Structure of Semliki Forest virus in complex with its receptor VLDLR.

Semliki Forest virus (SFV) is an alphavirus that uses the very-low-density lipoprotein receptor (VLDLR) as a receptor during infection of its vertebrate hosts and insect vectors. Herein, we used cryoelectron microscopy to study the structure of SFV in complex with VLDLR. We found that VLDLR binds multiple E1-DIII sites of SFV through its membrane-distal LDLR class A (LA) repeats. Among the LA repeats of the VLDLR, LA3 has the best binding affinity to SFV. The high-resolution structure shows that LA3 binds SFV E1-DIII through a small surface area of 378 Å2, with the main interactions at the interface involving salt bridges. Compared with the binding of single LA3s, consecutive LA repeats around LA3 promote synergistic binding to SFV, during which the LAs undergo a rotation, allowing simultaneous key interactions at multiple E1-DIII sites on the virion and enabling the binding of VLDLRs from divergent host species to SFV.

Neutralizing immunity in vaccine breakthrough infections from the SARS-CoV-2 Omicron and Delta variants.

Virus-like particle (VLP) and live virus assays were used to investigate neutralizing immunity against Delta and Omicron SARS-CoV-2 variants in 259 samples from 128 vaccinated individuals. Following Delta breakthrough infection, titers against WT rose 57-fold and 3.1-fold compared with uninfected boosted and unboosted individuals, respectively, versus only a 5.8-fold increase and 3.1-fold decrease for Omicron breakthrough infection. Among immunocompetent, unboosted patients, Delta breakthrough infections induced 10.8-fold higher titers against WT compared with Omicron (p = 0.037). Decreased antibody responses in Omicron breakthrough infections relative to Delta were potentially related to a higher proportion of asymptomatic or mild breakthrough infections (55.0% versus 28.6%, respectively), which exhibited 12.3-fold lower titers against WT compared with moderate to severe infections (p = 0.020). Following either Delta or Omicron breakthrough infection, limited variant-specific cross-neutralizing immunity was observed. These results suggest that Omicron breakthrough infections are less immunogenic than Delta, thus providing reduced protection against reinfection or infection from future variants.

B Cells Are the Dominant Antigen-Presenting Cells that Activate Naive CD4+ T Cells upon Immunization with a Virus-Derived Nanoparticle Antigen.

B cells can present antigens to CD4+ T cells, but it is thought that dendritic cells (DCs) are the primary initiators of naive CD4+ T cell responses. Nanoparticles, including virus-like particles (VLPs), are attractive candidates as carriers for vaccines and drug delivery. Using RNA phage Qß-derived VLP (Qß-VLP) as a model antigen, we found that antigen-specific B cells were the dominant antigen-presenting cells that initiated naive CD4+ T cell activation. B cells were sufficient to induce T follicular helper cell development in the absence of DCs. Qß-specific B cells promoted CD4+ T cell proliferation and differentiation via cognate interactions and through Toll-like receptor signaling-mediated cytokine production. Antigen-specific B cells were also involved in initiating CD4+ T cell responses during immunization with inactivated influenza virus. These findings have implications for the rational design of nanoparticles as vaccine candidates, particularly for therapeutic vaccines that aim to break immune tolerance.

Human paraneoplastic antigen Ma2 (PNMA2) forms icosahedral capsids that can be engineered for mRNA delivery.

A number of endogenous genes in the human genome encode retroviral gag-like proteins, which were domesticated from ancient retroelements. The paraneoplastic Ma antigen (PNMA) family members encode a gag-like capsid domain, but their ability to assemble as capsids and traffic between cells remains mostly uncharacterized. Here, we systematically investigate human PNMA proteins and find that a number of PNMAs are secreted by human cells. We determine that PNMA2 forms icosahedral capsids efficiently but does not naturally encapsidate nucleic acids. We resolve the cryoelectron microscopy (cryo-EM) structure of PNMA2 and leverage the structure to design engineered PNMA2 (ePNMA2) particles with RNA packaging abilities. Recombinantly purified ePNMA2 proteins package mRNA molecules into icosahedral capsids and can function as delivery vehicles in mammalian cell lines, demonstrating the potential for engineered endogenous capsids as a nucleic acid therapy delivery modality.

Virus-like Particles Identify an HIV V1V2 Apex-Binding Neutralizing Antibody that Lacks a Protruding Loop.

