Respiratory Syncytial Virus Entry Research Articles

Both chebulagic acid and punicalagin inhibit respiratory syncytial virus entry via multi-targeting glycoprotein and fusion protein.

Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infections, with no currently available small-molecule drugs that are both safe and effective. A major obstacle in antiviral drug development is the rapid emergence of drug-resistant viral strains. Targeting multiple viral compounds may help mitigate the development of resistance. Herein, we conducted a drug screening using the Antiviral Traditional Chinese Medicine Active Compound Library, aiming to identify compounds that simultaneously target the RSV fusion (F) protein, glycoprotein (G), and the host heparan sulfate proteoglycans (HSPGs). From this screening, 10 candidate compounds were identified for their ability to interact with all three targets. Among these 10 candidates, chebulagic acid (CHLA) and punicalagin (PUG) demonstrated the most potent inhibition of RSV replication. In vitro dose-response assays confirmed the antiviral efficacy of CHLA (IC50: 0.07864 µM) and PUG (IC50: 0.08065 µM). Further experiments revealed both CHLA and PUG disrupt RSV attachment and membrane fusion by targeting the RSV-F and G proteins, rather than HSPG. Notably, CHLA and PUG were found to bind to the CX3C motif of the RSV-G protein, with docking assays predicting their binding sites at cysteines 176 and 182. Additionally, CHLA enhanced the conformational stability of the RSV-F protein before fusion. In an in vivo study, both CHLA and PUG were shown to alleviate RSV-induced pulmonary pathology by reducing viral titers, mitigating lung injury, and suppressing the inflammatory responses in the lungs. Our findings suggest that CHLA and PUG hold potential as therapeutic agents for RSV infection.IMPORTANCEA significant challenge in developing anti-respiratory syncytial virus (RSV) agents is the rapid emergence of resistant viral strains. Designing drugs that target multiple viral components can effectively reduce the likelihood of developing resistant strains. In this study, we screened compounds from the Antiviral Traditional Chinese Medicine Active Compound Library, aiming to simultaneously targe the RSV fusion (F) protein, glycoprotein (G), and host heparan sulfate proteoglycans (HSPGs). Our findings revealed that chebulagic acid (CHLA) and punicalagin (PUG) significantly inhibited RSV replication both in vitro and in vivo and interacted with all three targets. Both CHLA and PUG were able to disrupt RSV attachment and membrane fusion. Mechanistically, CHLA and PUG were found to bind to the CX3C motif of the RSV-G protein, with CHLA also enhancing the conformational stability of the RSV-F protein before fusion. In conclusion, our study suggests that CHLA and PUG hold promise as therapeutic agents against RSV infection.

A new mechanism of respiratory syncytial virus entry inhibition by small-molecule to overcome K394R-associated resistance.

Respiratory syncytial virus (RSV) infection is a major public health concern, and many small-molecule candidates targeting the viral fusion (F) protein are associated with a considerable risk of inducing drug-resistant mutations. This study investigated virological features of the K394R variant, a mutant strain conferring resistance to multiple RSV fusion inhibitors. Our results demonstrated that the K394R variant is highly fusogenic in cell cultures and more pathogenic than the parental strain in mice. The small-molecule inhibitor CL-A3-7 substantially reduced in vitro and in vivo infections of both wild-type RSV and the K394R variant by blocking the interaction of viral F protein with its cellular receptor, showing a new mechanism of action for small-molecules to inhibit RSV infection and overcome K394R-associated resistance.

