Optimization of human papillomavirus-based pseudovirus techniques for efficient gene transfer

Abstract

Human papillomavirus (HPV) L1 and L2 capsid proteins self-assemble into virions capable of efficiently packaging either its 8 kilobase genome or non-viral DNA. The ability of HPV capsids to package non-viral DNA makes these a useful tool for delivering plasmids to study proteins of interest in a variety of cell types. We describe optimization of current methods and present new protocols for using HPV capsids to deliver non-viral DNA thereby providing an alternative to DNA transfection. Using keratinocyte generated extracellular matrices can enhance infection efficiency in keratinocytes, hepatocytes and neuronal cells. Furthermore, we describe a suspension-based efficient technique for infecting different cell types.

Cells infected with human papilloma pseudovirus display nuclear reorganization and heterogenous infection kinetics

“…We found a lower infection efficiency in HaCaT cells, but with mitosis being a prime requirement for HPV infection, we attribute this difference to their longer population doubling time [6, 7]. However, as pre‐binding HPV PsV with extracellular matrix (ECM) has been shown to significantly increase the infection efficiency in HaCaT cells, proliferation may not the only determinant [20].…”

“…To resolve this, recombinant viral particles such as pseudoviruses (PsV) have been conceived that are highly immunogenic and have an identical capsid composition as the authentic wild type HPV but lack the replicative part of the vDNA [17][18][19]. HPV PsV are primarily used for analyzing neutralization or infectious entry pathways, but can also be used as gene delivery vehicle [19,20]. Here, we have exploited HPV PsV harboring a fluorescent reporter (EGFP) to monitor early infection kinetics in human cells.…”