Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development

Puckette, Michael; Clark, Benjamin A.; Smith, Justin D.; Turecek, Traci; Martel, Erica; Gabbert, Lindsay; Pisano, Melia; Hurtle, William; Pacheco, Juan M.; Barrera, José; Neilan, John G.; Rasmussen, Max V.

doi:10.1128/jvi.00924-17

Cited by 22 publications

(41 citation statements)

References 34 publications

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“…Presumably, a host protease is responsible for the 3CD cleavage seen, which occurs at a sequence close to the natural cleavage site. It would be interesting to fully ablate this cleavage event in Pichia or introduce mutations into 3C pro to reduce toxicity, as shown previously for foot-and-mouth disease virus, and determine the impact of these modifications on VLP production (44). We demonstrated that all three expression cassettes led to the correct processing of P1 through the detection of VP-1 using monoclonal antibody (MAb) 8650.…”

Section: Discussionmentioning

confidence: 58%

Comparative Molecular Biology Approaches for the Production of Poliovirus Virus-Like Particles Using Pichia pastoris

Sherry

Grehan

Snowden

et al. 2020

mSphere

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For enteroviruses such as poliovirus (PV), empty capsids, which are antigenically indistinguishable from mature virions, are produced naturally during viral infection. The production of such capsids recombinantly, in heterologous systems such as yeast, have great potential as virus-like particle (VLP) vaccine candidates. Here, using PV as an exemplar, we show the production of VLPs in Pichia pastoris by coexpression of the structural precursor protein P1 and the viral protease 3CD. The level of expression of the potentially cytotoxic protease relative to that of the P1 precursor was modulated by three different approaches: expression of the P1 precursor and protease from different transcription units, separation of the P1 and protease proteins using the Thosea asigna virus (TaV) 2A translation interruption sequence, or separation of the P1 and protease-coding sequences by an internal ribosome entry site sequence from Rhopalosiphum padi virus (RhPV). We also investigate the antigenicity of VLPs containing previously characterized mutations when produced in Pichia. Finally, using transmission electron microscopy and two-dimensional classification, we show that Pichia-derived VLPs exhibited the classical icosahedral capsid structure displayed by enteroviruses. IMPORTANCE Although the current poliovirus immunization program has been extremely successful in reducing the number of cases of paralytic polio worldwide, now more cases are caused by vaccine-derived polioviruses than by wild poliovirus. Switching to inactivated poliovirus vaccines will reduce this over time; however, their production requires the growth of large amounts of virus. This biosafety concern can be addressed by producing just the virus capsid. The capsid serves to protect the genetic material, which causes disease when introduced into a cell. Therefore, empty capsids (virus-like particles [VLPs]), which lack the viral RNA genome, are safe both to make and to use. We exploit yeast as a versatile model expression system to produce VLPs, and here we specifically highlight the potential of this system to supply next-generation poliovirus vaccines to secure a polio-free world for the future.

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Section: Discussionmentioning

confidence: 58%

Comparative Molecular Biology Approaches for the Production of Poliovirus Virus-Like Particles Using Pichia pastoris

Sherry

Grehan

Snowden

et al. 2020

mSphere

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“…This result shows that the structural proteins are able to determine the antigenic specificity [3]. Studies have shown that the availability of free polypeptide chains in zone A and the tap-folding barrels of zone B and aspartic acid residue R56 in surface protein of VP3 could determine the binding with polyclonal antibodies [31]. Similarly, VP3 protein is crucial in recognizing the cellular heparin sulfate (HS) co-receptor of FMDV [32].…”

Section: Discussionmentioning

confidence: 97%

Antigenicity and Immunogenicity Analysis of the E. coli Expressed FMDV Structural Proteins; VP1, VP0, VP3 of the South African Territories Type 2 Virus