Farnesyltransferase inhibitor lonafarnib suppresses respiratory syncytial virus infection by blocking conformational change of fusion glycoprotein

Respiratory syncytial virus (RSV) is the major cause of bronchiolitis and pneumonia in young children and the elderly. There are currently no approved RSV-specific therapeutic small molecules available. Using high-throughput antiviral screening, we identified an oral drug, the prenylation inhibitor lonafarnib, which showed potent inhibition of the RSV fusion process. Lonafarnib exhibited antiviral activity against both the RSV A and B genotypes and showed low cytotoxicity in HEp-2 and human primary bronchial epithelial cells (HBEC). Time-of-addition and pseudovirus assays demonstrated that lonafarnib inhibits RSV entry, but has farnesyltransferase-independent antiviral efficacy. Cryo-electron microscopy revealed that lonafarnib binds to a triple-symmetric pocket within the central cavity of the RSV F metastable pre-fusion conformation. Mutants at the RSV F sites interacting with lonafarnib showed resistance to lonafarnib but remained fully sensitive to the neutralizing monoclonal antibody palivizumab. Furthermore, lonafarnib dose-dependently reduced the replication of RSV in BALB/c mice. Collectively, lonafarnib could be a potential fusion inhibitor for RSV infection.

Emodin inhibits respiratory syncytial virus entry by interactions with fusion protein.

Respiratory syncytial virus (RSV) fusion (F) protein is essential for facilitating virus entry into host cells, providing a hopeful path for combating viral diseases. However, F protein inhibitors can rapidly select for viral resistance. Thus, discovering new inhibitors of F-protein is necessary to enrich the RSV drug development pipeline. In this study, we screen 25 bioactive compounds from Chinese herbal medicines that exhibit a strong binding to the RSV-F protein using surface plasmon resonance. After screening, we found emodin could strongly bind to RSV-F protein, and could effectively curb RSV infection. Further investigations certificated that emodin specifically disrupts the attachment and internalization phases of RSV infection by targeting the RSV-F protein. In vivo studies with mice infected with RSV demonstrated that emodin effectively reduces lung pathology. This therapeutic effect is attributed to emodin's capacity to diminish pro-inflammatory cytokine production and reduce viral load in the lungs. In conclusion, our findings provide initial insights into the mechanism by which emodin counters RSV infection via engagement with the RSV-F protein, establishing it as a viable contender for the development of novel therapeutic agents aimed at RSV.

M6A Reader YTHDC1 Impairs Respiratory Syncytial Virus Infection by Downregulating Membrane CX3CR1 Expression.

Respiratory syncytial virus (RSV) is the most prevalent cause of acute lower respiratory infection in young children. Currently, the first RSV vaccines are approved by the FDA. Recently, N6-methyladenosine (m6A) RNA methylation has been implicated in the regulation of the viral life cycle and replication of many viruses, including RSV. m6A methylation of RSV RNA has been demonstrated to promote replication and prevent anti-viral immune responses by the host. Whether m6A is also involved in viral entry and whether m6A can also affect RSV infection via different mechanisms than methylation of viral RNA is poorly understood. Here, we identify m6A reader YTH domain-containing protein 1 (YTHDC1) as a novel negative regulator of RSV infection. We demonstrate that YTHDC1 abrogates RSV infection by reducing the expression of RSV entry receptor CX3C motif chemokine receptor 1 (CX3CR1) on the cell surface of lung epithelial cells. Altogether, these data reveal a novel role for m6A methylation and YTHDC1 in the viral entry of RSV. These findings may contribute to the development of novel treatment options to control RSV infection.

Exploring the secrets of virus entry: the first respiratory syncytial virus carrying beta lactamase.

Respiratory Syncytial Virus (RSV) presents a significant health threat, especially to young children. In-depth understanding of RSV entry mechanisms is essential for effective antiviral development. This study introduces an innovative RSV variant, featuring the fusion of the beta-lactamase (BlaM) enzyme with the RSV-P phosphoprotein, providing a versatile tool for dissecting viral entry dynamics. Using the AlphaFold2 algorithm, we modeled the tertiary structure of the P-BlaM chimera, revealing structural similarities with both RSV-P and BlaM. Functional assessments, utilizing flow cytometry, quantified beta-lactamase activity and GFP expression in infected bronchial epithelial cells. Western blot analysis confirmed the integrity of P-BlaM within virions. The modeled P-BlaM chimera exhibited structural parallels with RSV-P and BlaM. Functional assays demonstrated robust beta-lactamase activity in recombinant virions, confirming successful P-BlaM incorporation as a structural protein. Quercetin, known for its antiviral properties, impeded viral entry by affecting virion fusion. Additionally, Ulixertinib, an ERK-1/2 inhibitor, significantly curtailed viral entry, implicating ERK-1/2 pathway signaling. Our engineered RSV-P-BlaM chimera emerges as a valuable tool, illuminating RSV entry mechanisms. Structural and functional analyses unveil potential therapeutic targets. Quercetin and Ulixertinib, identified as distinct stage inhibitors, show promise for targeted antiviral strategies. Time-of-addition assays pinpoint quercetin's specific interference stage, advancing our comprehension of RSV entry and guiding future antiviral developments.

