Structural analysis of influenza protein-membrane interactions during entry

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2nd funding period (2018 – 06/2022)

Project description – Summary – 3rd funding period

We aim to structurally elucidate influenza A virus (IAV) entry and the interplay of viral proteins with host cellular factors during IAV cell-to-cell spread and entry. In the previous funding period, we established a cryo-focused ion beam milling and cryo-correlative light and electron microscopy (cryo-CLEM) workflow, which enables us to study organelles such as pulmonary lamellar bodies and virus-infected cells in close to native conditions and at molecular resolution (Klein S, Wimmer, B et al, Commun Biol. 2021).

We applied it to study IAV entry in collaboration with Brügger/Kräusslich and Schwarz/Lanzer. Moreover, we also applied our workflow to unravel the steps of SARS-CoV-2 assembly and replication in infected cells in collaboration with Bartenschlager/Höfer and Boulant (Klein S, Cortese M, Winter S, et al, Nat Commun, 2020).

In the current funding period, we continue to investigate how IFITM3, one of the major IAV entry inhibitors increases the energy barrier which needs to be overcome by IAV hemagglutinins to form a membrane fusion pore. We will further apply our cryo-CLEM workflow to structurally characterize IAV post-fusion entry steps focusing on the role of the viral M1 protein in viral cell-to-cell spread and entry. In addition, we will investigate the role of virion morphology determined by M1 protein in IAV cell-to-cell spread using a lung-mimicking environment. IAV is a pleiomorphic virus forming long filamentous virions, which are often found in human isolates, whereas spherical virions are typical for lab-adapted strains. To this end, we will establish a novel correlative light and scanning electron microscopy workflow to quantitatively assess the spread of IAV with spherical and filamentous morphology in collaboration with Funaya/Schwab.

In collaborative efforts with Dao Thi, we will use cryo-ET to provide a further understanding of the cell-to-cell spread of naked and quasi-enveloped hepatitis E virions (HEV). In addition, we will apply our cryo-CLEM workflow to continue our established collaboration with Bartenschlager/Höfer to study the replication of hepatitis C virus (HCV) in infected cells.

Project staff

Steffen Klein
PhD student