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Investigation of mumps and measles virus proteome and lipidome

Measles (MEV) and mumps (MUV) virus are enveloped, nonsegmented, 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. Lipid bilayer of enveloped viruses is not directly encoded by the virus but acquired from the host cell membranes. Lipid composition of some viruses was found to resemble that of host cell plasma membrane, but in other cases significant differences were noticed [2, 3]. Apart from virus-coded proteins, it has been established that various enveloped viruses incorporate also host cell proteins (HCPs) both inside the viral particle and in the lipid bilayer. Indications exist that some HCPs are specifically taken up by the viral particles by direct interactions with viral proteins, but HCPs might also be included by non-specific interactions, or as contamination arising from the presence of exosomes in virus preparations [4-6]. The aim of our research was investigation of MEV and MUV lipidome and proteome, consisting of 8 and 9 virus-coded proteins, respectively, and detection of HCPs potentially present in virus preparations. Viruses were grown in Vero cells, microfiltrated, purified by hydrophobic interaction chromatography (HIC) and eluates concentrated by ultracentrifugation. Alternatively, viruses were purified only by ultracentrifugation. Also, supernatants from non-infected Vero cells were purified and analyzed as described for virus samples. Obtained samples were resolved by SDS-PAGE, protein bands excised and in-gel digestion by trypsin was performed. Lipids were extracted from samples by Bligh-Dyer method and separated using 2D-HPTLC. Lipids and proteins were identified by MALDI-TOF/TOF-MS. Results implicate that there are no considerable differences between lipid composition of viral particles and Vero cells. All viral structural proteins were detected in viral samples, including fusion protein both for MUV and MEV which was not previously found. Also, various HCPs previously reported in other purified virus preparations such as actin, cyclophilin A, and annexin A2 were detected. References [1] C.L. Afonso, G.K. Amarasinghe, K. Banyai, Y. Bao et al., Arch. Virol. 161 (2016) 2351- 2360. [2] M. Brgles, M. Bonta, M. Šantak, M. Jagušić, D. Forčić, B. Halassy, G. Allmaier and M. Marchetti-Deschmann, Virology Journal 13 (2016) DOI:10.1186/s12985-016-0463-0. [3] B. Brügger, B. Glass, P. Haberkant, I. Leibrecht, F.T. Wieland and H.-G. Kräusslich, Proc. Natl. Acad. Sci. U.S.A. 103 (2006) 2641- 2646. [4] R. Cantin, S. Méthot and M.J. Tremblay, J. Virol. 79 (2005) 6577-6587. [5] E.K. Franke, H.E. Yuan and J. Luban, Nature 372 (1994) 359-362. [6] E. Nolte-‘t Hoen, T. Cremer, R.C. Gallo and L.B. Margolis, Proc. Natl. Acad. Sci. U.S.A. 113 (2016) 9155-9161.

mumps virus, measles virus, mass spectrometry, MALDI-TOF/TOF

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