Biology subjects

Jahun, A. S.

Publications and source records attributed to Jahun, A. S..

8 recordsLinked to original sources

bioRxiv2024

Encephalomyocarditis virus protein 2B* antagonises innate immune signalling by interacting with 14-3-3 protein family members

Encephalomyocarditis virus (EMCV) has for decades served as an important model RNA virus. Although most of the EMCV proteins are obtained via proteolytic cleavage of a long polyprotein, 2B* is expressed from a short overlapping open reading frame via an unusual protein-stimulated temporally dependent ribosomal frameshifting mechanism. The function of 2B* has not yet been characterised, though mutant viruses that are unable to express 2B* have a small plaque phenotype. Here we show that 2B* binds all seven members of the 14-3-3 protein family during virus infection. Binding is dependent on the 2B* C-terminal sequence RRNSS. IFN-{beta} and IL-6 signalling are impeded following overexpression of 2B* but not a truncated version lacking the RRNSS residues, thus suggesting a 14-3-3-dependent role for 2B* in inhibiting MAVS signalling. We also find that this function is distinct from the effect of 2B* on plaque size, as a virus in which 2B* was similarly truncated exhibited near-wildtype plaque size, thus indicating that 2B* also harbours additional functions. This work provides the first identification of a role of 2B* in innate immune antagonism and expands our knowledge of the protein complement of this important model virus. IMPORTANCEEncephalomyocarditis virus (EMCV) infects a range of species, causing economically important reproductive disorders in pigs and encephalitis and myocarditis in rodents. Due to its wide host range, it is an important model pathogen for investigating virus-host interactions. EMCV expresses an accessory protein, 2B*, from an overlapping open reading frame via an unusual ribosomal frameshifting mechanism. Although the frameshifting mechanism has been established, the function of the 2B* protein had not previously been explored. Here, we determined the host proteins to which 2B* binds and found that it specifically binds to the entire 14-3-3 protein family which, among other roles, contribute to the innate immune response to viral infection in mammalian cells. This interaction requires a specific stretch of amino acids at the end of 2B*. By interacting with the 14-3-3 proteins, 2B* blocks immune response activation. Thus, 2B* is a novel antagonist of innate immunity.

microbiology↗

bioRxiv2024

Variable rates of SARS-CoV-2 evolution in chronic infections

An important feature of the evolution of the SARS-CoV-2 virus has been the emergence of highly mutated novel variants, which are characterised by the gain of multiple mutations relative to viruses circulating in the general global population. Cases of chronic viral infection have been suggested as an explanation for this phenomenon, whereby an extended period of infection, with an increased rate of evolution, creates viruses with substantial genetic novelty. However, measuring a rate of evolution during chronic infection is made more difficult by the potential existence of compartmentalisation in the viral population, whereby the viruses in a host form distinct subpopulations. We here describe and apply a novel statistical method to study within-host virus evolution, identifying the minimum number of subpopulations required to explain sequence data observed from cases of chronic infection, and inferring rates for within-host viral evolution. Across nine cases of chronic SARS-CoV-2 infection in hospitalised patients we find that non-trivial population structure is relatively common, with four cases showing evidence of more than one viral population evolving independently within the host. We find cases of within-host evolution proceeding significantly faster, and significantly slower, than that of the global SARS-CoV-2 population, and of cases in which viral subpopulations in the same host have statistically distinguishable rates of evolution. Non-trivial population structure was associated with high rates of within-host evolution that were systematically underestimated by a more standard inference method.

