Genome-wide, bidirectional CRISPR screens identify mucins as critical host factors modulating SARS-CoV-2 infection

Abstract

SUMMARYSARS-CoV-2 can cause a range of symptoms in infected individuals, from mild respiratory illness to acute respiratory distress syndrome. A systematic understanding of the host factors mediating viral infection or restriction is critical to elucidate SARS-CoV-2 host-pathogen interactions and the progression of COVID-19. To this end, we conducted genome-wide CRISPR knockout and activation screens in human lung epithelial cells with endogenous expression of the SARS-CoV-2 entry factors ACE2 and TMPRSS2. These screens uncovered proviral and antiviral host factors across highly interconnected host pathways, including components implicated in clathrin transport, inflammatory signaling, cell cycle regulation, and transcriptional and epigenetic regulation. We further identified mucins, a family of high-molecular weight glycoproteins, as a prominent viral restriction network. We demonstrate that multiple membrane-anchored mucins are critical inhibitors of SARS-CoV-2 entry and are upregulated in response to viral infection. This functional landscape of SARS-CoV-2 host factors provides a physiologically relevant starting point for new host-directed therapeutics and suggests interactions between SARS-CoV-2 and airway mucins of COVID-19 patients as a host defense mechanism.

Inducible CRISPR activation screen for interferon-stimulated genes identifies OAS1 as a SARS-CoV-2 restriction factor

“…Given the apparently potent effects of OAS1 in our experiments, we were surprised that it had not been detected in other recently published genome-wide and ISG-focused screens 29,43,44,85,86 . This discrepancy could be due to the distinct features of the experimental systems used, including different cell types, different infection time points, and low M.O.I.…”

“…Using a similar genome-wide gRNA library in a different cell line, Dukhovny et al detected antiviral effectors with activity against Zika Virus 42 . Recent preprints also reported genome-wide CRISPRa screens for SARS-CoV-2 43,44 . While these examples demonstrate the utility of pooled CRISPRa screens for identifying viral restriction factors genome-wide, we aimed to establish a CRISPRa system optimized for efficiently evaluating ISG activities against respiratory viruses.…”

CRISPR screens for host factors critical for infection by SARS-CoV-2 variants of concern identify GATA6 as a central modulator of ACE2

“…AP1G1 (Adaptor Related Protein Complex 1 Subunit Gamma 1) encodes a component of clathrincoated vesicles that were shown to mediate endocytosis and intracellular trafficking of SARS-CoV-2 [50,51]. Of note, the clathrin-coating complex, to which AP1G1 belongs, was recently identified as important for SARS-CoV-2 infectivity in a screen conducted in Calu-3 cells but not in screens conducted in other cell lines [21]. Taken together, the data therefore suggest that some of the host entry factors necessary for SARS infection exhibit cell-line specificity illustrating the importance of using relevant cell models to maximize our understanding of SARS-CoV-2 entry.…”

“…Another strategy to explore SARS-CoV-2-host interactions is to disrupt individual genes of the entire host genome and screen for genes whose disruption results in resistance to viral infection and consequently to host-cell survival. Several reports documented genome-wide CRISPR-Cas9-mediated loss-of-function screens and identify host factors that are functionally required for SARS-CoV-2 infection in various cell types [10,[18][19][20][21][22][23][24][25].…”

Systematic genome-scale identification of host factors for SARS-CoV-2 infection across models yields a core single gene dependency; ACE2

“…This motif is also common in multiple viruses, including SARS-CoV-2, allowing viruses to exploit AP2M1 for internalization from the plasma membrane to intracellular sites (Minakshi and Padhan, 2014; Yuan et al, 2020). Calu-3 knockout screens reported by others (Biering et al, 2021; Rebendenne et al, 2021), also identified members of the clathrin adaptor complex; however, the genes were pro-viral and associated with the AP1 complex, which is found on the trans-Golgi-network and endosomes and is responsible for trafficking proteins between the Golgi and/or endosomes to lysosome (Biering et al ., 2021; Touz et al, 2004). Many viruses are known to hijack endocytic mechanisms for entry to the cell (Robinson et al, 2018).…”