Most HIV-1-specific neutralizing antibodies isolated to date exhibit unusual characteristics that complicate their elicitation. Neutralizing antibodies that target the V1V2 apex of the HIV-1 envelope (Env) trimer feature unusually long protruding loops, which enable them to penetrate the HIV-1 glycan shield. As antibodies with loops of requisite length are created through uncommon recombination events, an alternative mode of apex binding has been sought. Here, we isolated a lineage of Env apex-directed neutralizing antibodies, N90-VRC38.01-11, by using virus-like particles and conformationally stabilized Env trimers as B cell probes. A crystal structure of N90-VRC38.01 with a scaffolded V1V2 revealed a binding mode involving side-chain-to-side-chain interactions that reduced the distance the antibody loop must traverse the glycan shield, thereby facilitating V1V2 binding via a non-protruding loop. The N90-VRC38 lineage thus identifies a solution for V1V2-apex binding that provides a more conventional B cell pathway for vaccine design.

A tetravalent dengue virus-like particle vaccine induces high levels of neutralizing antibodies and reduces dengue replication in non-human primates.

Dengue virus (DENV) represents a significant global health burden, with 50% of the world's population at risk of infection, and there is an urgent need for next-generation vaccines. Virus-like particle (VLP)-based vaccines, which mimic the antigenic structure of the virus but lack the viral genome, are an attractive approach. Here, we describe a dengue VLP (DENVLP) vaccine which generates a neutralizing antibody response against all four DENV serotypes in 100% of immunized non-human primates for up to 1 year. Additionally, DENVLP vaccination produced no ADE response against any of four DENV serotypes in vitro. DENVLP vaccination reduces viral replication in a non-human primate challenge model. We also show that transfer of purified IgG from immunized monkeys into immunodeficient mice protects against subsequent lethal DENV challenge, indicating a humoral mechanism of protection. These results indicate that this DENVLP vaccine is immunogenic and can be considered for clinical evaluation. Immunization of non-human primates with a tetravalent DENVLP vaccine induces high levels of neutralizing antibodies and reduces the severity of infection for all four dengue serotypes.IMPORTANCEDengue is a viral disease that infects nearly 400 million people worldwide and causes dengue hemorrhagic fever, which is responsible for 10,000 deaths each year. Currently, there is no therapeutic drug licensed to treat dengue infection, which makes the development of an effective vaccine essential. Virus-like particles (VLPs) are a safe and highly immunogenic platform that can be used in young children, immunocompromised individuals, as well as healthy adults. In this study, we describe the development of a dengue VLP vaccine and demonstrate that it induces a robust immune response against the dengue virus for over 1 year in monkeys. The immunity induced by this vaccine reduced live dengue infection in both murine and non-human primate models. These results indicate that our dengue VLP vaccine is a promising vaccine candidate.

Virus-like particles displaying the mature C-terminal domain of filamentous hemagglutinin are immunogenic and protective against Bordetella pertussis respiratory infection in mice.

Bordetella pertussis, the bacterium responsible for whooping cough, remains a significant public health challenge despite the existing licensed pertussis vaccines. Current acellular pertussis vaccines, though having favorable reactogenicity and efficacy profiles, involve complex and costly production processes. In addition, acellular vaccines have functional challenges such as short-lasting duration of immunity and limited antigen coverage. Filamentous hemagglutinin (FHA) is an adhesin of B. pertussis that is included in all multivalent pertussis vaccine formulations. Antibodies to FHA have been shown to prevent bacterial attachment to respiratory epithelial cells, and T cell responses to FHA facilitate cell-mediated immunity. In this study, FHA's mature C-terminal domain (MCD) was evaluated as a novel vaccine antigen. MCD was conjugated to virus-like particles via SpyTag-SpyCatcher technology. Prime-boost vaccine studies were performed in mice to characterize immunogenicity and protection against the intranasal B. pertussis challenge. MCD-SpyVLP was more immunogenic than SpyTag-MCD antigen alone, and in Tohama I strain challenge studies, improved protection against challenge was observed in the lungs at day 3 and in the trachea and nasal wash at day 7 post-challenge. Furthermore, a B. pertussis strain encoding genetically inactivated pertussis toxin was used to evaluate MCD-SpyVLP vaccine immunity. Mice vaccinated with MCD-SpyVLP had significantly lower respiratory bacterial burden at both days 3 and 7 post-challenge compared to mock-vaccinated animals. Overall, these data support the use of SpyTag-SpyCatcher VLPs as a platform for use in vaccine development against B. pertussis and other pathogens.

