Nanopore guided annotation of transcriptome architectures.

Unlabelled: Nanopore direct RNA sequencing (DRS) enables the capture and full-length sequencing of native RNAs, without recoding or amplification bias. Resulting data sets may be interrogated to define the identity and location of chemically modified ribonucleotides, as well as the length of poly(A) tails, on individual RNA molecules. The success of these analyses is highly dependent on the provision of high-resolution transcriptome annotations in combination with workflows that minimize misalignments and other analysis artifacts.

Viral modulation of type II interferon increases T cell adhesion and virus spread.

During primary varicella zoster virus (VZV) infection, infected lymphocytes drive primary viremia, causing systemic dissemination throughout the host, including the skin. This results in cytokine expression, including interferons (IFNs), which partly limit infection. VZV also spreads from skin keratinocytes to lymphocytes prior to secondary viremia.

Nanopore Guided Annotation of Transcriptome Architectures.

High-resolution annotations of transcriptomes from all domains of life are essential for many sequencing-based RNA analyses, including Nanopore direct RNA sequencing (DRS), which would otherwise be hindered by misalignments and other analysis artefacts. DRS allows the capture and full-length sequencing of native RNAs, without recoding or amplification bias, and resulting data may be interrogated to define the identity and location of chemically modified ribonucleotides, as well as the length of poly(A) tails on individual RNA molecules. Existing software solutions for generating high-resolution transcriptome annotations are poorly suited to small gene dense organisms such as viruses due to the challenge of identifying distinct transcript isoforms where alternative splicing and overlapping RNAs are prevalent.

tRNA epitranscriptome determines pathogenicity of the opportunistic pathogen .

The success of bacterial pathogens depends on the coordinated expression of virulence determinants. Regulatory circuits that drive pathogenesis are complex, multilayered, and incompletely understood. Here, we reveal that alterations in tRNA modifications define pathogenic phenotypes in the opportunistic pathogen .

Histone H2A variant H2A.B is enriched in transcriptionally active and replicating HSV-1 lytic chromatin.

Unlabelled: Herpes simplex virus 1 (HSV-1) transcription is restricted in latently infected neurons and the genomes are in mostly silenced chromatin, whereas all viral genes are transcribed in lytically infected cells, in which the genomes are dynamically chromatinized. Epigenetic regulation modulates HSV-1 transcription during lytic, latent, and reactivating infections but the precise mechanisms are not fully defined. Nucleosomes are dynamic: they slide, breathe, assemble, and disassemble.

CCR4-NOT differentially controls host versus virus poly(a)-tail length and regulates HCMV infection.

Unlike most RNA and DNA viruses that broadly stimulate mRNA decay and interfere with host gene expression, human cytomegalovirus (HCMV) extensively remodels the host translatome without producing an mRNA decay enzyme. By performing a targeted loss-of-function screen in primary human fibroblasts, we here identify the host CCR4-NOT deadenylase complex members CNOT1 and CNOT3 as unexpected pro-viral host factors that selectively regulate HCMV reproduction. We find that the scaffold subunit CNOT1 is specifically required for late viral gene expression and genome-wide host responses in CCR4-NOT-disrupted cells.

Viral modulation of type II interferon increases T cell adhesion and virus spread.

During primary infection, varicella zoster virus (VZV) infects epithelial cells in the respiratory lymphoid organs and mucosa. Subsequent infection of lymphocytes, T cells in particular, causes primary viremia allowing systemic spread throughout the host, including the skin. This results in the expression of cytokines, including interferons (IFNs) which partly limit primary infection.

A Variant Allele in Varicella-Zoster Virus Glycoprotein B Selected during Production of the Varicella Vaccine Contributes to Its Attenuation.

Attenuation of the live varicella Oka vaccine (vOka) has been attributed to mutations in the genome acquired during cell culture passage of pOka (parent strain); however, the precise mechanisms of attenuation remain unknown. Comparative sequence analyses of several vaccine batches showed that over 100 single-nucleotide polymorphisms (SNPs) are conserved across all vaccine batches; 6 SNPs are nearly fixed, suggesting that these SNPs are responsible for attenuation. By contrast, prior analysis of chimeric vOka and pOka recombinants indicates that loci other than these six SNPs contribute to attenuation.

Epstein-Barr Virus-Encoded BILF1 Orthologues From Porcine Lymphotropic Herpesviruses Display Common Molecular Functionality.

Infection of immunosuppressed transplant patients with the human γ-herpesvirus Epstein-Barr virus (EBV) is associated with post-transplant lymphoproliferative disease (PTLD), an often fatal complication. Immunosuppressed miniature pigs infected with γ-herpesvirus porcine lymphotropic herpesvirus 1 (PLHV1) develop a similar disease, identifying pigs as a potential preclinical model for PTLD in humans. BILF1 is a G protein-coupled receptor (GPCR) encoded by EBV with constitutive activity linked to tumorigenesis and immunoevasive function downregulating MHC-I.