Advances in understanding RSV glycoproteins: Insights into viral entry and implications for vaccines and therapeutic development

Respiratory syncytial virus (RSV) causes a significant number of deaths in newborns annually due to respiratory infections. Since its first isolation in 1955, great progress has been made in the development of related vaccines and monoclonal antibodies. Currently, two RSV vaccines and two monoclonal antibodies have been approved for marketing. Understanding the structure, function, and mechanism of RSV glycoproteins interaction with their receptors involved in its entry is of great significance for the development of vaccines and therapeutic drugs. In this review, we will introduce the substantial progress made in the understanding of the structure of RSV glycoproteins, the receptors they bind with, and the process of RSV entry. Progress made in developing vaccines and neutralizing antibodies which benefit from these studies will also be briefly discussed. Finally, we will outline the unresolved issues and controversies in RSV glycoprotein and entry mechanism research, propose some possible approaches to address these issues, and provide insights into future drug and vaccine development.

Actin cytoskeleton remodeling disrupts physical barriers to infection and presents entry receptors to respiratory syncytial virus.

RSV is the leading cause of infant hospitalizations and a significant cause of paediatric and geriatric morbidity worldwide. Recently, we reported that insulin-like growth factor 1 receptor (IGF1R) was a receptor for respiratory syncytial virus (RSV) in airway epithelial cells and that activation of IGF1R recruited the coreceptor, nucleolin (NCL), to the cell surface. Cilia and mucus that line the airways pose a significant barrier to viral and bacterial infection. The cortical actin cytoskeleton has been shown by others to mediate RSV entry, so we studied the roles of the RSV receptors and actin remodelling during virus entry. We found that IGF1R expression and phosphorylation were associated with the ability of RSV to infect cells. Confocal immunofluorescence imaging showed that actin projections, a hallmark of macropinocytosis, formed around viral particles 30 min after infection. Consistent with prior reports we also found that virus particles were internalized into early endosome antigen-1 positive endosomes within 90 min. Inhibiting actin polymerization significantly reduced viral titre by approximately ten-fold. Inhibiting PI3 kinase and PKCζ in stratified air-liquid interface (ALI) models of the airway epithelium had similar effects on reducing the actin remodelling observed during infection and attenuating viral entry. Actin projections were associated with NCL interacting with RSV particles resting on apical cilia of the ALIs. We conclude that macropinocytosis-like actin projections protrude through normally protective cilia and mucus layers of stratified airway epithelium that helps present the IGF1R receptor and the NCL coreceptor to RSV particles waiting at the surface.

Respiratory syncytial virus entry mechanism in host cells: A general overview.

Respiratory syncytial virus (RSV) is a virus that causes acute respiratory infections in neonates and older adults. To infect host cells, the attachment glycoprotein (G) interacts with a cell surface receptor. This interaction determines the specific cell types that are susceptible to infection. RSV possesses a type I fusion protein F. Type I fusion proteins are metastable when rearrangement of the prefusion F occurs; the fusion peptide is exposed transforming the protein into postfusion form. The transition between the prefusion form and its postfusion form facilitates the viral envelope and the host cell membrane to fuse, enabling the virus to enter the host cell. Understanding the entry mechanism employed by RSV is crucial for developing effective antiviral therapies. In this review, we will discuss the various types of viral fusion proteins and explore the potential entry mechanisms utilized by RSV. A deeper understanding of these mechanisms will provide valuable insights for the development of novel approaches to treat RSV infections.