evolutionary biology↗

bioRxiv2023

Porcine sapovirus protease controls the innate immune response and targets TBK1

Human sapoviruses (HuSaVs) and noroviruses are considered the leading cause of acute gastroenteritis worldwide. While extensive research has focused on noroviruses, our understanding of sapoviruses (SaVs) and their interactions with the hosts immune response remains limited. HuSaVs have been challenging to propagate in vitro, making the porcine sapovirus (PSaV) Cowden strain a valuable model for studying SaV pathogenesis. In this study we show, for the first time, that PSaV Cowden strain has mechanisms to evade the hosts innate immune response. The virus 3C-like protease (NS6) inhibits type I IFN production by targeting TBK1. Catalytically active NS6, both during ectopic expression and during PSaV infection, targets TBK1 which is then led for rapid degradation by the proteasome. Moreover, deletion of TBK1 from porcine cells led to a significant increase in PSaV titres, emphasizing its role in regulating PSaV infection. Additionally, we successfully established PSaV infection in IPEC-J2 cells, an enterocytic cell line originating from the jejunum of a neonatal piglet. Overall, this study provides novel insights into PSaV evasion strategies, opening the way for future investigations for SaV-host interactions, and enabling the use of a new cell line model for PSaV research.

microbiology↗

bioRxiv2022

TRIM7 restricts Coxsackievirus and norovirus infection by detecting the C-terminalglutamine generated by 3C protease processing

TRIM7 catalyses the ubiquitination of multiple substrates with unrelated biological functions. This cross-reactivity is at odds with the specificity usually displayed by enzymes, including ubiquitin ligases. Here we show that TRIM7s extreme substrate promiscuity is due to a highly unusual binding mechanism, in which the PRYSPRY domain captures any ligand with a C-terminal helix that terminates in a hydrophobic residue followed by a glutamine. Many of the non-structural proteins found in RNA viruses contain C-terminal glutamines as a result of polyprotein cleavage by 3C protease. This viral processing strategy generates novel substrates for TRIM7 and explains its ability to inhibit Coxsackie virus and norovirus replication. In addition to viral proteins, cellular proteins such as glycogenin have evolved C-termini that make them a TRIM7 substrate. The helix-{Phi}Q degron motif recognised by TRIM7 is reminiscent of the N-end degron system and is found in [~] 1% of cellular proteins. These features, together with TRIM7s restricted tissue expression and lack of immune regulation suggest that viral restriction may not be its physiological function.

biochemistry↗

bioRxiv2021

Leaked genomic and mitochondrial DNA contribute to the host response to noroviruses in a STING-dependent manner

The cGAS-STING pathway is central to the IFN response against DNA viruses. However, recent studies are increasingly demonstrating its role in the restriction of some RNA viruses. Here we show that the cGAS-STING pathway also contributes to the IFN response against noroviruses, positive-sense single-stranded RNA viruses that are now one of the most common causes of infectious gastroenteritis world-wide. We show a significant reduction in IFN-{beta} induction and a corresponding increase in viral replication in norovirus-infected cells following STING inhibition, knockdown or deletion. Upstream of STING, we show that cells lacking either cGAS or IFI16 also have severely impaired IFN responses. Further, we demonstrate that immunostimulatory host genome-derived DNA, and to a lesser extent mitochondrial DNA, accumulate in the cytosol of norovirus-infected cells. And lastly, overexpression of the viral NS4 protein was sufficient to drive the accumulation of cytosolic DNA. Together, our data elucidate a role for cGAS, IFI16 and STING in the restriction of noroviruses, and demonstrate for the first time the utility of host genomic DNA as a damage-associated molecular pattern in cells infected with an RNA virus. HighlightsO_LIcGAS, IFI16 and STING are required for a robust IFN response against noroviruses C_LIO_LINuclear and mitochondrial DNA accumulate in the cytosols of infected cells C_LIO_LIViral NS4 mediates accumulation of cytosolic DNA C_LI

