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Open AccessEditorial

Special Issue “Next-Generation Technologies to Understand Mechanisms of Virus Infections”

by Allan R. Brasier

Viruses 2023, 15(1), 33; https://doi.org/10.3390/v15010033 - 22 Dec 2022

Cited by 1 | Viewed by 1173

Abstract

RNA viruses are responsible for substantial morbidity and health burden [...] Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

Research

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18 pages, 5097 KiB

Open AccessArticle

Circular RNA Profiles in Viremia and ART Suppression Predict Competing circRNA–miRNA–mRNA Networks Exclusive to HIV-1 Viremic Patients

by Dora Zucko, Abdullgadir Hayir, Kelsey Grinde and Kathleen Boris-Lawrie

Viruses 2022, 14(4), 683; https://doi.org/10.3390/v14040683 - 25 Mar 2022

Cited by 3 | Viewed by 2921

Abstract

Since the onset of the HIV-1/AIDS epidemic in 1981, 75 million people have been infected with the virus, and the disease remains a public health crisis worldwide. Circular RNAs (circRNAs) are derived from excised exons and introns during backsplicing, a form of alternative [...] Read more.

Since the onset of the HIV-1/AIDS epidemic in 1981, 75 million people have been infected with the virus, and the disease remains a public health crisis worldwide. Circular RNAs (circRNAs) are derived from excised exons and introns during backsplicing, a form of alternative splicing. The relevance of unconventional, non-capped, and non-poly(A) transcripts to transcriptomics studies remains to be routinely investigated. Knowledge gaps to be filled are the interface between host-encoded circRNAs and viral replication in chronically progressed patients and upon treatment with antiviral drugs. We implemented a bioinformatic pipeline and repurpose publicly archived RNA sequence reads from the blood of 19 HIV-1-positive patients that previously compared transcriptomes during viremia and viremia suppression by antiretroviral therapy (ART). The in silico analysis identified viremic patients’ circRNA that became undetectable after ART. The circRNAs originated from a subset of host genes enriched in the HDAC biological pathway. These circRNAs and parental mRNAs held in common a small collection of miRNA response elements (MREs), some of which were present in HIV-1 mRNAs. The function of the MRE-containing target mRNA enriched the RNA polymerase II GO pathway. To visualize the interplay between individual circRNA–miRNA–target mRNA, important for HIV-1 and potentially other diseases, an Interactive Circos tool was developed to efficiently parse the intricately competing endogenous network of circRNA–miRNA–mRNA interactions originating from seven circRNA singled out in viremic versus non-viremic patients. The combined downregulation of the identified circRNAs warrants investigation as a novel antiviral targeting strategy. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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13 pages, 2001 KiB

Open AccessArticle

Metabolic Reprogramming of Nasal Airway Epithelial Cells Following Infant Respiratory Syncytial Virus Infection

by Andrew R. Connelly, Brian M. Jeong, Mackenzie E. Coden, Jacob Y. Cao, Tatiana Chirkova, Christian Rosas-Salazar, Jacqueline-Yvonne Cephus, Larry J. Anderson, Dawn C. Newcomb, Tina V. Hartert and Sergejs Berdnikovs

Viruses 2021, 13(10), 2055; https://doi.org/10.3390/v13102055 - 13 Oct 2021

Cited by 11 | Viewed by 3797

Abstract

Respiratory syncytial virus (RSV) is a seasonal mucosal pathogen that infects the ciliated respiratory epithelium and results in the most severe morbidity in the first six months of life. RSV is a common cause of acute respiratory infection during infancy and is an [...] Read more.