A Pseudovirus-Based Entry Assay to Evaluate Neutralizing Activity against Respiratory Syncytial Virus.

Respiratory syncytial virus (RSV) infection can cause life-threatening pneumonia and bronchiolitis, posing a significant threat to human health worldwide, especially to children and the elderly. Currently, there is no specific treatment for RSV infection. The most effective measures for preventing RSV infection are vaccines and prophylactic medications. However, not all population groups are eligible for the approved vaccines or antibody-based preventive medications. Therefore, there is an urgent need to develop novel vaccines and prophylactic drugs available for people of all ages. High-throughput assays that evaluate the efficacy of viral entry inhibitors or vaccine-induced neutralizing antibodies in blocking RSV entry are crucial for evaluating vaccine and prophylactic drug candidates. We developed an efficient entry assay using a lentiviral pseudovirus carrying the fusion (F) protein of type A or B RSV. In addition, the essential parameters were systematically optimized, including the number of transfected plasmids, storage conditions of the pseudovirus, cell types, cell numbers, virus inoculum, and time point of detection. Furthermore, the convalescent sera exhibited comparable inhibitory activity in this assay as in the authentic RSV virus neutralization assay. We established a robust pseudovirus-based entry assay for RSV, which holds excellent promise for studying entry mechanisms, evaluating viral entry inhibitors, and assessing vaccine-elicited neutralizing antibodies against RSV.

The RSV F p27 peptide: current knowledge, important questions.

Respiratory syncytial virus (RSV) remains a leading cause of hospitalizations and death for young children and adults over 65. The worldwide impact of RSV has prioritized the search for an RSV vaccine, with most targeting the critical fusion (F) protein. However, questions remain about the mechanism of RSV entry and RSV F triggering and fusion promotion. This review highlights these questions, specifically those surrounding a cleaved 27 amino acids long peptide within F, p27.

Discovery of N-Containing (-)-Borneol Esters as Respiratory Syncytial Virus Fusion Inhibitors.

Respiratory syncytial virus (RSV) causes acute respiratory infections, thus, posing a serious threat to the health of infants, children, and elderly people. In this study, we have discovered a series of potent RSV entry inhibitors with the (-)-borneol scaffold. The active compounds 3b, 5a, 5c, 7b, 9c, 10b, 10c, and 14b were found to exhibit activity against RSV A strain A2 in HEp-2 cells. The most active substances, 3b (IC50 = 8.9 μM, SI = 111) and 5a (IC50 = 5.0 μM, SI = 83), displayed more potency than the known antiviral agent Ribavirin (IC50 = 80.0 μM, SI = 50). Time-of-addition assay and temperature shift studies demonstrated that compounds 3b, 5a, and 6b inhibited RSV entry, probably by interacting with the viral F protein that mediated membrane fusion, while they neither bound to G protein nor inhibited RSV attachment to the target cells. Appling procedures of molecular modeling and molecular dynamics, the binding mode of compounds 3b and 5a was proposed. Taken together, the results of this study suggest (-)-borneol esters to be promising lead compounds for developing new anti-RSV agents.

Prefusion-specific antibody-derived peptides trivalently presented on DNA-nanoscaffolds as an innovative strategy against RSV entry

