Baculovirus as a highly efficient gene delivery vector for the expression of hepatitis delta virus antigens in mammalian cells

Abstract

Baculovirus has been employed for a wide variety of applications. In this study, we further expanded the application to the high-level expression of hepatitis delta virus (HDV) antigens and the formation of virus-like particles (VLP) in transduced mammalian cells. To this end, two recombinant baculoviruses were constructed to express large hepatitis delta antigen (L-HDAg) and hepatitis B surface antigen (HBsAg) under mammalian promoters. With a simplified transduction protocol using unconcentrated virus, high transduction efficiencies were achieved in hepatoma cells, in which L-HDAg and HBsAg were expressed abundantly, allowing for easy colorimetric detection in Western blots. L-HDAg alone was nucleus-bound and HBsAg alone was secreted; formation and secretion of HDV-like particles were readily detected upon coexpression, indicating that the baculovirus-expressed proteins were processed correctly as the authentic proteins. Quantitative real-time PCR (Q-PCR) analyses quantitatively revealed that baculovirus transduction was more efficient than plasmid transfection with respect to DNA uptake and DNA transport to the nucleus. Furthermore, superinfection introduced more baculovirus DNA into cells in the long-term culture as revealed by Q-PCR, thereby enhancing and prolonging the expression. In summary, baculovirus transduction can be an attractive method as an alternative to the plasmid transfection commonly employed for HDV research thanks to the significantly higher gene delivery efficiencies as well as the abundant expression and proper processing. Baculovirus can also be envisaged as a useful tool for investigating protein-cell interactions and virus assembly.

Baculovirus‐mediated gene transfer is attenuated by sodium bicarbonate

“…Thanks to the nonreplicating nature and low cytotoxicity inside mammalian cells, the large genome (≈130 kb) capable of harboring multiple genes or large inserts [4], and ease of recombinant virus construction and production, increasing efforts have been directed towards employing baculovirus for in vitro and in vivo gene delivery [5][6][7]. Further, baculovirus-mediated gene delivery into mammalian cells has been exploited for the development of cell-based assays [8], delivery of small interfering RNA [9,10], surface display of eucaryotic proteins [11], and high-level production of virus-like particles [12].…”

“…To enhance the gene delivery efficiency, we have previously developed a protocol by which virus transduction is performed by incubating the cells with unconcentrated virus supernatant for 4-8 h at 25 • C or 27 • C using Ca 2+ -and Mg 2+ -free Dulbecco's PBS as the surrounding solution to adjust the final liquid volume [16][17][18]. This protocol has been employed to effectively enhance baculovirusmediated gene delivery into HepG2, HeLa and BHK cells [12,19,20].…”

A Cholesterol Recognition Amino Acid Consensus Domain in GP64 Fusion Protein Facilitates Anchoring of Baculovirus to Mammalian Cells

“…Unlike most viruses studied so far, most interestingly, AcMNPV can enter a wide variety of cells from different organisms and drive the expression of foreign genes under the control of mammalian promoters (7,8,9).…”

Baculovirus as a new gene delivery vector for stem cell engineering and bone tissue engineering

“…In contrast to these vectors, baculovirus neither replicates nor is toxic inside the transduced cells. 15 Baculoviral DNA degrades in the cells over time 36,37 and no evidence currently exists that baculoviral DNA integrates into host chromosomes in the absence of selective pressure. 38 These attributes reduce the potential side effects and ease the safety concerns.…”