Respiratory syncytial virus (RSV) is a seasonal mucosal pathogen that infects the ciliated respiratory epithelium and results in the most severe morbidity in the first six months of life. RSV is a common cause of acute respiratory infection during infancy and is an important early-life risk factor strongly associated with asthma development. While this association has been repeatedly demonstrated, limited progress has been made on the mechanistic understanding in humans of the contribution of infant RSV infection to airway epithelial dysfunction. An active infection of epithelial cells with RSV in vitro results in heightened central metabolism and overall hypermetabolic state; however, little is known about whether natural infection with RSV in vivo results in lasting metabolic reprogramming of the airway epithelium in infancy. To address this gap, we performed functional metabolomics, 13C glucose metabolic flux analysis, and RNA-seq gene expression analysis of nasal airway epithelial cells (NAECs) sampled from infants between 2–3 years of age, with RSV infection or not during the first year of life. We found that RSV infection in infancy was associated with lasting epithelial metabolic reprogramming, which was characterized by (1) significant increase in glucose uptake and differential utilization of glucose by epithelium; (2) altered preferences for metabolism of several carbon and energy sources; and (3) significant sexual dimorphism in metabolic parameters, with RSV-induced metabolic changes most pronounced in male epithelium. In summary, our study supports the proposed phenomenon of metabolic reprogramming of epithelial cells associated with RSV infection in infancy and opens exciting new venues for pursuing mechanisms of RSV-induced epithelial barrier dysfunction in early life. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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16 pages, 2487 KiB

Open AccessArticle

Impact of Analytical Treatment Interruption on Burden and Diversification of HIV Peripheral Reservoir: A Pilot Study

by Rossana Scutari, Valentino Costabile, Laura Galli, Maria Concetta Bellocchi, Luca Carioti, Silvia Barbaliscia, Andrea Poli, Andrea Galli, Carlo Federico Perno, Maria Mercedes Santoro, Antonella Castagna, Francesca Ceccherini-Silberstein, Claudia Alteri and Vincenzo Spagnuolo

Viruses 2021, 13(7), 1403; https://doi.org/10.3390/v13071403 - 19 Jul 2021

Cited by 3 | Viewed by 2232

Abstract

Background: If analytical antiretroviral-treatment (ART) interruption (ATI) might significantly impact quantitative or qualitative peripheral-total HIV-DNA is still debated. Methods: Six chronically HIV-1 infected patients enrolled in APACHE-study were analysed for peripheral-total HIV-DNA and residual viremia, major-resistance-mutations (MRMs) and C2-V3-C3 evolution at pre-ATI (T1), [...] Read more.

Background: If analytical antiretroviral-treatment (ART) interruption (ATI) might significantly impact quantitative or qualitative peripheral-total HIV-DNA is still debated. Methods: Six chronically HIV-1 infected patients enrolled in APACHE-study were analysed for peripheral-total HIV-DNA and residual viremia, major-resistance-mutations (MRMs) and C2-V3-C3 evolution at pre-ATI (T1), during ATI (T2) and at achievement of virological success after ART-resumption (post-ATI, T3). These data were obtained at three comparable time-points in five chronically HIV-1 infected patients on suppressive ART for ≥1 year, enrolled in MODAt-study. Results: At T1, APACHE and MODAt individuals had similar peripheral-total HIV-DNA and residual viremia (p = 0.792 and 0.662, respectively), and no significant changes for these parameters were observed between T1 and T3 in both groups. At T1, 4/6 APACHE and 2/5 MODAt carried HIV-DNA MRMs. MRMs disappeared at T3 in 3/4 APACHE. All disappearing MRMs were characterized by T1 intra-patient prevalence <80%, and mainly occurred in APOBEC3-related sites. All MRMs persisted over-time in the 2 MODAt. C2-V3-C3 genetic-distance significantly changed from T1 to T3 in APACHE individuals (+0.36[0.11–0.41], p = 0.04), while no significant changes were found in MODAt. Accordingly, maximum likelihood trees (bootstrap > 70%) and genealogical sorting indices (GSI > 0.50 with p-value < 0.05) showed that T1 C2-V3-C3 DNA sequences were distinct from T2 and T3 viruses in 4/6 APACHE. Virus populations at all three time-points were highly interspersed in MODAt. Conclusions: This pilot study indicates that short ATI does not alter peripheral-total HIV-DNA burden and residual viremia, but in some cases could cause a genetic diversification of peripheral viral reservoir in term of both MRMs rearrangement and viral evolution. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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16 pages, 10891 KiB

Open AccessArticle

A Fluorescent Real-Time Plaque Assay Enables Single-Cell Analysis of Virus-Induced Cytopathic Effect by Live-Cell Imaging

by Jorge L. Arias-Arias, Eugenia Corrales-Aguilar and Rodrigo A. Mora-Rodríguez

Viruses 2021, 13(7), 1193; https://doi.org/10.3390/v13071193 - 22 Jun 2021

Cited by 5 | Viewed by 4497

Abstract

Conventional plaque assays rely on the use of overlays to restrict viral infection allowing the formation of distinct foci that grow in time as the replication cycle continues leading to countable plaques that are visualized with standard techniques such as crystal violet, neutral [...] Read more.