Human respiratory syncytial virus (RSV) is the primary cause of acute lower respiratory tract infections in children and the elderly worldwide, for which neither a vaccine nor an effective therapy is approved. The entry of RSV into the host cell is mediated by stepwise structural changes in the surface RSV fusion (RSV-F) glycoprotein. Recent progress in structural and functional studies of RSV-F glycoprotein revealed conformation-dependent neutralizing epitopes which have become attractive targets for vaccine and therapeutic development. As RSV-F is present on viral surface in a trimeric form, a trivalent binding interaction between a candidate fusion inhibitor and the respective epitopes on each of the three monomers is expected to prevent viral infection at higher potency than a monovalent or bivalent inhibitor. Here we demonstrate a novel RSV entry inhibitory approach by implementing a trimeric DNA nanostructure as a template to display up to three linear peptide moieties that simultaneously target an epitope on the surface of the prefusion RSV-F protein. In order to design synthetic binding peptides that can be coupled to the DNA nanostructure, the prefusion RSV-F-specific monoclonal antibody (D25) was selected. Complementarity-determining region 3 (CDR3) derived peptides underwent truncation and alanine-scanning mutagenesis analysis, followed by systematic sequence modifications using non-canonical amino acids. The most effective peptide candidate was used as a binding moiety to functionalize the DNA nanostructure. The designed DNA-peptide construct was able to block RSV infection on cells more efficiently than the monomeric peptides, however a more moderate reduction of viral load was observed in the lungs of infected mice upon intranasal application, likely due to dissociation or absorption of the underlying DNA structure by cells in the lungs. Taken together, our results point towards the inhibitory potential of a novel trimeric DNA-peptide based approach against RSV and open the possibility to apply this platform to target other viral infections.

Green Synthesis-Mediated Nanoparticles and Their Curative Character Against Post COVID-19 Skin Diseases.

Purpose of ReviewThis article provides the explanatory manuscript regarding the SARS-Corona virus 2. Sub-titled as the history of single-stranded RNA, internal characters of COVID-19, resource, the life cycle of COVID-19, reservoir of the disease, secondary infections of COVID-19 and nano herbal remedy.Recent FindingsThe skin is not the main target of the SARS-corona virus 2 infections but somehow directly or indirectly, it causes exacerbating eruptions on the skin. Recent research shows that curcumin–mediated synthesized AgNPs show its potential character in the entry of respiratory syncytial virus (RSV), blocks interaction with the viral surface, and damages the viral protein. In modern days, molecular docking studies fabricated copper iodide flower extract (CuI-FE) which shows tough inhibitory action against COVID-19. Many articles show green synthesis–mediated nanoparticles like silver, gold, zinc, copper, iron, titanium dioxide, selenium, and cadmium which possess high anti-viricidal activity.SummaryThe anti-oxidant, anti-viral, anti-inflammatory, anti-hive rich plant–mediated nanoparticle synthesis might be an alternative betterment, cost-effective, and eco-friendly medication for the skin disease caused by SARS-corona virus 2 (the viral clinical signs are itchy, hives, rashes, papules, psoriasis, and inflammation) and (non-viral clinical signs-pressure urticaria, contact dermatitis, and acne) that occurred as the result of COVID-19.

Inhibition of Respiratory Syncytial Virus Infection by Small Non-Coding RNA Fragments

Respiratory syncytial virus (RSV) causes acute lower respiratory tract infection in infants, immunocompromised individuals and the elderly. As the only current specific treatment options for RSV are monoclonal antibodies, there is a need for efficacious antiviral treatments against RSV to be developed. We have previously shown that a group of synthetic non-coding single-stranded DNA oligonucleotides with lengths of 25–40 nucleotides can inhibit RSV infection in vitro and in vivo. Based on this, herein, we investigate whether naturally occurring single-stranded small non-coding RNA (sncRNA) fragments present in the airways have antiviral effects against RSV infection. From publicly available sequencing data, we selected sncRNA fragments such as YRNAs, tRNAs and rRNAs present in human bronchoalveolar lavage fluid (BALF) from healthy individuals. We utilized a GFP-expressing RSV to show that pre-treatment with the selected sncRNA fragments inhibited RSV infection in A549 cells in vitro. Furthermore, by using a flow cytometry-based binding assay, we demonstrate that these naturally occurring sncRNAs fragments inhibit viral infection most likely by binding to the RSV entry receptor nucleolin and thereby preventing the virus from binding to host cells, either directly or via steric hindrance. This finding highlights a new function of sncRNAs and displays the possibility of using naturally occurring sncRNAs as treatments against RSV.

Evaluation of Small Molecule Combinations against Respiratory Syncytial Virus In Vitro.