A SARS-CoV-2 peptide antigen purified from bacteria and displayed in a high-density repetitive manner on a virus-like particle could generate anti-SARS-CoV-2 neutralizing antibodies unlike free peptide.

SARS-CoV-2 is an enveloped virus. Proteins of the lipid based envelope are not rigidly arranged as compared to non-enveloped viruses lacking lipid based outer layer. Rigidly arranged multivalent display has been reported to be important for potent immune stimulation. We assembled himeric virus-like-particle (VLP) where the core of the particle is composed of the coat protein of Acinetobacter phage. Peptide-based antigen from receptor binding domain (RBD) of SARS-CoV-2 spike protein has been displayed on the coat based VLP matrix using spy-tag/spy-catcher split inteine conjugation system that allows spontaneous, irreversible isopeptide bond formation. Both the matrix as well as peptide have been purified from bacteria which is an easy platform for protein purification. We used the closed state of spike trimer for epitope mapping as this is the conformation of the spike that the immune system visualizes unlike the open state that appears when the virus tries to attach to receptor. Mice based immunization studies were used to test immunogenicity of the antigens. The work demonstrates that peptide-based antigens displayed in high densities can induce neutralizing antibody production unlike free peptide. Careful choice of peptides can deliver better candidates. Also, small size allows easy improvisation. Production in bacteria offers cheaper and robust purification option.

Distinct motifs in the E protein are required for SARS-CoV-2 virus particle formation and lysosomal deacidification in host cells.

IMPORTANCE: Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2), the virus responsible for coronavirus disease 2019 (COVID-19), has caused a global public health crisis. The E protein, a structural protein found in this virus particle, is also known to be a viroporin. As such, it forms oligomeric ion channels or pores in the host cell membrane. However, the relationship between these two functions is poorly understood. In this study, we showed that the roles of E protein in virus particle and viroporin formation are distinct. This study contributes to the development of drugs that inhibit SARS-CoV-2 virus particle formation. Additionally, we designed a highly sensitive and high-throughput virus-like particle detection system using the HiBiT tag, which is a useful tool for studying the release of SARS-CoV-2.

Intraocular mRNA delivery with endogenous MmPEG10-based virus-like particles.

Virus-like particles (VLP) are a promising tool for intracellular gene delivery, yet their potential in ocular gene therapy remains underexplored. In this study, we bridged this knowledge gap by demonstrating the successful generation and application of vesicular stomatitis virus glycoprotein (VSVG)-pseudotyped mouse PEG10 (MmPEG10)-VLP for intraocular mRNA delivery. Our findings revealed that PEG10-VLP can efficiently deliver GFP mRNA to adult retinal pigment epithelial cell line-19 (ARPE-19) cells, leading to transient expression. Moreover, we showed that MmPEG10-VLP can transfer SMAD7 to inhibit epithelial-mesenchymal transition (EMT) in RPE cells effectively. In vivo experiments further substantiated the potential of these vectors, as subretinal delivery into adult mice resulted in efficient transduction of retinal pigment epithelial (RPE) cells and GFP reporter gene expression without significant immune response. However, intravitreal injection did not yield efficient ocular expression. We also evaluated the transduction characteristics of MmPEG10-VLP following intracameral delivery, revealing transient GFP protein expression in corneal endothelial cells without significant immunotoxicities. In summary, our study established that VSVG pseudotyped MmPEG10-based VLP can transduce mitotically inactive RPE cells and corneal endothelial cells in vivo without triggering an inflammatory response, underscoring their potential utility in ocular gene therapy.

Production of VP3-only virus-like particles of Adeno-associated virus 2 in E. coli cells.

Adeno-associated Virus (AAV) is a promising vector for gene therapy. However, few studies have focused on producing virus-like particles (VLPs) of AAV in cells, especially in E. coli. In this study, we describe a method to produce empty VP3-only VLPs of AAV2 in E. coli by co-expressing VP3 and assembly-activating protein (AAP) of AAV2. Although the yields of VLPs produced with our method were low, the VLPs were able to self-assemble in E. coli without the need of in vitro capsid assembly. The produced VLPs were characterized by immunological detection and transmission electron microscopy (TEM). In conclusion, this study demonstrated that capsid assembly of AAV2 is possible in E. coli, and E. coli may be a candidate system for production of VLPs of AAV.

Significance of VLPs in Vlp-circRNA vaccines: a vaccine candidate or delivery vehicle?

