Viral Modulation of Host Translation and Implications for Vaccine Development

Translation of mRNAs into protein is an essential mechanism of regulating gene expression—and a step exploited by viruses for their own propagation. In this article, we review mechanisms that govern translation and provide an overview of the translation machinery, discuss some of the components involved in this process, and discuss how viruses modulate host translational controls and implications in vaccine design

Drug repositioning of Clopidogrel or Triamterene to inhibit influenza virus replication in vitro

Influenza viruses cause respiratory tract infections and substantial health concerns. Infection may result in mild to severe respiratory disease associated with morbidity and some mortality. Several anti-influenza drugs are available, but these agents target viral components and are susceptible to drug resistance. There is a need for new antiviral drug strategies that include repurposing of clinically approved drugs. Drugs that target cellular machinery necessary for influenza virus replication can provide a means for inhibiting influenza virus replication. We used RNA interference screening to identify key host cell genes required for influenza replication, and then FDA-approved drugs that could be repurposed for targeting host genes. We examined the effects of Clopidogrel and Triamterene to inhibit A/WSN/33 (EC(50) 5.84 uM and 31.48 uM, respectively), A/CA/04/09 (EC(50) 6.432 uM and 3.32 uM, respectively), and B/Yamagata/16/1988 (EC(50) 0.28 uM and 0.11 uM, respectively) replication. Clopidogrel and Triamterene provide a druggable approach to influenza treatment across multiple strains and subtypes

MicroRNA and Nonsense Transcripts as Putative Viral Evasion Mechanisms

Viral proteins encode numerous antiviral activities to modify the host immunity. In this article, we hypothesize that viral genomes and gene transcripts interfere with host gene expression using passive mechanisms to deregulate host microRNA (miRNA) activity. We postulate that various RNA viruses mimic or block binding between a host miRNA and its target transcript, a phenomenon mediated by the miRNA seed site at the 5′ end of miRNA. Virus-encoded miRNA seed sponges (vSSs) can potentially bind to host miRNA seed sites and prevent interaction with their native targets thereby relieving native miRNA suppression. In contrast, virus-encoded miRNA seed mimics (vSMs) may mediate considerable downregulation of host miRNA activity. We analyzed genomes from diverse RNA viruses for vSS and vSM signatures and found an abundance of these motifs indicating that this may be a mechanism of deceiving host immunity. Employing respiratory syncytial virus and measles virus as models, we reveal that regions surrounding vSS or vSM motifs have features characteristics of pre-miRNA templates and show that RSV viral transcripts are processed into small RNAs that may behave as vSS or vSM effectors. These data suggest that complex molecular interactions likely occur at the host-virus interface. Identifying the mechanisms in the network of interactions between the host and viral transcripts can help uncover ways to improve vaccine efficacy, therapeutics, and potentially mitigate the adverse events that may be associated with some vaccines

MicroRNAs affect GPCR and Ion channel genes needed for influenza replication

Influenza virus causes seasonal epidemics and sporadic pandemics resulting in morbidity, mortality, and economic losses worldwide. Understanding how to regulate influenza virus replication is important for developing vaccine and therapeutic strategies. Identifying microRNAs (miRs) that affect host genes used by influenza virus for replication can support an antiviral strategy. In this study, G-protein coupled receptor (GPCR) and ion channel (IC) host genes in human alveolar epithelial (A549) cells used by influenza virus for replication (Orr-Burks et al., 2021) were examined as miR target genes following A/CA/04/09- or B/Yamagata/16/1988 replication. Thirty-three miRs were predicted to target GPCR or IC genes and their miR mimics were evaluated for their ability to decrease influenza virus replication. Paired miR inhibitors were used as an ancillary measure to confirm or not the antiviral effects of a miR mimic. Fifteen miRs lowered influenza virus replication and four miRs were found to reduce replication irrespective of virus strain and type differences. These findings provide evidence for novel miR disease intervention strategies for influenza viruses

The central conserved region (CCR) of respiratory syncytial virus (RSV) G protein modulates host miRNA expression and alters the cellular response to infection

Respiratory Syncytial Virus (RSV) infects respiratory epithelial cells and deregulates host gene expression by many mechanisms including expression of RSV G protein (RSV G). RSV G protein encodes a central conserved region (CCR) containing a CX3C motif that functions as a fractalkine mimic. Disruption of the CX3C motif (a.a. 182–186) located in the CCR of the G protein has been shown to affect G protein function in vitro and the severity of RSV disease pathogenesis in vivo. We show that infection of polarized Calu3 respiratory cells with recombinant RSV having point mutations in Cys173 and 176 (C173/176S) (rA2-GC12), or Cys186 (C186S) (rA2-GC4) is associated with a decline in the integrity of polarized Calu-3 cultures and decreased virus production. This is accompanied with downregulation of miRNAs let-7f and miR-24 and upregulation of interferon lambda (IFNλ), a primary antiviral cytokine for RSV in rA2-GC12/rA2-GC4 infected cells. These results suggest that residues in the cysteine noose region of RSV G protein can modulate IFN λ expression accompanied by downregulation of miRNAs, and are important for RSV G protein function and targeting

