Quantitative proteomic analysis of extracellular vesicles in response to baculovirus infection of a Trichoplusia ni cell line

Abstract

Due to its outstanding suitability to produce complex biopharmaceutical products including virus-like particles and subunit vaccines, the baculovirus/insect cell expression system has developed into a highly popular production platform in the biotechnological industry. For high productivity, virus-cell communication and an efficient spreading of the viral infection are crucial, and, in this context, extracellular vesicles (EVs) might play a significant role. EVs are small particles, utilized by cells to transfer biologically active compounds such as proteins, lipids as well as nucleic acids to recipient cells for intracellular communication. Studies in mammalian cells showed that the release of EVs is altered in response to infection with many viruses, ultimately either limiting or fostering infection spreading. In this study we isolated and characterized EVs, from both uninfected and baculovirus infected Tnms42 insect cells. Via quantitative proteomic analysis we identified more than 3000 T. ni proteins in Tnms42 cell derived EVs, of which more than 400 were significantly differentially abundant upon baculovirus infection. Subsequent gene set enrichment analysis revealed a depletion of proteins related to the extracellular matrix in EVs from infected cultures. Our findings show a significant change of EV protein cargo upon baculovirus infection, suggesting a major role of EVs as stress markers. Our study might serve in designing new tools for process monitoring and control to further improve biopharmaceutical production within the baculovirus/insect cell expression system.

Production of norovirus-, rotavirus-, and enterovirus-like particles in insect cells is simplified by plasmid-based expression

“…In all cases the VLP purification was simpler and no contamination by baculoviral or alphanodaviral particles is expected 18 nor was observed in TEM, DLS or PAGE. Even contamination by extracellular vesicles is reported to be lower in transfected Trichoplusia ni insect cells, as compared to baculovirus-infected cells 52 . In this work, we did not compare production of enveloped VLPs, whose expression with the plasmid-based insect cell expression system has been validated 22 .…”

Characterisation of extracellular vesicles in baculovirus infection of Spodoptera frugiperda cells

“…While EV secretion by S. frugiperda cells had been described previously, no characterisation of these EVs has yet been reported (Hashimoto et al., 2017 ; Ishikawa et al., 2020 ; Puente‐Massaguer et al., 2022 ). Small EVs released by Trichoplusia ni cells during baculovirus infection were recently analysed by mass spectrometry, yet viral proteins were not analysed due to possible contamination by virions (Hausjell et al., 2023 ). To characterise sEVs of AcMNPV‐infected cells, these vesicles had to be separated from the BV.…”

“…Though the presence of EVs has been described in S. frugiperda cells, little research has been performed on the role they may play in baculovirus infection (Hashimoto et al., 2017 ; Hodgson et al., 2022 ; Ishikawa et al., 2020 ; Puente‐Massaguer et al., 2022 ). Recently, sEVs were characterised by mass spectrometry in AcMNPV‐infected T. ni cells, although the presence of viral proteins was not evaluated since no full separation of sEVs from BV was achieved (Hausjell et al., 2023 ). Here, sEVs were characterised from both uninfected and baculovirus‐infected S. frugiperda cells.…”

Increased efficacy of influenza virus vaccine candidate through display of recombinant neuraminidase on virus like particles

“…For particle visualization, samples were prepared as described in Hausjell et al. (2023) ( 33 ). Briefly, formvar-coated copper grids were floated on VLP suspension diluted 1:50 in particle-free water.…”