Dataset Information

SARS-CoV-2 hijacks macropinocytosis to facilitate its entry and promote viral spike-mediated cell-to-cell fusion.

ABSTRACT: Revealing the mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry and cell-to-cell spread might provide insights for understanding the underlying mechanisms of viral pathogenesis, tropism, and virulence. The signaling pathways involved in SARS-CoV-2 entry and viral spike-mediated cell-to-cell fusion remain elusive. In the current study, we found that macropinocytosis inhibitors significantly suppressed SARS-CoV-2 infection at both the entry and viral spike-mediated cell-to-cell fusion steps. We demonstrated that SARS-CoV-2 entry required the small GTPase Rac1 and its effector kinase p21-activated kinase 1 by dominant-negative and RNAi assays in human embryonic kidney 293T-angiotensin-converting enzyme 2 cells and that the serine protease transmembrane serine protease 2 reversed the decrease in SARS-CoV-2 entry caused by the macropinocytosis inhibitors. Moreover, in the cell-to-cell fusion assay, we confirmed that macropinocytosis inhibitors significantly decreased viral spike-mediated cell-to-cell fusion. Overall, we provided evidence that SARS-CoV-2 utilizes a macropinocytosis pathway to enter target cells and to efficiently promote viral spike-mediated cell-to-cell fusion.

SUBMITTER: Zhang YY

PROVIDER: S-EPMC9484108 | biostudies-literature | 2022 Nov

REPOSITORIES: biostudies-literature

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SARS-CoV-2 hijacks macropinocytosis to facilitate its entry and promote viral spike-mediated cell-to-cell fusion.

Zhang Yu-Yuan YY Liang Ronghui R Wang Shu-Jie SJ Ye Zi-Wei ZW Wang Tong-Yun TY Chen Meng M Liu Jianbo J Na Lei L Yang Yue-Lin YL Yang Yong-Bo YB Yuan Shuofeng S Yin Xin X Cai Xue-Hui XH Tang Yan-Dong YD

The Journal of biological chemistry 20220919 11

Revealing the mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry and cell-to-cell spread might provide insights for understanding the underlying mechanisms of viral pathogenesis, tropism, and virulence. The signaling pathways involved in SARS-CoV-2 entry and viral spike-mediated cell-to-cell fusion remain elusive. In the current study, we found that macropinocytosis inhibitors significantly suppressed SARS-CoV-2 infection at both the entry and viral spike-mediated c ...[more]

PMID: 36259516

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Project description:Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike is the fusion machine for host cell entry. Still, the mechanism by which spike protein interacts with the target lipid membrane to facilitate membrane fusion during entry is not fully understood. Here, using steady-state membrane fusion and single-molecule fluorescence resonance energy transfer imaging of spike trimers on the surface of SARS-CoV-2 pseudovirion, we directly show that spike protein interacts with phosphatidylserine (PS) lipid in the target membrane for mediating fusion. We observed that the fusion peptide of the spike S2 domain interacts with the PS lipid of the target membrane. Low pH and Ca2+ trigger the spike conformational change and bring fusion peptide in close proximity to the PS lipid of the membrane. The binding of the spike with PS lipid of its viral membrane (cis interaction) impedes the fusion activation. PS on the target membrane promotes spike binding via trans interaction, prevents the cis interaction, and accelerates fusion. Sequestering or absence of PS lipid abrogates the spike-mediated fusion process and restricts SARS-CoV-2 infectivity. We found that PS-dependent interaction for fusion is conserved across all the SARS-CoV-2 spike variants of concern (D614G, Alpha, Beta, Delta, and Omicron). Our study suggests that PS lipid is indispensable for SARS-CoV-2 spike-mediated virus and target membrane fusion for entry, and restricting PS interaction with spike inhibits the SARS-CoV-2 spike-mediated entry. Therefore, PS is an important cofactor and acts as a molecular beacon in the target membrane for SARS-CoV-2 entry.ImportanceThe role of lipids in the host cell target membrane for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry is not clear. We do not know whether SARS-CoV-2 spike protein has any specificity in terms of lipid for membrane fusion reaction. Here, using in vitro reconstitution of membrane fusion assay and single-molecule fluorescence resonance energy transfer imaging of SARS-CoV-2 spike trimers on the surface of the virion, we have demonstrated that phosphatidylserine (PS) lipid plays a key role in SARS-CoV-2 spike-mediated membrane fusion reaction for entry. Membrane-externalized PS lipid strongly promotes spike-mediated membrane fusion and COVID-19 infection. Blocking externalized PS lipid with PS-binding protein or in the absence of PS, SARS-CoV-2 spike-mediated fusion is strongly inhibited. Therefore, PS is an important target for restricting viral entry and intervening spike, and PS interaction presents new targets for COVID-19 interventions.

Dataset Information

SARS-CoV-2 hijacks macropinocytosis to facilitate its entry and promote viral spike-mediated cell-to-cell fusion.

Publications

SARS-CoV-2 hijacks macropinocytosis to facilitate its entry and promote viral spike-mediated cell-to-cell fusion.

Similar Datasets

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