Conventional plaque assays rely on the use of overlays to restrict viral infection allowing the formation of distinct foci that grow in time as the replication cycle continues leading to countable plaques that are visualized with standard techniques such as crystal violet, neutral red, or immunolabeling. This classical approach takes several days until large enough plaques can be visualized and counted with some variation due to subjectivity in plaque recognition. Since plaques are clonal lesions produced by virus-induced cytopathic effect, we applied DNA fluorescent dyes with differential cell permeability to visualize them by live-cell imaging. We could observe different stages of that cytopathic effect corresponding to an early wave of cells with chromatin-condensation followed by a wave of dead cells with membrane permeabilization within plaques generated by different animal viruses. This approach enables an automated plaque identification using image analysis to increase single plaque resolution compared to crystal violet counterstaining and allows its application to plaque tracking and plaque reduction assays to test compounds for both antiviral and cytotoxic activities. This fluorescent real-time plaque assay sums to those next-generation technologies by combining this robust classical method with modern fluorescence microscopy and image analysis approaches for future applications in virology. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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15 pages, 2105 KiB

Open AccessArticle

Late Relapse and Reinfection in HCV Patients Treated with Direct-Acting Antiviral (DAA) Drugs

by Claudia Minosse, Cesare E. M. Gruber, Martina Rueca, Chiara Taibi, Mauro Zaccarelli, Elisabetta Grilli, Marzia Montalbano, Maria R. Capobianchi, Andrea Antinori, Gianpiero D’Offizi, Fiona McPhee and Anna Rosa Garbuglia

Viruses 2021, 13(6), 1151; https://doi.org/10.3390/v13061151 - 16 Jun 2021

Cited by 6 | Viewed by 2543

Abstract

The risk of hepatitis C virus (HCV) recurrence after direct-acting antiviral (DAA) treatment is <0.5%. However, the distinction between HCV RNA late relapse and reinfection still represents a challenge in virological diagnostics. The aim of this study was to employ next-generation sequencing (NGS) [...] Read more.

The risk of hepatitis C virus (HCV) recurrence after direct-acting antiviral (DAA) treatment is <0.5%. However, the distinction between HCV RNA late relapse and reinfection still represents a challenge in virological diagnostics. The aim of this study was to employ next-generation sequencing (NGS) to investigate HCV RNA recurrence in patients achieving a sustained virologic response (SVR) at least six months post-treatment. NGS was performed on plasma samples from six HCV-positive patients (Pt1–6) treated with DAA. NGS of HCV NS5B was analyzed before treatment (T0), after HCV RNA rebound (T1), and, for Pt3, after a second rebound (T2). Reinfection was confirmed for Pt5, and for the first rebound observed in Pt3. Conversely, viral relapse was observed when comparing T0 and T1 for Pt6 and T1 and T2 for Pt3. Z-scores were calculated and used to predict whether HCV-positive patient samples at different time points belonged to the same quasispecies population. A low Z-score of <2.58 confirmed that viral quasispecies detected at T0 and T1 were closely related for both Pt1 and Pt2, while the Z-score for Pt4 was suggestive of possible reinfection. NGS data analyses indicate that the Z-score may be a useful parameter for distinguishing late relapse from reinfection. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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19 pages, 2947 KiB

Open AccessArticle

Discovery of RSV-Induced BRD4 Protein Interactions Using Native Immunoprecipitation and Parallel Accumulation—Serial Fragmentation (PASEF) Mass Spectrometry

by Morgan Mann, David S. Roberts, Yanlong Zhu, Yi Li, Jia Zhou, Ying Ge and Allan R. Brasier

Viruses 2021, 13(3), 454; https://doi.org/10.3390/v13030454 - 11 Mar 2021

Cited by 17 | Viewed by 3376

Abstract

Respiratory Syncytial Virus (RSV) causes severe inflammation and airway pathology in children and the elderly by infecting the epithelial cells of the upper and lower respiratory tract. RSV replication is sensed by intracellular pattern recognition receptors upstream of the IRF and NF-κB transcription [...] Read more.

