SARS-CoV-2 requires cholesterol for viral entry and pathological syncytia formation
- Princeton University, United States
- Boston University, United States
- University of Virginia, United States
- Washington University School of Medicine, United States
- Harvard Medical School, United States
Abstract
Many enveloped viruses induce multinucleated cells (syncytia), reflective of membrane fusion events caused by the same machinery that underlies viral entry. These syncytia are thought to facilitate replication and evasion of the host immune response. Here, we report that co-culture of human cells expressing the receptor ACE2 with cells expressing SARS-CoV-2 spike, results in synapse-like intercellular contacts that initiate cell-cell fusion, producing syncytia resembling those we identify in lungs of COVID-19 patients. To assess the mechanism of spike/ACE2-driven membrane fusion, we developed a microscopy-based, cell-cell fusion assay to screen ~6000 drugs and >30 spike variants. Together with quantitative cell biology approaches, the screen reveals an essential role for biophysical aspects of the membrane, particularly cholesterol-rich regions, in spike-mediated fusion, which extends to replication-competent SARS-CoV-2 isolates. Our findings potentially provide a molecular basis for positive outcomes reported in COVID-19 patients taking statins, and suggest new strategies for therapeutics targeting the membrane of SARS-CoV-2 and other fusogenic viruses.
Data availability
All data generated or analyzed during this study are included in the manuscript and supporting files with the exception of raw imaging data (>400,000 Nikon ND2 files), which is not feasible to post online given its massive size (>1.5 TB). This data is available from the lead contact upon request, assuming the interested party provides a server with sufficient storage capacity. Raw data (computed fusion scores) from the drug repurposing screen is available in Supplemental File 1; bioinformatics, Supplemental File 3.
Article and author information
Author details
Clifford P Brangwynne
Funding
National Institute of General Medical Sciences (GM095467)
- Bruce D Levy
National Heart, Lung, and Blood Institute (HL122531)
- Bruce D Levy
National Institute of General Medical Sciences (GM134949)
- Ilya Levental
National Institute of General Medical Sciences (GM124072)
- Ilya Levental
Howard Hughes Medical Institute
- Clifford P Brangwynne
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Human subjects: Human pathology studies were performed with the approval of the Institutional Review Board at Brigham and Women's Hospital. Clinical autopsies with full anatomic dissection were performed on SARS-CoV-2 decedents by a board-certified anatomic pathologist (RFP) with appropriateinfectious precautions.
Copyright
© 2021, Sanders et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 18,882
- views
-
- 1,738
- downloads
-
- 172
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
SARS-CoV-2 requires cholesterol for viral entry and pathological syncytia formation
eLife 10:e65962.
https://doi.org/10.7554/eLife.65962
Further reading
-
- Cell Biology
- Neuroscience
The claustrum complex is viewed as fundamental for higher-order cognition; however, the circuit organization and function of its neuroanatomical subregions are not well understood. We demonstrated that some of the key roles of the CLA complex can be attributed to the connectivity and function of a small group of neurons in its ventral subregion, the endopiriform (EN). We identified a subpopulation of EN neurons by their projection to the ventral CA1 (ENvCA1-proj. neurons), embedded in recurrent circuits with other EN neurons and the piriform cortex. Although the ENvCA1-proj. neuron activity was biased toward novelty across stimulus categories, their chemogenetic inhibition selectively disrupted the memory-guided but not innate responses of mice to novelty. Based on our functional connectivity analysis, we suggest that ENvCA1-proj. neurons serve as an essential node for recognition memory through recurrent circuits mediating sustained attention to novelty, and through feed-forward inhibition of distal vCA1 neurons shifting memory-guided behavior from familiarity to novelty.
-
- Cell Biology
- Computational and Systems Biology
Induced pluripotent stem cell (iPSC) technology is revolutionizing cell biology. However, the variability between individual iPSC lines and the lack of efficient technology to comprehensively characterize iPSC-derived cell types hinder its adoption in routine preclinical screening settings. To facilitate the validation of iPSC-derived cell culture composition, we have implemented an imaging assay based on cell painting and convolutional neural networks to recognize cell types in dense and mixed cultures with high fidelity. We have benchmarked our approach using pure and mixed cultures of neuroblastoma and astrocytoma cell lines and attained a classification accuracy above 96%. Through iterative data erosion, we found that inputs containing the nuclear region of interest and its close environment, allow achieving equally high classification accuracy as inputs containing the whole cell for semi-confluent cultures and preserved prediction accuracy even in very dense cultures. We then applied this regionally restricted cell profiling approach to evaluate the differentiation status of iPSC-derived neural cultures, by determining the ratio of postmitotic neurons and neural progenitors. We found that the cell-based prediction significantly outperformed an approach in which the population-level time in culture was used as a classification criterion (96% vs 86%, respectively). In mixed iPSC-derived neuronal cultures, microglia could be unequivocally discriminated from neurons, regardless of their reactivity state, and a tiered strategy allowed for further distinguishing activated from non-activated cell states, albeit with lower accuracy. Thus, morphological single-cell profiling provides a means to quantify cell composition in complex mixed neural cultures and holds promise for use in the quality control of iPSC-derived cell culture models.
