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Mass spectrometry-based investigation of mumps and measles virus proteome

Introduction Mumps (MUV) and measles (MEV) virus are enveloped, non-segmented, negative single- stranded RNA viruses of the family Paramyxoviridae [1]. They are the cause of mumps and measles, respectively, both of which can be prevented by vaccination. Aside from proteins coded by viral genome, viruses also incorporate host cell proteins (HCPs), through specific or non-specific interactions [2]. The presence of extracellular vesicles (EVs) which are co-purified with viruses due to their similarity in size and density also contributes to EVs in virus preparations and this has often been overlooked [3]. Methods MUV, MEV and EVs were prepared in Vero cells, microfiltrated and purified by one-step ultracentrifugation (UC) 2 h at 141, 000 × g, hydrophobic interaction chromatography (HIC) [4] or immunoaffinity chromatography (IAC) on anti-MUV column [5]. Samples obtained by chromatography were additionally concentrated by UC prior to SDS- PAGE under reducing conditions. Protein bands were excised, subjected to in-gel digestion by trypsin, and identified by MALDI-TOF/TOF-MS (peptide mass fingerprint and peptide sequencing). Results Almost all virus coded proteins were detected in MUV and MEV by the applied proteomic approach, but the purification procedure affected which of them were detected. All HCPs present in virus preparations were also detected in EVs, but the comparison of HCPs present in EVs and virus preparations obtained by different purification procedures implies that actin, annexins, cyclophilin A, moesin and integrin β1 are present inside virions, while other HCPs are more likely to be contaminants, most likely arising from co-purified EVs. Innovative aspects • Comparative analysis of HCPs in viruses and EVs was carried out to differentiate between contaminating HCPs and those present inside the virions References [1] R.A. Lamb, G.D. Parks, Paramyxoviridae, in: D.M. Knipe, P.M. Howley (Eds.), Fields Virology, 6th ed., Lippincot Williams & Wilkins, Philadelphia, 2013: pp. 957–995. [2] R. Cantin, S. Méthot, M.J. Tremblay, Plunder and Stowaways : Incorporation of cellular proteins by enveloped viruses, J. Virol. 79 (2005) 6577–6587. [3] E. Nolte-’t Hoen, T. Cremer, R.C. Gallo, L.B. Margolis, Extracellular vesicles and viruses: Are they close relatives?, Proc. Natl. Acad. Sci. U. S. A. 113 (2016) 9155–61. [4] D. Sviben, D. Forcic, J. Ivancic-Jelecki, B. Halassy, M. Brgles, Recovery of infective virus particles in ion-exchange and hydrophobic interaction chromatography is influenced by particle charge and total-to- infective particle ratio, J. Chromatogr. B Anal. Technol. Biomed. Life Sci. 1054 (2017) 10–19. [5] M. Brgles, D. Sviben, D. Forcic, B. Halassy, Nonspecific native elution of proteins and mumps virus in immunoaffinity chromatography, J. Chromatogr. A. 1447 (2016) 107–114.

measles virus, mumps virus, mass spectrometry, proteome, host cell proteins, extracellular vesicles

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