Respiratory syncytial virus (RSV) is a major pathogen that causes severe lower respiratory tract infection in infants, the elderly and the immunocompromised worldwide. At present no approved specific drugs or vaccines are available to treat this pathogen. Recently, several promising candidates targeting RSV entry and multiplication steps are under investigation. However, it is possible to lead to drug resistance under the long-term treatment. Therapeutic combinations constitute an alternative to prevent resistance and reduce antiviral doses. Therefore, we tested in vitro two-drug combinations of fusion inhibitors (GS5806, Ziresovir and BMS433771) and RNA-dependent RNA polymerase complex (RdRp) inhibitors (ALS8176, RSV604, and Cyclopamine). The statistical program MacSynergy II was employed to determine synergism, additivity or antagonism between drugs. From the result, we found that combinations of ALS8176 and Ziresovir or GS5806 exhibit additive effects against RSV in vitro, with interaction volume of 50 µM2% and 31 µM2% at 95% confidence interval, respectively. On the other hand, all combinations between fusion inhibitors showed antagonistic effects against RSV in vitro, with volume of antagonism ranging from −50 µM2 % to −176 µM2 % at 95% confidence interval. Over all, our results suggest the potentially therapeutic combinations in combating RSV in vitro could be considered for further animal and clinical evaluations.

Respiratory Syncytial Virus and Human Metapneumovirus Infections in Three-Dimensional Human Airway Tissues Expose an Interesting Dichotomy in Viral Replication, Spread, and Inhibition by Neutralizing Antibodies.

Respiratory syncytial virus (RSV) and human metapneumovirus (HMPV) are two of the leading causes of respiratory infections in children and elderly and immunocompromised patients worldwide. There is no approved treatment for HMPV and only one prophylactic treatment against RSV, palivizumab, for high-risk infants. Better understanding of the viral lifecycles in a more relevant model system may help identify novel therapeutic targets. By utilizing three-dimensional (3-D) human airway tissues to examine viral infection in a physiologically relevant model system, we showed that RSV infects and spreads more efficiently than HMPV, with the latter requiring higher multiplicities of infection (MOIs) to yield similar levels of infection. Apical ciliated cells were the target for both viruses, but RSV apical release was significantly more efficient than HMPV. In RSV- or HMPV-infected cells, cytosolic inclusion bodies containing the nucleoprotein, phosphoprotein, and respective viral genomic RNA were clearly observed in human airway epithelial (HAE) culture. In HMPV-infected cells, actin-based filamentous extensions were more common (35.8%) than those found in RSV-infected cells (4.4%). Interestingly, neither RSV nor HMPV formed syncytia in HAE tissues. Palivizumab and nirsevimab effectively inhibited entry and spread of RSV in HAE tissues, with nirsevimab displaying significantly higher potency than palivizumab. In contrast, 54G10 completely inhibited HMPV entry but only modestly reduced viral spread, suggesting HMPV may use alternative mechanisms for spread. These results represent the first comparative analysis of infection by the two pneumoviruses in a physiologically relevant model, demonstrating an interesting dichotomy in the mechanisms of infection, spread, and consequent inhibition of the viral lifecycles by neutralizing monoclonal antibodies.IMPORTANCE Respiratory syncytial virus and human metapneumovirus are leading causes of respiratory illness worldwide, but limited treatment options are available. To better target these viruses, we examined key aspects of the viral life cycle in three-dimensional (3-D) human airway tissues. Both viruses establish efficient infection through the apical surface, but efficient spread and apical release were seen for respiratory syncytial virus (RSV) but not human metapneumovirus (HMPV). Both viruses form inclusion bodies, minimally composed of nucleoprotein (N), phosphoprotein (P), and viral RNA (vRNA), indicating that these structures are critical for replication in this more physiological model. HMPV formed significantly more long, filamentous actin-based extensions in human airway epithelial (HAE) tissues than RSV, suggesting HMPV may promote cell-to-cell spread via these extensions. Lastly, RSV entry and spread were fully inhibited by neutralizing antibodies palivizumab and the novel nirsevimab. In contrast, while HMPV entry was fully inhibited by 54G10, a neutralizing antibody, spread was only modestly reduced, further supporting a cell-to-cell spread mechanism.

