RNA seq analysis comparing uninfected to MCPyV-infected 2D monolayer or 3D spheroids

However, studies on the life cycle of small DNA tumor viruses have been hampered primarily due to the lack of virus culture systems that allow for robust replication and high-level infection in host cells. Thus, studies involving Merkel cell polyomavirus (MCPyV), a human oncogenic polyomavirus, infections primarily rely on utilizing quasiviruses, virions produced through the artificial complementation of the viral genome with exogenous MCPyV proteins.

However, recombinant quasiviruses might not fully reflect the native MCPyV virions or infection conditions. To mimic the natural skin infection of MCPyV, we developed a simple and efficient method for producing MCPyV virions derived from primary human dermal fibroblasts without the requirement for exogenous MCPyV protein supplementation using a 3D spheroid culture system. Here, we found that the 3D spheroid microenvironment enabled the production of MCPyV virions and their release into the supernatant that were packaged inside of extracellular vesicles (EVs) from natural host cells.