Circular RNAs (circRNAs) are a class of single-stranded RNAs with a closed loop lacking 5' and 3' ends. These circRNAs are translatable and, therefore, have a potential in developing vaccine. CircRNA vaccines have been shown to be more stable, safe, easy to manufacture and scale-up production when compared to mRNA vaccines. However, these vaccines also suffer from several drawbacks such as low circularization efficiency for longer RNA precursor and usage of lipid nano particles (LNPs) in their delivery. LNPs have been shown to require large amounts of RNA due to their indirect delivery from endosome to cytosol. Besides, individual components of LNPs provide reactogenicity. Usage of virus like particles (VLPs) can improve the increased production and targeted delivery of circRNA vaccines and show no reactogenicity. Moreover, VLPs has also been used to produce vaccines against several diseases such as hepatitis C virus (HCV) etc. In this article, we will discuss about the methods used to enhance synthesis or circularization efficiency of circRNA. Moreover, we will also discuss about the significance of VLPs as the delivery vehicle for circRNA and their possible usage as the dual vaccine.

Asynchronous Code Division Multiplexing-Based Visible Light Positioning and Communication Network Using Successive Interference Cancellation Decoding.

In the evolving landscape of sixth-generation wireless communication, the integration of visible light communication (VLC) and visible light positioning (VLP), known as visible light positioning and communication (VLPC), emerges as a pivotal technology. This study addresses the challenges of asynchronous code division multiplexing (ACDM) in VLPC networks, focusing on the enhancement of data transmission quality and positioning accuracy. Firstly, we propose an orthogonal pseudo-random code (OPRC) for ACDM-based VLP systems. Leveraging its excellent correlation properties, VLP signals preserve orthogonality even amidst asynchronous transmissions, achieving sub-centimeter average positioning errors. Next, by combining OPRC with successive interference cancellation decoding (SICD), we propose an enhanced ACDM-based VLPC system that utilizes OPRC for improved signal orthogonality and SICD for progressive elimination of multiple access interference (MAI) among VLPC signals. The results show substantial improvements in bit-error rate (BER) and positioning error (PE), approaching the performance levels observed in synchronized VLPC systems. Specifically, the SICD-OPRC scheme reduces average BER to 4.3 × 10-4 and average PE to 2.7 cm, demonstrating its robustness and superiority in complex asynchronous scenarios.

Real-Time Indoor Visible Light Positioning (VLP) Using Long Short Term Memory Neural Network (LSTM-NN) with Principal Component Analysis (PCA).

New applications such as augmented reality/virtual reality (AR/VR), Internet-of-Things (IOT), autonomous mobile robot (AMR) services, etc., require high reliability and high accuracy real-time positioning and tracking of persons and devices in indoor areas. Among the different visible-light-positioning (VLP) schemes, such as proximity, time-of-arrival (TOA), time-difference-of-arrival (TDOA), angle-of-arrival (AOA), and received-signal-strength (RSS), the RSS scheme is relatively easy to implement. Among these VLP methods, the RSS method is simple and efficient. As the received optical power has an inverse relationship with the distance between the LED transmitter (Tx) and the photodiode (PD) receiver (Rx), position information can be estimated by studying the received optical power from different Txs. In this work, we propose and experimentally demonstrate a real-time VLP system utilizing long short-term memory neural network (LSTM-NN) with principal component analysis (PCA) to mitigate high positioning error, particularly at the positioning unit cell boundaries. Experimental results show that in a positioning unit cell of 100 × 100 × 250 cm3, the average positioning error is 5.912 cm when using LSTM-NN only. By utilizing the PCA, we can observe that the positioning accuracy can be significantly enhanced to 1.806 cm, particularly at the unit cell boundaries and cell corners, showing a positioning error reduction of 69.45%. In the cumulative distribution function (CDF) measurements, when using only the LSTM-NN model, the positioning error of 95% of the experimental data is >15 cm; while using the LSTM-NN with PCA model, the error is reduced to <5 cm. In addition, we also experimentally demonstrate that the proposed real-time VLP system can also be used to predict the direction and the trajectory of the moving Rx.

Accurate Low Complexity Quadrature Angular Diversity Aperture Receiver for Visible Light Positioning.

