Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells
Background/Objectives: Baculoviruses represent promising gene delivery vectors for mammalian systems, combining high safety profiles with substantial cargo capacity. While pseudotyping with vesicular stomatitis virus G-protein (VSV-G) enhances transduction efficiency, optimal expression strategies d...
| Autores principales: | , , , , , , , |
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| Formato: | info:ar-repo/semantics/artículo |
| Lenguaje: | Inglés |
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MDPI
2025
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| Materias: | |
| Acceso en línea: | http://hdl.handle.net/20.500.12123/23052 https://www.mdpi.com/2076-393X/13/7/693 https://doi.org/10.3390/vaccines13070693 |
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| author | Simonin, Jorge Alejandro Cuccovia Warlet, Franco Uriel Bauzá, María del Rosario Plastine, María Del Pilar Alfonso, Victoria Olea, Fernanda Daniela Cerrudo, Carolina Susana Belaich, Mariano Nicolás |
| author_browse | Alfonso, Victoria Bauzá, María del Rosario Belaich, Mariano Nicolás Cerrudo, Carolina Susana Cuccovia Warlet, Franco Uriel Olea, Fernanda Daniela Plastine, María Del Pilar Simonin, Jorge Alejandro |
| author_facet | Simonin, Jorge Alejandro Cuccovia Warlet, Franco Uriel Bauzá, María del Rosario Plastine, María Del Pilar Alfonso, Victoria Olea, Fernanda Daniela Cerrudo, Carolina Susana Belaich, Mariano Nicolás |
| author_sort | Simonin, Jorge Alejandro |
| collection | INTA Digital |
| description | Background/Objectives: Baculoviruses represent promising gene delivery vectors for mammalian systems, combining high safety profiles with substantial cargo capacity. While pseudotyping with vesicular stomatitis virus G-protein (VSV-G) enhances transduction efficiency, optimal expression strategies during the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) infection cycle remain unexplored. This study investigates how VSV-G expression timing affects pseudotype incorporation into budded virions (BVs) and subsequent transduction efficacy. Methods: Three recombinant AcMNPV constructs were generated, each expressing VSV-G under distinct baculoviral promoters (ie1, gp64, and p10) and GFP via a CMV promoter. VSV-G incorporation was verified by Western blot, while transduction efficiency was quantified in mammalian cell lines (fluorescence microscopy/flow cytometry) and rat hind limbs. Viral productivity was assessed through production kinetics and plaque assays. Results: All the pseudotyped viruses showed significantly enhanced transduction capacity versus controls, strongly correlating with VSV-G incorporation levels. The p10 promoter drove the highest VSV-G expression and transduction efficiency. Crucially, BV production yields and infectivity remained unaffected by VSV-G expression timing. The in vivo results mirrored the cell culture findings, with p10-driven constructs showing greater GFP expression at low doses (104 virions). Conclusions: Strategic VSV-G expression via very late promoters (particularly p10) maximizes baculoviral transduction without compromising production yields. This study establishes a framework for optimizing pseudotyped BV systems, demonstrating that late-phase glycoprotein expression balances high mammalian transduction with preserved insect-cell productivity—a critical advancement for vaccine vector development. |
| format | info:ar-repo/semantics/artículo |
| id | INTA23052 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | MDPI |
| publisherStr | MDPI |
| record_format | dspace |
| spelling | INTA230522025-07-17T10:27:26Z Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells Simonin, Jorge Alejandro Cuccovia Warlet, Franco Uriel Bauzá, María del Rosario Plastine, María Del Pilar Alfonso, Victoria Olea, Fernanda Daniela Cerrudo, Carolina Susana Belaich, Mariano Nicolás Baculovirus Virions Cells Vesicular Stomatitis Virus Promoters Vaccines Virión Células Virus de la Estomatitis Vesicular Autographa californica Promotora Vacuna Background/Objectives: Baculoviruses represent promising gene delivery vectors for mammalian systems, combining high safety profiles with substantial cargo capacity. While pseudotyping with vesicular stomatitis virus G-protein (VSV-G) enhances transduction efficiency, optimal expression strategies during the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) infection cycle remain unexplored. This study investigates how VSV-G expression timing affects pseudotype incorporation into budded virions (BVs) and subsequent transduction efficacy. Methods: Three recombinant AcMNPV constructs were generated, each expressing VSV-G under distinct baculoviral promoters (ie1, gp64, and p10) and GFP via a CMV promoter. VSV-G incorporation was verified by Western blot, while transduction efficiency was quantified in mammalian cell lines (fluorescence microscopy/flow cytometry) and rat hind limbs. Viral productivity was assessed through production kinetics and plaque assays. Results: All the pseudotyped viruses showed significantly enhanced transduction capacity versus controls, strongly correlating with VSV-G incorporation levels. The p10 promoter drove the highest VSV-G expression and transduction efficiency. Crucially, BV production yields and infectivity remained unaffected by VSV-G expression timing. The in vivo results mirrored the cell culture findings, with p10-driven constructs showing greater GFP expression at low doses (104 virions). Conclusions: Strategic VSV-G expression via very late promoters (particularly p10) maximizes baculoviral transduction without compromising production yields. This study establishes a framework for optimizing pseudotyped BV systems, demonstrating that late-phase glycoprotein expression balances high mammalian transduction with preserved insect-cell productivity—a critical advancement for vaccine vector development. Instituto de Biotecnología Fil: Simonin, Jorge Alejandro. Universidad Nacional de Quilmes. Instituto de Microbiología Básica y Aplicada. Laboratorio de Ingeniería Genética y Biología Celular y Molecular (LIGBCM); Argentina Fil: Simonin, Jorge Alejandro. Comisión de Investigaciones Científicas de la Provincia de Buenos Aires (CIC); Argentina Fil: Cuccovia Warlet, Franco Uriel. Universidad Nacional de Quilmes. Instituto de Microbiología Básica y Aplicada. Laboratorio de Ingeniería Genética y Biología Celular y Molecular (LIGBCM); Argentina Fil: Cuccovia Warlet, Franco Uriel. Comisión de Investigaciones Científicas de la Provincia de Buenos Aires (CIC); Argentina Fil: Bauzá, María del Rosario. Universidad Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería (IMETTYB). Laboratorio de Medicina Regenerativa Cardiovascular; Argentina Fil: Bauzá, María del Rosario. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Plastine, María Del Pilar. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular (IABIMO); Argentina Fil: Plastine, María Del Pilar. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Alfonso, Victoria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular (IABIMO); Argentina Fil: Alfonso, Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Olea, Fernanda Daniela. Universidad Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería (IMETTYB). Laboratorio de Medicina Regenerativa Cardiovascular; Argentina Fil: Olea, Fernanda Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Cerrudo, Carolina Susana. Universidad Nacional de Quilmes. Instituto de Microbiología Básica y Aplicada. Laboratorio de Ingeniería Genética y Biología Celular y Molecular (LIGBCM); Argentina Fil: Cerrudo, Carolina Susana. Comisión de Investigaciones Científicas de la Provincia de Buenos Aires (CIC); Argentina Fil: Belaich, Mariano Nicolás. Universidad Nacional de Quilmes. Instituto de Microbiología Básica y Aplicada. Laboratorio de Ingeniería Genética y Biología Celular y Molecular (LIGBCM); Argentina Fil: Belaich, Mariano Nicolás. Comisión de Investigaciones Científicas de la Provincia de Buenos Aires (CIC); Argentina 2025-07-17T10:20:16Z 2025-07-17T10:20:16Z 2025-07 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion http://hdl.handle.net/20.500.12123/23052 https://www.mdpi.com/2076-393X/13/7/693 2076-393X https://doi.org/10.3390/vaccines13070693 eng info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-sa/4.0/ Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0) application/pdf MDPI Vaccines 13 (7) : 693 (July 2025) |
| spellingShingle | Baculovirus Virions Cells Vesicular Stomatitis Virus Promoters Vaccines Virión Células Virus de la Estomatitis Vesicular Autographa californica Promotora Vacuna Simonin, Jorge Alejandro Cuccovia Warlet, Franco Uriel Bauzá, María del Rosario Plastine, María Del Pilar Alfonso, Victoria Olea, Fernanda Daniela Cerrudo, Carolina Susana Belaich, Mariano Nicolás Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| title | Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| title_full | Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| title_fullStr | Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| title_full_unstemmed | Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| title_short | Early to late VSV-G expression in AcMNPV BV enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| title_sort | early to late vsv g expression in acmnpv bv enhances transduction in mammalian cells but does not affect virion yield in insect cells |
| topic | Baculovirus Virions Cells Vesicular Stomatitis Virus Promoters Vaccines Virión Células Virus de la Estomatitis Vesicular Autographa californica Promotora Vacuna |
| url | http://hdl.handle.net/20.500.12123/23052 https://www.mdpi.com/2076-393X/13/7/693 https://doi.org/10.3390/vaccines13070693 |
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