0000000000125427
AUTHOR
Anssi J. Mähönen
Enhanced Gene Delivery by Avidin-Displaying Baculovirus
Flexible alteration of virus surface properties would be beneficial for enhanced and targeted gene delivery. A useful approach could be based on a high-affinity receptor–ligand pair, such as avidin and biotin. In this study, we have constructed an avidin-displaying baculovirus, Baavi. Avidin display was expected to enhance cell transduction due to the high positive charge of avidin in physiological pH and to provide a binding site for covering the virus with desired biotinylated ligands. Successful incorporation of avidin on the virus envelope was detected by immunoblotting and electron microscopy. Multiple biotin-binding sites per virus were detected with fluorescence-correlation spectrosc…
Culture medium induced vimentin reorganization associates with enhanced baculovirus-mediated gene delivery.
Baculoviruses can express transgenes under mammalian promoters in a wide range of vertebrate cells. However, the success of transgene expression is dependent on both the appropriate cell type and culture conditions. We studied the mechanism behind the substantial effect of the cell culture medium on efficiency of the baculovirus transduction in different cell lines. We tested six cell culture mediums; the highest transduction efficiency was detected in the presence of RPMI 1640 medium. Vimentin, a major component of type III intermediate filaments, was reorganized in the optimized medium, which associated with enhanced nuclear entry of baculoviruses. Accordingly, the phosphorylation pattern…
Baculovirus capsid display: a novel tool for transduction imaging
Baculoviruses are enveloped insect viruses that can carry large quantities of foreign DNA in their genome. Baculoviruses have proved to be very promising gene therapy vectors but little is known about their transduction mechanisms in mammalian cells. We show in this study that Autographa californica multiple nuclear polyhedrosis virus capsid is compatible with the incorporation of desired proteins in large quantities. Fusions can be made to the N-terminus or C-terminus of the major capsid protein vp39 without compromising the viral titer or functionality. As an example of the baculovirus capsid display we show a tracking of the baculovirus transduction in mammalian cells by an enhanced gree…
Coxsackievirus B3 VLPs purified by ion exchange chromatography elicit strong immune responses in mice
Coxsackievirus B3 (CVB3) is an important cause of acute and chronic viral myocarditis, and dilated cardiomyopathy (DCM). Although vaccination against CVB3 could significantly reduce the incidence of serious or fatal viral myocarditis and various other diseases associated with CVB3 infection, there is currently no vaccine or therapeutic reagent in clinical use. In this study, we contributed towards the development of a CVB3 vaccine by establishing an efficient and scalable ion exchange chromatography-based purification method for CVB3 virus and baculovirus-insect cell-expressed CVB3 virus-like particles (VLPs). This purification system is especially relevant for vaccine development and produ…
Generation of lentivirus vectors using recombinant baculoviruses
In spite of advances in conventional four-plasmid transient transfection methods and development of inducible stable production cell lines, production of replication-defective lentiviral vectors in clinical scale has been challenging. Baculovirus technology offers an alternative to scalable virus production as a result of fast and easy production of baculoviruses, efficient transduction of mammalian cells and safety of the baculoviruses. As a first step toward scalable lentiviral production system, we have constructed four recombinant baculoviruses: the BAC-transfer virus expresses green fluorescent protein (GFP) as a transgene and BAC-gag-pol, BAC-vesicular stomatitis virus glycoprotein G …