title: Synchrotron Infrared and Deep UV Fluorescent Microspectroscopy Study of PB1-F2 β-Aggregated Structures in Influenza A Virus-infected Cells.
abstract: PB1-F2 is a virulence factor of influenza A virus (IAV) whose functions remain misunderstood. The different roles of PB1-F2 may be linked to its structural polymorphism and to its propensity to assemble into oligomers and amyloid fibers in the vicinity of the membrane of IAV-infected cells. Here, we monitored the impact of PB1-F2 on the biochemical composition and protein structures of human epithelial pulmonary cells (A549) and monocytic cells (U937) upon IAV infection using synchrotron Fourier-transform infrared (FTIR) and deep UV (DUV) microscopies at the single-cell level. Cells were infected with a wild-type IAV and its PB1-F2 knock-out mutant for analyses at different times post-infection. IR spectra were recorded in each condition and processed to evaluate the change in the component band of the spectra corresponding to the amide I (secondary structure) and the CH stretching region (membrane). The IR spectra analysis revealed that expression of PB1-F2 in U937 cells, but not in A549 cells, results in the presence of a specific β-aggregate signature. Furthermore, the lipid membrane composition of U937 cells expressing PB1-F2 was also altered in a cell type-dependent manner. Using DUV microscopy and taking advantage of the high content of tryptophan residues in the sequence of PB1-F2 (5/90 aa), we showed that the increase of the autofluorescent signal recorded in monocytic cells could be correlated with the IR detection of β-aggregates. Altogether, our results constitute an important step forward in the understanding of the cell type-dependent function of PB1-F2.
authors.affiliation: From the VIM, UR892, INRA, Université PARIS-SACLAY, 78350 Jouy-en-Josas and christophe.chevalier@jouy.inra.fr.
authors.forename: Christophe
authors.initials: C
authors.lastname: Chevalier
authors.affiliation: From the VIM, UR892, INRA, Université PARIS-SACLAY, 78350 Jouy-en-Josas and.
authors.forename: Ronan
authors.initials: R
authors.lastname: Le Goffic
authors.affiliation: the DISCO and SMIS Beamlines, Synchrotron SOLEIL, L'Orme des Merisiers, 91190 Saint-Aubin, Gif-sur Yvette, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Jamme
authors.affiliation: From the VIM, UR892, INRA, Université PARIS-SACLAY, 78350 Jouy-en-Josas and.
authors.forename: Olivier
authors.initials: O
authors.lastname: Leymarie
authors.affiliation: the DISCO and SMIS Beamlines, Synchrotron SOLEIL, L'Orme des Merisiers, 91190 Saint-Aubin, Gif-sur Yvette, France.
authors.forename: Matthieu
authors.initials: M
authors.lastname: Réfrégiers
authors.affiliation: From the VIM, UR892, INRA, Université PARIS-SACLAY, 78350 Jouy-en-Josas and.
authors.forename: Bernard
authors.initials: B
authors.lastname: Delmas
mesh_terms: D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism / Q000821:virology; D006367:HeLa Cells; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity*; D007251:Influenza, Human / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D066329:Protein Aggregates; D020298:U937 Cells; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: C442816:PB1-F2 protein, Influenza A virus; D066329:Protein Aggregates; D014764:Viral Proteins
keywords: Fourier transform IR (FTIR); amyloid; fluorescence imaging; influenza virus; membrane; microscopy; oligomerization; synchrotron radiation experiment