Despite the many potential applications of an accurate indoor positioning system (IPS), no universal, readily available system exists. Much of the IPS research to date has been based on the use of radio transmitters as positioning beacons. Visible light positioning (VLP) instead uses LED lights as beacons. Either cameras or photodiodes (PDs) can be used as VLP receivers, and position estimates are usually based on either the angle of arrival (AOA) or the strength of the received signal. Research on the use of AOA with photodiode receivers has so far been limited by the lack of a suitable compact receiver. The quadrature angular diversity aperture receiver (QADA) can fill this gap. In this paper, we describe a new QADA design that uses only three readily available parts: a quadrant photodiode, a 3D-printed aperture, and a programmable system on a chip (PSoC). Extensive experimental results demonstrate that this design provides accurate AOA estimates within a room-sized test chamber. The flexibility and programmability of the PSoC mean that other sensors can be supported by the same PSoC. This has the potential to allow the AOA estimates from the QADA to be combined with information from other sensors to form future powerful sensor-fusion systems requiring only one beacon.

A Multifunctionalized Potyvirus-Derived Nanoparticle That Targets and Internalizes into Cancer Cells.

Plant viral nanoparticles (VNPs) are attractive to nanomedicine researchers because of their safety, ease of production, resistance, and straightforward functionalization. In this paper, we developed and successfully purified a VNP derived from turnip mosaic virus (TuMV), a well-known plant pathogen, that exhibits a high affinity for immunoglobulins G (IgG) thanks to its functionalization with the Z domain of staphylococcal Protein A via gene fusion. We selected cetuximab as a model IgG to demonstrate the versatility of this novel TuMV VNP by developing a fluorescent nanoplatform to mark tumoral cells from the Cal33 line of a tongue squamous cell carcinoma. Using confocal microscopy, we observed that fluorescent VNP-cetuximab bound selectively to Cal33 and was internalized, revealing the potential of this nanotool in cancer research.

A Bivalent Human Norovirus Vaccine Induces Homotypic and Heterotypic Neutralizing Antibodies.

A GII.2 outbreak in an efficacy study of a bivalent virus-like particle (VLP) norovirus vaccine, TAK-214, in healthy US adults provided an opportunity to examine GII.4 homotypic vs. GII.2 heterotypic responses to vaccination and infection. Three serological assays (VLP-binding, histoblood group antigen-blocking, and neutralizing) were performed for each genotype. Results were highly correlated within a genotype but not between genotypes. Although the vaccine provided protection from GII.2-associated disease, little GII.2-specific neutralization occurred after vaccination. Choice of antibody assay can affect assessments of human norovirus vaccine immunogenicity.

Production of antigenically stable enterovirus A71 virus-like particles in Pichia pastoris as a vaccine candidate.

Enterovirus A71 (EVA71) causes widespread disease in young children with occasional fatal consequences. In common with other picornaviruses, both empty capsids (ECs) and infectious virions are produced during the viral lifecycle. While initially antigenically indistinguishable from virions, ECs readily convert to an expanded conformation at moderate temperatures. In the closely related poliovirus, these conformational changes result in loss of antigenic sites required to elicit protective immune responses. Whether this is true for EVA71 remains to be determined and is the subject of this investigation.We previously reported the selection of a thermally resistant EVA71 genogroup B2 population using successive rounds of heating and passage. The mutations found in the structural protein-coding region of the selected population conferred increased thermal stability to both virions and naturally produced ECs. Here, we introduced these mutations into a recombinant expression system to produce stabilized virus-like particles (VLPs) in Pichia pastoris.The stabilized VLPs retain the native virion-like antigenic conformation as determined by reactivity with a specific antibody. Structural studies suggest multiple potential mechanisms of antigenic stabilization, however, unlike poliovirus, both native and expanded EVA71 particles elicited antibodies able to directly neutralize virus in vitro. Therefore, anti-EVA71 neutralizing antibodies are elicited by sites which are not canonically associated with the native conformation, but whether antigenic sites specific to the native conformation provide additional protective responses in vivo remains unclear. VLPs are likely to provide cheaper and safer alternatives for vaccine production and these data show that VLP vaccines are comparable with inactivated virus vaccines at inducing neutralising antibodies.

RG2-VLP: a Vaccine Designed to Broadly Protect against Anogenital and Skin Human Papillomaviruses Causing Human Cancer.