Respiratory Syncytial Virus (RSV) causes severe inflammation and airway pathology in children and the elderly by infecting the epithelial cells of the upper and lower respiratory tract. RSV replication is sensed by intracellular pattern recognition receptors upstream of the IRF and NF-κB transcription factors. These proteins coordinate an innate inflammatory response via Bromodomain-containing protein 4 (BRD4), a protein that functions as a scaffold for unknown transcriptional regulators. To better understand the pleiotropic regulatory function of BRD4, we examine the BRD4 interactome and identify how RSV infection dynamically alters it. To accomplish these goals, we leverage native immunoprecipitation and Parallel Accumulation—Serial Fragmentation (PASEF) mass spectrometry to examine BRD4 complexes isolated from human alveolar epithelial cells in the absence or presence of RSV infection. In addition, we explore the role of BRD4’s acetyl-lysine binding bromodomains in mediating these interactions by using a highly selective competitive bromodomain inhibitor. We identify 101 proteins that are significantly enriched in the BRD4 complex and are responsive to both RSV-infection and BRD4 inhibition. These proteins are highly enriched in transcription factors and transcriptional coactivators. Among them, we identify members of the AP1 transcription factor complex, a complex important in innate signaling and cell stress responses. We independently confirm the BRD4/AP1 interaction in primary human small airway epithelial cells. We conclude that BRD4 recruits multiple transcription factors during RSV infection in a manner dependent on acetyl-lysine binding domain interactions. This data suggests that BRD4 recruits transcription factors to target its RNA processing complex to regulate gene expression in innate immunity and inflammation. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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16 pages, 2464 KiB

Open AccessArticle

SARS-CoV-2 Variant of Concern 202012/01 Has about Twofold Replicative Advantage and Acquires Concerning Mutations

by Frederic Grabowski, Grzegorz Preibisch, Stanisław Giziński, Marek Kochańczyk and Tomasz Lipniacki

Viruses 2021, 13(3), 392; https://doi.org/10.3390/v13030392 - 1 Mar 2021

Cited by 73 | Viewed by 7687

Abstract

The novel SARS-CoV-2 Variant of Concern (VOC)-202012/01 (also known as B.1.1.7), first collected in United Kingdom on 20 September 2020, is a rapidly growing lineage that in January 2021 constituted 86% of all SARS-CoV-2 genomes sequenced in England. The VOC has been detected [...] Read more.

The novel SARS-CoV-2 Variant of Concern (VOC)-202012/01 (also known as B.1.1.7), first collected in United Kingdom on 20 September 2020, is a rapidly growing lineage that in January 2021 constituted 86% of all SARS-CoV-2 genomes sequenced in England. The VOC has been detected in 40 out of 46 countries that reported at least 50 genomes in January 2021. We have estimated that the replicative advantage of the VOC is in the range 1.83–2.18 [95% CI: 1.71–2.40] with respect to the 20A.EU1 variant that dominated in England in November 2020, and in range 1.65–1.72 [95% CI: 1.46–2.04] in Wales, Scotland, Denmark, and USA. As the VOC strain will likely spread globally towards fixation, it is important to monitor its molecular evolution. We have estimated growth rates of expanding mutations acquired by the VOC lineage to find that the L18F substitution in spike has initiated a fast growing VOC substrain. The L18F substitution is of significance because it has been found to compromise binding of neutralizing antibodies. Of concern are immune escape mutations acquired by the VOC: E484K, F490S, S494P (in the receptor binding motif of spike) and Q677H, Q675H (in the proximity of the polybasic cleavage site at the S1/S2 boundary). These mutants may hinder efficiency of existing vaccines and expand in response to the increasing after-infection or vaccine-induced seroprevalence. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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22 pages, 10791 KiB

Open AccessArticle

Alternative mRNA Processing of Innate Response Pathways in Respiratory Syncytial Virus (RSV) Infection

by Xiaofang Xu, Morgan Mann, Dianhua Qiao and Allan R. Brasier

Viruses 2021, 13(2), 218; https://doi.org/10.3390/v13020218 - 31 Jan 2021

Cited by 12 | Viewed by 2594

Abstract

The innate immune response (IIR) involves rapid genomic expression of protective interferons (IFNs) and inflammatory cytokines triggered by intracellular viral replication. Although the transcriptional control of the innate pathway is known in substantial detail, little is understood about the complexity of alternative splicing [...] Read more.