Isothermal amplification and fluorescent detection of SARS-CoV-2 and SARS-CoV-2 variant virus in nasopharyngeal swabs

The COVID-19 pandemic caused by the SARS-CoV-2 is a serious health threat causing worldwide morbidity and mortality. Real-time reverse transcription PCR (RT-qPCR) is currently the standard for SARS-CoV-2 detection. Although various nucleic acid-based assays have been developed to aid the detection of SARS-CoV-2 from COVID-19 patient samples, the objective of this study was to develop a diagnostic test that can be completed in 30 minutes without having to isolate RNA from the samples. Here, we present an RNA amplification detection method performed using reverse transcription loop-mediated isothermal amplification (RT-LAMP) reactions to achieve specific, rapid (30 min), and sensitive (<100 copies) fluorescent detection in real-time of SARS-CoV-2 directly from patient nasopharyngeal swab (NP) samples. When compared to RT-qPCR, positive NP swab samples assayed by fluorescent RT-LAMP had 98% (n = 41/42) concordance and negative NP swab samples assayed by fluorescent RT-LAMP had 87% (n = 59/68) concordance for the same samples. Importantly, the fluorescent RT-LAMP results were obtained without purification of RNA from the NP swab samples in contrast to RT-qPCR. We also show that the fluorescent RT-LAMP assay can specifically detect live virus directly from cultures of both SARS-CoV-2 wild type (WA1/2020), and a SARS-CoV-2 B.1.1.7 (alpha) variant strain with equal sensitivity to RT-qPCR. RT-LAMP has several advantages over RT-qPCR including isothermal amplification, speed (<30 min), reduced costs, and similar sensitivity and specificity

Small Non-coding RNA Expression Following Respiratory Syncytial Virus or Measles Virus Infection of Neuronal Cells

Publication history: Accepted - 3 August 2021; Published - 3 September 2021.Respiratory syncytial virus (RSV) or measles virus (MeV) infection modifies host responses through small non-coding RNA (sncRNA) expression. We show that RSV or MeV infection of neuronal cells induces sncRNAs including various microRNAs and transfer RNA fragments (tRFs). We show that these tRFs originate from select tRNAs (GCC and CAC for glycine, CTT and AAC for Valine, and CCC and TTT for Lysine). Some of the tRNAs are rarely used by RSV or MeV as indicated by relative synonymous codon usage indices suggesting selective cleavage of the tRNAs occurs in infected neuronal cells. The data implies that differentially expressed sncRNAs may regulate host gene expression via multiple mechanisms in neuronal cells.The studies were supported by a grant awarded to SC and RT (IE140652) to study “Paramyxovirus miRNA biogenesis and implications for host and viral gene expression” and the Georgia Research Alliance Foundation for RT

MicroRNA Regulation of Human Protease Genes Essential for Influenza Virus Replication

Influenza A virus causes seasonal epidemics and periodic pandemics threatening the health of millions of people each year. Vaccination is an effective strategy for reducing morbidity and mortality, and in the absence of drug resistance, the efficacy of chemoprophylaxis is comparable to that of vaccines. However, the rapid emergence of drug resistance has emphasized the need for new drug targets. Knowledge of the host cell components required for influenza replication has been an area targeted for disease intervention. In this study, the human protease genes required for influenza virus replication were determined and validated using RNA interference approaches. The genes validated as critical for influenza virus replication were ADAMTS7, CPE, DPP3, MST1, and PRSS12, and pathway analysis showed these genes were in global host cell pathways governing inflammation (NF-κB), cAMP/calcium signaling (CRE/CREB), and apoptosis. Analyses of host microRNAs predicted to govern expression of these genes showed that eight miRNAs regulated gene expression during virus replication. These findings identify unique host genes and microRNAs important for influenza replication providing potential new targets for disease intervention strategies

The Central Conserved Region (CCR) of Respiratory Syncytial Virus (RSV) G Protein Modulates Host miRNA Expression and Alters the Cellular Response to Infection

Respiratory Syncytial Virus (RSV) infects respiratory epithelial cells and deregulates host gene expression by many mechanisms including expression of RSV G protein (RSV G). RSV G protein encodes a central conserved region (CCR) containing a CX3C motif that functions as a fractalkine mimic. Disruption of the CX3C motif (a.a. 182–186) located in the CCR of the G protein has been shown to affect G protein function in vitro and the severity of RSV disease pathogenesis in vivo. We show that infection of polarized Calu3 respiratory cells with recombinant RSV having point mutations in Cys173 and 176 (C173/176S) (rA2-GC12), or Cys186 (C186S) (rA2-GC4) is associated with a decline in the integrity of polarized Calu-3 cultures and decreased virus production. This is accompanied with downregulation of miRNAs let-7f and miR-24 and upregulation of interferon lambda (IFNλ), a primary antiviral cytokine for RSV in rA2-GC12/rA2-GC4 infected cells. These results suggest that residues in the cysteine noose region of RSV G protein can modulate IFN λ expression accompanied by downregulation of miRNAs, and are important for RSV G protein function and targeting