A Family of Viral Satellites Manipulates Invading Virus Gene Expression and Can Affect Cholera Toxin Mobilization

Abstract

Many viruses possess temporally unfolding gene expression patterns aimed at subverting host defenses, commandeering host metabolism, and ultimately producing a large number of progeny virions. High-throughput omics tools, such as RNA sequencing (RNA-seq), have dramatically enhanced the resolution of expression patterns during infection. Less studied have been viral satellites, mobile genomes that parasitize viruses. By performing RNA-seq on infection time courses, we have obtained the first time-resolved transcriptomes for bacteriophage satellites during lytic infection. Specifically, we have acquired transcriptomes for the lytic Vibrio cholerae phage ICP1 and all five known variants of ICP1’s parasite, the phage inducible chromosomal island-like elements (PLEs). PLEs rely on ICP1 for both DNA replication and mobilization and abolish production of ICP1 progeny in infected cells. We investigated PLEs’ impact on ICP1 gene expression and found that PLEs did not broadly restrict or reduce ICP1 gene expression. A major exception occurred in ICP1’s capsid morphogenesis operon, which was downregulated by each of the PLE variants. Surprisingly, PLEs were also found to alter the gene expression of CTXΦ, the integrative phage that encodes cholera toxin and is necessary for virulence of toxigenic V. cholerae. One PLE, PLE1, upregulated CTXΦ genes involved in replication and integration and boosted CTXΦ mobility following induction of the SOS response. IMPORTANCE Viral satellites are found in all domains of life and can have profound fitness effects on both the viruses they parasitize and the cells they reside in. In this study, we have acquired the first RNA sequencing (RNA-seq) transcriptomes of viral satellites outside plants, as well as the transcriptome of the phage ICP1, a predominant predator of pandemic Vibrio cholerae. Capsid downregulation, previously observed in an unrelated phage satellite, is conserved among phage inducible chromosomal island-like elements (PLEs), suggesting that viral satellites are under strong selective pressure to reduce the capsid expression of their larger host viruses. Despite conserved manipulation of capsid expression, PLEs exhibit divergent effects on CTXΦ transcription and mobility. Our results demonstrate that PLEs can influence both their hosts’ resistance to phage and the mobility of virulence-encoding elements, suggesting that PLEs can play a substantial role in shaping Vibrio cholerae evolution.

A phage satellite tunes inducing phage gene expression using a domesticated endonuclease to balance inhibition and virion hijacking

“…Western blot analysis of total protein from PLE(−) and PLE(+) hosts over the course of infection demonstrated detectable capsid protein by 12 minutes post-infection and confirmed that PLE(+) infections produce approximately one-third as much ICP1 capsid protein as PLE(−) infections at all timepoints where capsid protein was detected (Figure 1D, Supplementary Figure S3A). These results together with the capsid operon reporter phage validate the observation of capsid operon transcriptional repression mediated by PLE (21) and confirm that this transcriptional repression results in a reduction in total ICP1 capsid monomer production during the late stages of a PLE(+) infection.…”

“…This requirement was dictated by the unique challenges presented by the lifecycle of ICP1 within a V. cholerae host cell, which is rapid and highly destructive: infections produce ~90 viable phage progeny per infected cell in under 20 minutes (15). The bacterial host chromosome is rapidly degraded (26) and transcriptional profiling during infection did not reveal any ICP1, PLE, or V. cholerae genes with consistent transcriptional activity over the infection time-course (21). This limits feasibility of evaluating the targeted repression of the capsid operon by RT-qPCR, which would require a proper normalization control.…”

“…We observed that a single operon in the ICP1 genome is transcriptionally downregulated in a PLE(+) host. The three-fold downregulated operon ( gp126-gp122 , from here forward referred to as the capsid operon) encodes ICP1’s capsid morphogenesis proteins including the major capsid subunit (Gp122), putative capsid decoration protein (Gp123), putative capsid maturation protease (Gp125) and other proteins with unknown function (Figure 1A) (21). This subtle and specific impact on ICP1 transcription was surprising given that PLE activity during ICP1 infection restricts ICP1 DNA replication and completely eliminates production of phage progeny (15,17).…”

“…This subtle and specific impact on ICP1 transcription was surprising given that PLE activity during ICP1 infection restricts ICP1 DNA replication and completely eliminates production of phage progeny (15,17). Capsid operon repression is a conserved feature of PLEs: five PLEs with unique allelic variations identified in V. cholerae isolates all reduce transcript and protein produced from the capsid operon (21). This work will refer to the modern dominant PLE1 as “PLE” from here forward unless otherwise specified.…”

IncC helper dependent plasmid-like replication of Salmonella Genomic Island 1

“…Further similarities are that the excised PLE overreplicates its helper phage, outperforms the phage genome in packaging and consequently, highly reduces the spread of the helper phage ( 59 ). Thereby PLEs provide efficient defence for their host against its lytic phage invaders and ensure their own horizontal transfer at the expense of the helper phage ( 59 , 60 ). PLE also has its own replication origin and encodes a RepA resembling Gram+ plasmid replication initiation factors.…”