The innate immune response (IIR) involves rapid genomic expression of protective interferons (IFNs) and inflammatory cytokines triggered by intracellular viral replication. Although the transcriptional control of the innate pathway is known in substantial detail, little is understood about the complexity of alternative splicing (AS) and alternative polyadenylation (APA) of mRNAs underlying the cellular IIR. In this study, we applied single-molecule, real-time (SMRT) sequencing with mRNA quantitation using short-read mRNA sequencing to characterize changes in mRNA processing in the epithelial response to respiratory syncytial virus (RSV) replication. Mock or RSV-infected human small-airway epithelial cells (hSAECs) were profiled using SMRT sequencing and the curated transcriptome analyzed by structural and quality annotation of novel transcript isoforms (SQANTI). We identified 113,082 unique isoforms; 28,561 represented full splice matches, and 45% of genes expressed six or greater AS mRNA isoforms. Identification of differentially expressed AS isoforms was accomplished by mapping a short-read RNA sequencing expression matrix to the curated transcriptome, and 905 transcripts underwent differential polyadenylation site analysis enriched in protein secretion, translation, and mRNA degradation. We focused on 355 genes showing differential isoform utilization (DIU), indicating where a new AS isoform becomes a major fraction of mRNA isoforms expressed. In pathway and network enrichment analyses, we observed that DIU transcripts are substantially enriched in cell cycle control and IIR pathways. Interestingly, the RelA/IRF7 innate regulators showed substantial DIU where major transcripts included distinct isoforms with exon occlusion, intron inclusion, and alternative transcription start site utilization. We validated the presence of RelA and IRF7 AS isoforms as well as their induction by RSV using eight isoform-specific RT-PCR assays. These isoforms were identified in both immortalized and primary small-airway epithelial cells. We concluded that the cell cycle and IIR are differentially spliced in response to RSV. These data indicate that substantial post-transcriptional complexity regulates the antiviral response. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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14 pages, 888 KiB

Open AccessArticle

NanoString Technology for Human Papillomavirus Typing

by Mangalathu S. Rajeevan, Sonya Patel, Tengguo Li and Elizabeth R. Unger

Viruses 2021, 13(2), 188; https://doi.org/10.3390/v13020188 - 27 Jan 2021

Cited by 2 | Viewed by 2307

Abstract

High-throughput HPV typing assays with increased automation, faster turnaround and type-specific digital readout would facilitate studies monitoring the impact of HPV vaccination. We evaluated the NanoString nCounter^® platform for detection and digital readout of 48 HPV types in a single reaction. NanoString [...] Read more.

High-throughput HPV typing assays with increased automation, faster turnaround and type-specific digital readout would facilitate studies monitoring the impact of HPV vaccination. We evaluated the NanoString nCounter^® platform for detection and digital readout of 48 HPV types in a single reaction. NanoString (NS) used proprietary software to design CodeSets: type-specific probe pairs targeting 48 HPV types and the globin gene. We tested residual DNA extracts from epidemiologic specimens and defined samples (HPV plasmids at 10 to 10⁴ copies/reaction) directly (No-PCR) as well as after L1 consensus PCR of 45 (PCR-45) or 15 cycles (PCR-15). Assay and interpretation followed NS recommendations. We evaluated analytic performance by comparing NanoString results for types included in prior assays: Roche Linear Array (LA) or HPV TypeSeq assay. No-PCR results on 40 samples showed good type-specific agreement with LA (k = 0.621) but sensitivity was 65% with lower limit of detection (LOD) at 10⁴ plasmid copies. PCR-45 results showed almost perfect type-specific agreement with LA (k = 0.862), 82% sensitivity and LOD at 10 copies. PCR-15 results on 75 samples showed substantial type-specific agreement with LA (k = 0.796, 92% sensitivity) and TypeSeq (k = 0.777, 87% sensitivity), and LOD at 10 copies of plasmids. This proof-of-principle study demonstrates the efficacy of the NS platform with HPV CodeSet for type-specific detection using a low number of PCR cycles (PCR-15). Studies are in progress to evaluate assay reproducibility and analytic validation with a larger number of samples. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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9 pages, 2860 KiB

