Biology subjects

Lozach, P.-Y.

Publications and source records attributed to Lozach, P.-Y..

4 recordsLinked to original sources

bioRxiv2023

A high-content RNA-based imaging assay reveals integrin beta 1 as a cofactor for cell entry of non-enveloped hepatitis E virus

Hepatitis E virus (HEV) is a major cause of acute hepatitis and mainly transmitted faecal-orally. HEV particles in faeces are non-enveloped, while those in the blood possess a cell-derived lipid envelope. Despite being a global health concern, there is limited understanding of the steps in the HEV life cycle, particularly cell entry. A previous study proposed integrin alpha 3 (ITGA3) as a potential host factor for nHEV entry, but the {beta}-integrin partner that co-mediates HEV entry has not been described. To address this knowledge gap and resolve the existing controversies surrounding HEV cell entry, we developed an RNA-FISH-based high-content imaging assay alllowing investigation of the entry pathways of both naked and enveloped HEV particles. Our observations indicate that naked HEV particles interact with the surface receptor integrin beta 1 (ITGB1), which likely facilitates their trafficking through the recycling endosome. In contrast, enveloped HEV particles do not interact with ITGB1 and instead use the classical endocytic pathway via the early endosome. Importantly, both forms of HEV require endosomal acidification and proteolytic cleavage by lysosomal cathepsins, which ultimately results in delivery of the HEV genome to the cytoplasm.

microbiology↗

bioRxiv2023

The phenuivirus Toscana virus makes an atypical use of vacuolar acidity to enter host cells

Toscana virus is a major cause of arboviral disease in humans in the Mediterranean basin during summer. However, early virus-host cell interactions and entry mechanisms remain poorly characterized. Investigating iPSC-derived human neurons and cell lines, we found that virus binding to the cell surface was specific but inefficient, and 50% of bound virions were endocytosed within 10 min. Virions entered Rab5a+ early endosomes and, subsequently, Rab7a+ and LAMP-1+ late endosomal compartments. Penetration required intact late endosomes and occurred within 30 min following internalization. Virus entry relied on vacuolar acidification, with an optimal pH for viral membrane fusion at pH 5.5. The pH threshold increased to 5.8 with longer pre-exposure of virions to the slightly acidic pH in early endosomes. Strikingly, the particles remained infectious after entering late endosomes with a pH below the fusion threshold. Overall, our study establishes Toscana virus as a late-penetrating virus and reveals an atypical use of vacuolar acidity by this virus to enter host cells.

microbiology↗

bioRxiv2022

The glycolipid GlcCer is recruited into the viral envelope to promote phenuivirus binding to host cells

Virus-receptor interactions largely contribute to the tropism and outcome of an infection. Here, we found that the glycolipid glucosylceramide (GlcCer) is a major component of Uukuniemi phenuivirus and allows viral binding to host cells. A lipidomic analysis with mass spectrometry revealed the lipidome of UUKV particles and indicated that GlcCer was enriched in both infected cells and viral particles. In addition, the infectivity of UUKV depended on the conversion of ceramide (Cer) into GlcCer in the Golgi network of producer cells. In contrast, depletion of GlcCer in virions profoundly impaired the attachment of UUKV and other related viruses to target cells. Furthermore, competing GlcCer ligands prevented virus binding to various cell types. Altogether, our results demonstrate that glycolipids are essential structural determinants of the virions necessary for virus attachment to host cells and have strong implications for future work on the identification of virus receptors.

microbiology↗

bioRxiv2020

Host Cell Proteases Drive Early or Late SARS-CoV-2 Penetration

SARS-CoV-2 is a newly emerged coronavirus (CoV) that spread through human populations worldwide in early 2020. CoVs rely on host cell proteases for activation and infection. The trypsin-like protease TMPRSS2 at the cell surface, cathepsin L in endolysosomes, and furin in the Golgi have all been implicated in the SARS-CoV-2 proteolytic processing. Whether SARS-CoV-2 depends on endocytosis internalization and vacuolar acidification for infectious entry remains unclear. Here, we examined the dynamics of SARS-CoV-2 activation during the cell entry process in tissue culture. Using four cell lines representative of lung, colon, and kidney epithelial tissues, we found that TMPRSS2 determines the SARS-CoV-2 entry pathways. In TMPRSS2-positive cells, infection was sensitive to aprotinin, a TMPRSS2 inhibitor, but not to SB412515, a drug that impairs cathepsin L. Infectious penetration was marginally dependent on endosomal acidification, and the virus passed the protease-sensitive step within 10 min. In a marked contrast, in TMPRSS2-negative cells cathepsin L and low pH were required for SARS-CoV-2 entry. The cathepsin L-activated penetration occurred within 40-60 min after internalization and required intact endolysosomal functions. Importantly, pre-activation of the virus allowed it to bypass the need for endosomal acidification for viral fusion and productive entry. Overall, our results indicate that SARS-CoV-2 shares with other CoVs a strategy of differential use of host cell proteases for activation and infectious penetration. This study also highlights the importance of TMPRSS2 in dictating the entry pathway used by SARS-CoV-2. SignificancePreventing SARS-CoV-2 spread requires approaches affecting early virus-host cell interactions before the virus enters and infects target cells. Host cell proteases are critical for coronavirus activation and infectious entry. Here, we reconcile apparent contradictory observations from recent reports on endosomal acidification and the role of furin, TMPRSS2, and cathepsin L in the productive entry and fusion process of SARS-CoV-2. Investigating authentic virus in various cell types, we demonstrated that SARS-CoV-2 developed the ability to use different entry pathways, depending on the proteases expressed by the target cell. Our results have strong implications for future research on the apparent broad tropism of the virus in vivo. This study also provides a handle to develop novel antiviral strategies aiming to block virus entry, as illustrated with the several drugs that we identified to prevent SARS-CoV-2 infection, some with low IC50.

microbiology↗