DLK-Dependent Biphasic Reactivation of Herpes Simplex Virus Latency Established in the Absence of Antivirals.

Understanding the molecular mechanisms of herpes simplex virus 1 (HSV-1) latent infection and reactivation in neurons requires the use of model systems. Establishing a quiescent infection in cultured neurons is problematic, as any infectious virus released can superinfect the cultures. Previous studies have used the viral DNA replication inhibitor acyclovir to prevent superinfection and promote latency establishment.

Nanopore-Based Detection of Viral RNA Modifications.

The chemical modification of ribonucleotides plays an integral role in the biology of diverse viruses and their eukaryotic host cells. Mapping the precise identity, location, and abundance of modified ribonucleotides remains a key goal of many studies aimed at characterizing the function and importance of a given modification. While mapping of specific RNA modifications through short-read sequencing approaches has powered a wealth of new discoveries in the past decade, this approach is limited by inherent biases and an absence of linkage information.

An eIF3d-dependent switch regulates HCMV replication by remodeling the infected cell translation landscape to mimic chronic ER stress.

Regulated loading of eIF3-bound 40S ribosomes on capped mRNA is generally dependent upon the translation initiation factor eIF4E; however, mRNA translation often proceeds during physiological stress, such as virus infection, when eIF4E availability and activity are limiting. It remains poorly understood how translation of virus and host mRNAs are regulated during infection stress. While initially sensitive to mTOR inhibition, which limits eIF4E-dependent translation, we show that protein synthesis in human cytomegalovirus (HCMV)-infected cells unexpectedly becomes progressively reliant upon eIF3d.

DRUMMER-rapid detection of RNA modifications through comparative nanopore sequencing.

Motivation: The chemical modification of ribonucleotides regulates the structure, stability and interactions of RNAs. Profiling of these modifications using short-read (Illumina) sequencing techniques provides high sensitivity but low-to-medium resolution i.e.

Single-cell transcriptomics identifies Gadd45b as a regulator of herpesvirus-reactivating neurons.

Single-cell RNA sequencing (scRNA-seq) is a powerful technique for dissecting the complexity of normal and diseased tissues, enabling characterization of cell diversity and heterogeneous phenotypic states in unprecedented detail. However, this technology has been underutilized for exploring the interactions between the host cell and viral pathogens in latently infected cells. Herein, we use scRNA-seq and single-molecule sensitivity fluorescent in situ hybridization (smFISH) technologies to investigate host single-cell transcriptome changes upon the reactivation of a human neurotropic virus, herpes simplex virus-1 (HSV-1).

Mutagenesis of the Varicella-Zoster Virus Genome Demonstrates That VLT and VLT-ORF63 Proteins Are Dispensable for Lytic Infection.

Primary varicella-zoster virus (VZV) infection leads to varicella and the establishment of lifelong latency in sensory ganglion neurons. Reactivation of latent VZV causes herpes zoster, which is frequently associated with chronic pain. Latent viral gene expression is restricted to the VZV latency-associated transcript (VLT) and VLT-ORF63 (VLT63) fusion transcripts.

The architecture of the simian varicella virus transcriptome.

Primary infection with varicella-zoster virus (VZV) causes varicella and the establishment of lifelong latency in sensory ganglion neurons. In one-third of infected individuals VZV reactivates from latency to cause herpes zoster, often complicated by difficult-to-treat chronic pain. Experimental infection of non-human primates with simian varicella virus (SVV) recapitulates most features of human VZV disease, thereby providing the opportunity to study the pathogenesis of varicella and herpes zoster in vivo.

ICTV Virus Taxonomy Profile: 2021.

Members of the family have enveloped, spherical virions with characteristic complex structures consisting of symmetrical and non-symmetrical components. The linear, double-stranded DNA genomes of 125-241 kbp contain 70-170 genes, of which 43 have been inherited from an ancestral herpesvirus. In general, herpesviruses have coevolved with and are highly adapted to their hosts, which comprise many mammalian, avian and reptilian species.

Varicella-Zoster Virus-Genetics, Molecular Evolution and Recombination.

This chapter first details the structure, organization and coding content of the VZV genome to provide a foundation on which the molecular evolution of the virus can be projected. We subsequently describe the evolution of molecular profiling approaches from restriction fragment length polymorphisms to single nucleotide polymorphism profiling to modern day high-throughput sequencing approaches. We describe how the application of these methodologies led to our current model of VZV phylogeograpy including the number and structure of geographic clades and the role of recombination in reshaping these.

Widespread remodeling of the mA RNA-modification landscape by a viral regulator of RNA processing and export.

-methyladenosine (mA) is the most abundant internal messenger RNA (mRNA) modification, contributing to the processing, stability, and function of methylated RNAs. Methylation occurs in the nucleus during pre-mRNA synthesis and requires a core methyltransferase complex consisting of METTL3, METTL14, and WTAP. During herpes simplex virus (HSV-1) infection, cellular gene expression is profoundly suppressed, allowing the virus to monopolize the host transcription and translation apparatus and antagonize antiviral responses.