Open AccessArticle

Library Preparation Based on Transposase Assisted RNA/DNA Hybrid Co-Tagmentation for Next-Generation Sequencing of Human Noroviruses

by Zilei Zhang, Danlei Liu, Dapeng Wang and Qingping Wu

Viruses 2021, 13(1), 65; https://doi.org/10.3390/v13010065 - 6 Jan 2021

Cited by 3 | Viewed by 3313

Abstract

Human noroviruses (HuNoVs) are one of the leading causes of foodborne illnesses globally. The viral genome is the most essential information for viral source tracing and viral transmission pattern monitoring. However, whole genome sequencing of HuNoVs is still challenging due to the sequence [...] Read more.

Human noroviruses (HuNoVs) are one of the leading causes of foodborne illnesses globally. The viral genome is the most essential information for viral source tracing and viral transmission pattern monitoring. However, whole genome sequencing of HuNoVs is still challenging due to the sequence heterogeneity among different genotypes and low titer in samples. To address this need, in this study, the Transposase assisted RNA/DNA hybrid Co-tagmentation (TRACE-seq) method was established for next generation sequencing library preparation of HuNoVs. Our data demonstrated that almost the whole HuNoVs genome (>7 kb) could be obtained from all of the 11 clinical samples tested. Twelve genotypes including GI.3, GI.4, GI.5, GI.8, GII.2, GII.3, GII.4, GII.6, GII.12, GII.13, GII.14, and GII.21 were involved. Compared with the traditional method for viral metagenomics library preparation, optimized TRACE-seq greatly reduced the interference from the host’s and bacterial RNAs. In addition, viral genome sequences can be assembled by using less raw data with sufficient depth along the whole genome. Therefore, for the high versatility and reliability, this method is promising for whole viral genome attainment. It is particularly applicable for the viruses with a low titer that are mixed with a complicated host background and are unable to be cultured in vitro, like the HuNoVs utilized in this study. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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Review

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23 pages, 1161 KiB

Open AccessReview

Top-Down and Bottom-Up Proteomics Methods to Study RNA Virus Biology

by Yogy Simanjuntak, Kira Schamoni-Kast, Alice Grün, Charlotte Uetrecht and Pietro Scaturro

Viruses 2021, 13(4), 668; https://doi.org/10.3390/v13040668 - 13 Apr 2021

Cited by 4 | Viewed by 3881

Abstract

RNA viruses cause a wide range of human diseases that are associated with high mortality and morbidity. In the past decades, the rise of genetic-based screening methods and high-throughput sequencing approaches allowed the uncovering of unique and elusive aspects of RNA virus replication [...] Read more.

RNA viruses cause a wide range of human diseases that are associated with high mortality and morbidity. In the past decades, the rise of genetic-based screening methods and high-throughput sequencing approaches allowed the uncovering of unique and elusive aspects of RNA virus replication and pathogenesis at an unprecedented scale. However, viruses often hijack critical host functions or trigger pathological dysfunctions, perturbing cellular proteostasis, macromolecular complex organization or stoichiometry, and post-translational modifications. Such effects require the monitoring of proteins and proteoforms both on a global scale and at the structural level. Mass spectrometry (MS) has recently emerged as an important component of the RNA virus biology toolbox, with its potential to shed light on critical aspects of virus–host perturbations and streamline the identification of antiviral targets. Moreover, multiple novel MS tools are available to study the structure of large protein complexes, providing detailed information on the exact stoichiometry of cellular and viral protein complexes and critical mechanistic insights into their functions. Here, we review top-down and bottom-up mass spectrometry-based approaches in RNA virus biology with a special focus on the most recent developments in characterizing host responses, and their translational implications to identify novel tractable antiviral targets. Full article

(This article belongs to the Special Issue Next-Generation Technologies to Understand Mechanisms of Virus Infections)

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