, such circuits usually require regulatory genes whose only purpose is to regulate the expression of other genes. When designing very small genetic constructs, such as viral genomes, we may want to avoid introducing such auxiliary gene products. To this end, here we demonstrate that varying only the placement and strengths of promoters, terminators, and RNase cleavage sites in a computational model of a bacteriophage genome is sufficient to achieve solutions to a variety of basic expression patterns. We discover these solutions by computationally evolving genomes to reproduce desired target expression patterns. Our approach shows non-trivial patterns can be evolved, including patterns in which the relative ordering of genes by abundance changes over time. We find that some patterns are easier to evolve than others, and different genomes that express comparable expression patterns may differ in their genetic architecture. Our work opens up a novel avenue to genome engineering via fine-tuning the balance of gene expression and gene degradation rates.

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“…The historic case of overlapped genes in phage ϕX174 (Sanger et al, 1977) has been updated recently. Data from modified genomes led us to conclude that gene overlapping in the phage genome is essential for the correct expression of proteins and DNA replication (Wright et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

Transcriptional analysis in bacteriophage Fc02 of Pseudomonas aeruginosa revealed two overlapping genes with exclusion activity

Ramírez-Sánchez

Magos-Castro²,

Guarneros³

2023

Front. Microbiol.

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Little is known about the gene expression program during the transition from lysogenic to lytic cycles of temperate bacteriophages in Pseudomonas aeruginosa. To investigate this issue, we developed a thermo-sensitive repressor mutant in a lysogen and analyzed the phage transcriptional program by strand-specific RNA-Seq before and after thermo-induction. As expected, the repressor gene located on the phage DNA forward strand is transcribed in the lysogen at the permissive temperature of 30°C. Upstream the repressor gene, we noticed the presence of two overlapped ORFs apparently in the same transcript. One ORF is a gene that encodes a protein of 7.9 kDa mediating the exclusion of various super-infecting phages. The other ORF, placed in an alternate reading frame with a possible AUG initiation codon at 25 nucleotide downstream of the AUG of the first gene, is expected to encode a 20.7 kDa polypeptide of yet an unknown function. Upon lifting repression at 40°C, the transcription of an operon which is involved in the lytic cycle is started from a promoter on the reverse phage DNA strand. The first gene in the operon is a homolog of the antirepresor ner, a common gene in the lysis–lysogeny regulation region of other phages. Interestingly, the next gene after ner is gene 10 that on the reverse strand overlaps the overlapped gene olg1 on the forward strand. Curiously, gene 10 expression also shows superinfection exclusion. Strand-specific RNA-Seq also has uncovered the transcription succession of gene modules expressed during the phage lytic stage. The conservation of overlapped genes with similar functions may be evolutionarily selected.

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Genome Modularization Reveals Overlapped Gene Topology Is Necessary for Efficient Viral Reproduction

Cited by 17 publications

References 111 publications

Proteomic and Transcriptomic Analysis of Microviridae φX174 Infection Reveals Broad Upregulation of Host Escherichia coli Membrane Damage and Heat Shock Responses

Proteomic and Transcriptomic Analysis of Microviridae φX174 Infection Reveals Broad Upregulation of Host Escherichia coli Membrane Damage and Heat Shock Responses

Generating complex patterns of gene expression without regulatory circuits

Transcriptional analysis in bacteriophage Fc02 of Pseudomonas aeruginosa revealed two overlapping genes with exclusion activity

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