Inhibitory Effect of PIK-24 on Respiratory Syncytial Virus Entry by Blocking Phosphatidylinositol-3 Kinase Signaling.

Phosphoinositide-3 kinase signaling modulates many cellular processes, including cell survival, proliferation, differentiation, and apoptosis. Currently, it is known that the establishment of respiratory syncytial virus infection requires phosphoinositide-3 kinase signaling. However, the regulatory pattern of phosphoinositide-3 kinase signaling or its corresponding molecular mechanism during respiratory syncytial virus entry remains unclear. Here, the involvement of phosphoinositide-3 kinase signaling in respiratory syncytial virus entry was studied. PIK-24, a novel compound designed with phosphoinositide-3 kinase as a target, had potent anti-respiratory syncytial virus activity both in vitro and in vivo PIK-24 significantly reduced viral entry into the host cell through blocking the late stage of the fusion process. In a mouse model, PIK-24 effectively reduced the viral load and alleviated inflammation in lung tissue. Subsequent studies on the antiviral mechanism of PIK-24 revealed that viral entry was accompanied by phosphoinositide-3 kinase signaling activation, downstream RhoA and cofilin upregulation, and actin cytoskeleton rearrangement. PIK-24 treatment significantly reversed all these effects. The disruption of actin cytoskeleton dynamics or the modulation of phosphoinositide-3 kinase activity by knockdown also affected viral entry efficacy. Altogether, it is reasonable to conclude that the antiviral activity of PIK-24 depends on the phosphoinositide-3 kinase signaling and that the use of phosphoinositide-3 kinase signaling to regulate actin cytoskeleton rearrangement plays a key role in respiratory syncytial virus entry.

IGF1R is an entry receptor for respiratory syncytial virus.

Pneumonia resulting from infection is one of the leading causes of death worldwide. Pulmonary infection by the respiratory syncytial virus (RSV) is a large burden on human health, for which there are few therapeutic options1. RSV targets ciliated epithelial cells in the airways, but how viruses such as RSV interact with receptors on these cells is not understood. Nucleolin is an entry coreceptor for RSV2 and also mediates the cellular entry of influenza, the parainfluenza virus, some enteroviruses and the bacterium that causes tularaemia3,4. Here we show a mechanism of RSV entry into cells in which outside-in signalling, involving binding of the prefusion RSV-F glycoprotein withthe insulin-like growth factor-1 receptor, triggers the activation of protein kinase C zeta (PKCζ). This cellular signalling cascade recruits nucleolin from the nuclei of cells to the plasma membrane, where it also binds to RSV-F on virions. We find that inhibiting PKCζ activation prevents the trafficking of nucleolin to RSV particles on airway organoid cultures, and reduces viral replication and pathology in RSV-infected mice. These findings reveal a mechanism of virus entry in which receptor engagement and signal transduction bring the coreceptor to viral particles at the cell surface, and could form the basis of new therapeutics to treat RSV infection.

Structural Insight into Paramyxovirus and Pneumovirus Entry Inhibition.

Paramyxoviruses and pneumoviruses infect cells through fusion (F) protein-mediated merger of the viral envelope with target membranes. Members of these families include a range of major human and animal pathogens, such as respiratory syncytial virus (RSV), measles virus (MeV), human parainfluenza viruses (HPIVs), and highly pathogenic Nipah virus (NiV). High-resolution F protein structures in both the metastable pre- and the postfusion conformation have been solved for several members of the families and a number of F-targeting entry inhibitors have progressed to advanced development or clinical testing. However, small-molecule RSV entry inhibitors have overall disappointed in clinical trials and viral resistance developed rapidly in experimental settings and patients, raising the question of whether the available structural information may provide a path to counteract viral escape through proactive inhibitor engineering. This article will summarize current mechanistic insight into F-mediated membrane fusion and examine the contribution of structural information to the development of small-molecule F inhibitors. Implications are outlined for future drug target selection and rational drug engineering strategies.