molecular biology↗

bioRxiv2021

Evolution of enhanced innate immune evasion by the SARS-CoV-2 B.1.1.7 UK variant

Emergence of SARS-CoV-2 variants, including the globally successful B.1.1.7 lineage, suggests viral adaptations to host selective pressures resulting in more efficient transmission. Although much effort has focused on Spike adaptation for viral entry and adaptive immune escape, B.1.1.7 mutations outside Spike likely contribute to enhance transmission. Here we used unbiased abundance proteomics, phosphoproteomics, mRNA sequencing and viral replication assays to show that B.1.1.7 isolates more effectively suppress host innate immune responses in airway epithelial cells. We found that B.1.1.7 isolates have dramatically increased subgenomic RNA and protein levels of Orf9b and Orf6, both known innate immune antagonists. Expression of Orf9b alone suppressed the innate immune response through interaction with TOM70, a mitochondrial protein required for RNA sensing adaptor MAVS activation, and Orf9b binding and activity was regulated via phosphorylation. We conclude that B.1.1.7 has evolved beyond the Spike coding region to more effectively antagonise host innate immune responses through upregulation of specific subgenomic RNA synthesis and increased protein expression of key innate immune antagonists. We propose that more effective innate immune antagonism increases the likelihood of successful B.1.1.7 transmission, and may increase in vivo replication and duration of infection.

microbiology↗

bioRxiv2020

Patterns of within-host genetic diversity in SARS-CoV-2

Monitoring the spread of SARS-CoV-2 and reconstructing transmission chains has become a major public health focus for many governments around the world. The modest mutation rate and rapid transmission of SARS-CoV-2 prevents the reconstruction of transmission chains from consensus genome sequences, but within-host genetic diversity could theoretically help identify close contacts. Here we describe the patterns of within-host diversity in 1,181 SARS-CoV-2 samples sequenced to high depth in duplicate. 95% of samples show within-host mutations at detectable allele frequencies. Analyses of the mutational spectra revealed strong strand asymmetries suggestive of damage or RNA editing of the plus strand, rather than replication errors, dominating the accumulation of mutations during the SARS-CoV-2 pandemic. Within and between host diversity show strong purifying selection, particularly against nonsense mutations. Recurrent within-host mutations, many of which coincide with known phylogenetic homoplasies, display a spectrum and patterns of purifying selection more suggestive of mutational hotspots than recombination or convergent evolution. While allele frequencies suggest that most samples result from infection by a single lineage, we identify multiple putative examples of co-infection. Integrating these results into an epidemiological inference framework, we find that while sharing of within-host variants between samples could help the reconstruction of transmission chains, mutational hotspots and rare cases of superinfection can confound these analyses.

genomics↗

bioRxiv2019

Murine norovirus virulence factor 1 (VF1) protein contributes to viral fitness during persistent infection

BackgroundMurine norovirus (MNV) is widely used as a model for studying norovirus biology. While MNV isolates vary in their pathogenesis, infection of immunocompetent mice mostly results in persistent infection. The ability of a virus to establish a persistent infection is dependent on its ability to subvert or avoid the host immune response. Previously, we described the identification and characterization of virulence factor 1 (VF1) in MNV, and established that it acts as an innate immune antagonist. Here, we explore the role of VF1 during persistent MNV infection in an immunocompetent host.\n\nMethodsUsing reverse genetics, we generated VF1-knockout MNV-3 that contained a single or a triple termination codon inserted in the VF1 open reading frame.\n\nResultsVF1-knockout MNV-3 replicated to comparable levels to the wildtype virus in tissue culture. Comparative studies between MNV-3 and an acute MNV-1 strain show that MNV-3 VF1 exerts the same functions as MNV-1 VF1, but with reduced potency. Mice infected with VF1-knockout MNV-3 showed significantly reduced replication kinetics during the acute phase of the infection, but viral loads rapidly reached the levels seen in mice infected with wildtype virus after phenotypic restoration of VF1 expression. Infection with an MNV-3 mutant that had three termination codons inserted into VF1, in which reversion was suppressed, resulted in consistently lower replication throughout a three-month persistent infection in mice, suggesting a role for VF1 in viral fitness in vivo.\n\nConclusionOur results indicate that VF1 expressed by a persistent strain of MNV also functions to antagonise the innate response to infection. We found that VF1 is not essential for viral persistence, but instead contributes to viral fitness in mice. These data fit with the hypothesis that noroviruses utilise multiple mechanisms to avoid and/or control the host response to infection and that VF1 is just one component of this.

microbiology↗