Wubshet

Ding

et al. 2021

Viruses

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An alternative vaccine design approach and diagnostic kits are highly required against the anticipated pandemicity caused by the South African Territories type 2 (SAT2) Foot and Mouth Disease Virus (FMDV). However, the distinct antigenicity and immunogenicity of VP1, VP0, and VP3 of FMDV serotype SAT2 are poorly understood. Similarly, the particular roles of the three structural proteins in novel vaccine design and development remain unexplained. We therefore constructed VP1, VP0, and VP3 encoding gene (SAT2:JX014256 strain) separately fused with His-SUMO (histidine-small ubiquitin-related modifier) inserted into pET-32a cassette to express the three recombinant proteins and separately evaluated their antigenicity and immunogenicity in mice. The fusion protein was successfully expressed and purified by the Ni-NTA resin chromatography. The level of serum antibody, spleen lymphocyte proliferation, and cytokines against the three distinct recombinant proteins were analyzed. Results showed that the anti-FMDV humoral response was triggered by these proteins, and the fusion proteins did enhance the splenocyte immune response in the separately immunized mice. We observed low variations among the three fusion proteins in terms of the antibody and cytokine production in mice. Hence, in this study, results demonstrated that the structural proteins of SAT2 FMDV could be used for the development of immunodiagnostic kits and subunit vaccine designs.

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“…FMDV capsid precursor P1 is the precursor protein of VP1. P1 is finally processed into mature VP0, VP1, and VP3 proteins under the action of FMDV 3C protease (3C pro ) (32,33). In order to clarify whether the degradation of TPL2 is caused by the specificity of the VP1 protein in the P1 precursor, the effect of P1 on the expression level of TPL2 was first detected by Western blotting.…”

Section: Fmdv Vp1 Protein Promotes K48-linked Polyubiquitination Of Tpl2 and Degrades Tpl2 By The Proteasome Pathwaymentioning

confidence: 99%

Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

Zhang

Yan

Hao

et al. 2021

J Virol

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Tumor progression locus 2 (TPL2) is a serine/threonine kinase that belongs to the mitogen-activated protein 3 kinase (MAP3K) family, and it plays an important role in pathogen infection. The trimer complex of TPL2, p105, and ABIN2 is essential for maintenance of TPL2 steady-state levels and host cell response to pathogens. Foot-and-mouth disease virus (FMDV) is a positive-strand RNA virus of the family Picornaviridae that encodes proteins capable of antagonizing host immune responses to achieve infection. The VP1 protein of FMDV is a multifunctional protein that can bind host cells and induce an immune response as well as cell apoptosis. However, the role and mechanisms of TPL2 in FMDV infection remain unknown. Here, we determined that FMDV infection could inhibit TPL2, p105, and ABIN2 at the transcription and protein levels, while VP1 could only inhibit TPL2, p105 and ABIN2 at protein level. TPL2 inhibited the replication of FMDV in vivo and in vitro, the 268 to 283 amino-acid region in the TPL2 kinase domain was essential for interaction with VP1. Moreover, VP1 promoted K48-linked polyubiquitination of TPL2 and degraded TPL2 by the proteasome pathway. However, VP1-induced degradation of p105 and ABIN2 was independent of proteasome, autophagy, lysosome, and caspase-dependent pathways. Further studies showed that VP1 destroyed the stability of the TPL2-p105-ABIN2 complex. Taken together, these results revealed that VP1 antagonized TPL2-meditated antivirus activity by degrading TPL2 and destroying its complex. These findings may contribute to understand FMDV-host interactions and improve development of a novel vaccine to prevent FMDV infection. Importance Virus-host interactions are critical for virus infection. This study was the first to demonstrate the antiviral effect of host TPL2 during FMDV replication by increasing production of interferons and antiviral cytokines. Both FMDV and VP1 protein can reduce host TPL2, ABIN2 and p105 to destroy TPL2-p105-ABIN2 trimer complex. VP1 interacted with TPL2 and degrade TPL2 via proteasome pathway to repress TPL2-mediated antivirus activity. This study provided new insights into FMDV immune evasion mechanisms, elucidating new informations regarding FMDV counteraction of host antivirus activity.

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Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development

Cited by 22 publications

References 34 publications

Comparative Molecular Biology Approaches for the Production of Poliovirus Virus-Like Particles Using Pichia pastoris

Comparative Molecular Biology Approaches for the Production of Poliovirus Virus-Like Particles Using Pichia pastoris

Antigenicity and Immunogenicity Analysis of the E. coli Expressed FMDV Structural Proteins; VP1, VP0, VP3 of the South African Territories Type 2 Virus

Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

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