The family of human papillomaviruses (HPV) includes over 400 genotypes. Genus α genotypes generally infect the anogenital mucosa, and a subset of these HPV are a necessary, but not sufficient, cause of cervical cancer. Of the 13 high-risk (HR) and 11 intermediate-risk (IR) HPV associated with cervical cancer, genotypes 16 and 18 cause 50% and 20% of cases, respectively, whereas HPV16 dominates in other anogenital and oropharyngeal cancers. A plethora of ßHPVs are associated with cutaneous squamous cell carcinoma (CSCC), especially in sun-exposed skin sites of epidermodysplasia verruciformis (EV), AIDS, and immunosuppressed patients. Licensed L1 virus-like particle (VLP) vaccines, such as Gardasil 9, target a subset of αHPV but no ßHPV. To comprehensively target both α- and ßHPVs, we developed a two-component VLP vaccine, RG2-VLP, in which L2 protective epitopes derived from a conserved αHPV epitope (amino acids 17 to 36 of HPV16 L2) and a consensus ßHPV sequence in the same region are displayed within the DE loop of HPV16 and HPV18 L1 VLP, respectively. Unlike vaccination with Gardasil 9, vaccination of wild-type and EV model mice (Tmc6Δ/Δ or Tmc8Δ/Δ) with RG2-VLP induced robust L2-specific antibody titers and protected against ß-type HPV5. RG2-VLP protected rabbits against 17 αHPV, including those not covered by Gardasil 9. HPV16- and HPV18-specific neutralizing antibody responses were similar between RG2-VLP- and Gardasil 9-vaccinated animals. However, only transfer of RG2-VLP antiserum effectively protected naive mice from challenge with all ßHPVs tested. Taken together, these observations suggest RG2-VLP's potential as a broad-spectrum vaccine to prevent αHPV-driven anogenital, oropharyngeal, and ßHPV-associated cutaneous cancers. IMPORTANCE Licensed preventive HPV vaccines are composed of VLPs derived by expression of major capsid protein L1. They confer protection generally restricted to infection by the αHPVs targeted by the up-to-9-valent vaccine, and their associated anogenital cancers and genital warts, but do not target ßHPV that are associated with CSCC in EV and immunocompromised patients. We describe the development of a two-antigen vaccine protective in animal models against known oncogenic αHPVs as well as diverse ßHPVs by incorporation into HPV16 and HPV18 L1 VLP of 20-amino-acid conserved protective epitopes derived from minor capsid protein L2.

WWOX-Mediated Degradation of AMOTp130 Negatively Affects Egress of Filovirus VP40 Virus-Like Particles.

Ebola virus (EBOV) and Marburg virus (MARV) continue to emerge and cause severe hemorrhagic disease in humans. A comprehensive understanding of the filovirus-host interplay will be crucial for identifying and developing antiviral strategies. The filoviral VP40 matrix protein drives virion assembly and egress, in part by recruiting specific WW domain-containing host interactors via its conserved PPxY late (L) domain motif to positively regulate virus egress and spread. In contrast to these positive regulators of virus budding, a growing list of WW domain-containing interactors that negatively regulate virus egress and spread have been identified, including BAG3, YAP/TAZ, and WWOX. In addition to host WW domain regulators of virus budding, host PPxY-containing proteins also contribute to regulating this late stage of filovirus replication. For example, angiomotin (AMOT) is a multi-PPxY-containing host protein that functionally interacts with many of the same WW domain-containing proteins that regulate virus egress and spread. In this report, we demonstrate that host WWOX, which negatively regulates egress of VP40 virus-like particles (VLPs) and recombinant vesicular stomatitis virus (VSV) M40 virus, interacts with and suppresses the expression of AMOT. We found that WWOX disrupts AMOT's scaffold-like tubular distribution and reduces AMOT localization at the plasma membrane via lysosomal degradation. In sum, our findings reveal an indirect and novel mechanism by which modular PPxY-WW domain interactions between AMOT and WWOX regulate PPxY-mediated egress of filovirus VP40 VLPs. A better understanding of this modular network and competitive nature of protein-protein interactions will help to identify new antiviral targets and therapeutic strategies. IMPORTANCE Filoviruses (Ebola virus [EBOV] and Marburg virus [MARV]) are zoonotic, emerging pathogens that cause outbreaks of severe hemorrhagic fever in humans. A fundamental understanding of the virus-host interface is critical for understanding the biology of these viruses and for developing future strategies for therapeutic intervention. Here, we reveal a novel mechanism by which host proteins WWOX and AMOTp130 interact with each other and with the filovirus matrix protein VP40 to regulate VP40-mediated egress of virus-like particles (VLPs). Our results highlight the biological impact of competitive interplay of modular virus-host interactions on both the virus life cycle and the host cell.