Genomewide CRISPR knockout screen identified PLAC8 as an essential factor for SADS-CoVs infection

Tse, Longping V.; Meganck, Rita M.; Araba, Kenza C.; Yount, Boyd; Shaffer, Kendall M.; Hou, Yixuan J.; Munt, Jennifer E.; Adams, Lily E.; Wykoff, Jason A.; Morowitz, Jeremy M; Dong, Stephanie; Magness, Scott T.; Marzluff, William F.; Gonzalez, Liara M.; Ehré, Camille; Baric, Ralph S.

doi:10.1073/pnas.2118126119

Cited by 18 publications

(14 citation statements)

References 69 publications

(88 reference statements)

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“…Similarly, it has been shown that deletion of the retromer component VPS29 inhibits SARS‐CoV‐2 infection and results in entrapment of VSV/SARS‐CoV‐2 chimeric viruses in the endosomes and the loss of endosomal cathepsin activity (Poston et al , 2022). It is worth noting that, during the final round of revision of this work, another genome‐wide CRISPR screen has identified PLAC8 as an essential factor for swine acute diarrhea syndrome coronavirus (SADS‐CoV), an alpha‐CoV‐1 virus closely related to beta‐CoVs (Tse et al , 2022). Interestingly, they also concluded that most likely PLAC8 participates in vesicle trafficking in the early stages of the SADS‐CoV life cycle, but it does not affect virion binding or endocytosis.…”

Section: Discussionmentioning

confidence: 99%

Autophagy‐linked plasma and lysosomal membrane protein PLAC8 is a key host factor for SARS‐CoV‐2 entry into human cells

et al. 2022

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Better understanding on interactions between SARS-CoV-2 and host cells should help to identify host factors that may be targetable to combat infection and COVID-19 pathology. To this end, we have conducted a genome-wide CRISPR/Cas9-based loss-offunction screen in human lung cancer cells infected with SARS-CoV-2-pseudotyped lentiviruses. Our results recapitulate many findings from previous screens that used full SARS-CoV-2 viruses, but also unveil two novel critical host factors: the lysosomal efflux transporter SPNS1 and the plasma and lysosomal membrane protein PLAC8. Functional experiments with full SARS-CoV-2 viruses confirm that loss-of-function of these genes impairs viral entry. We find that PLAC8 is a key limiting host factor, whose overexpression boosts viral infection in eight different human lung cancer cell lines. Using single-cell RNA-Seq data analyses, we demonstrate that PLAC8 is highly expressed in ciliated and secretory cells of the respiratory tract, as well as in gut enterocytes, cell types that are highly susceptible to SARS-CoV-2 infection. Proteomics and cell biology studies suggest that PLAC8 and SPNS1 regulate the autophagolysosomal compartment and affect the intracellular fate of endocytosed virions.

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Section: Discussionmentioning

confidence: 99%

Autophagy‐linked plasma and lysosomal membrane protein PLAC8 is a key host factor for SARS‐CoV‐2 entry into human cells

et al. 2022

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“…In agreement with the KYCG probe enrichment analysis, chromatin state discovery and characterization (chromHMM) identified enhancers as heavily enriched at as a top marker for monocyte dysregulation in scRNA-seq datasets collected from both sepsis and severe Covid-19 patients, and its role as an autophagy-linked plasma and lysosomal membrane protein is believed to be essential for SARS-Cov2 infection (Reyes et al, 2020;Schulte-Schrepping et al, 2020;Ugalde et al, 2022;Tse et al, 2022).…”

Section: Dna Methylation Reprogramming At Gene Regulatory Elements In...mentioning

confidence: 73%

“…Given the extent of genome-wide H3K4me1/3 reprogramming normally observed during sepsis and LPS tolerization, it is reasonable to speculate that DNA methylation serves to reinforce changes to the H3K4 methyl landscape in exhausted monocytes (Saeed et al, 2014;Foster et al, 2007;Novakovic et al, 2016). Furthermore, two recent studies demonstrated Plac8 expression as essential for coronavirus infection by regulating either viral entry or transcription in host cells (Tse et al, 2022;Ugalde et al, 2022). Whereas previous studies have implicated Plac8 hypomethylation in endothelial colony-forming cells as a driver for Plac8 upregulation and downstream transcriptional alterations in gestational diabetes mellitus, to our knowledge our study is the first to report DNA methylation as potential driver for Plac8 monocytic dysregulation during sepsis progression (Blue et al, 2015).…”

Section: Discussionmentioning

confidence: 99%

Altered DNA methylation underlies monocyte dysregulation and innate exhaustion memory in sepsis

Caldwell,

Wu,

Wang

et al. 2023

Preprint

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Innate immune memory is the process by which pathogen exposure elicits cell-intrinsic states to alter the strength of future immune challenges. Such altered memory states drive monocyte dysregulation during sepsis, promoting pathogenic behavior characterized by pro-inflammatory, immunosuppressive gene expression in concert with emergency hematopoiesis. Epigenetic changes, notably in the form of histone modifications, have been shown to underlie innate immune memory, but the contribution of DNA methylation to this process remains poorly understood. Using an ex vivo sepsis model, we discovered broad changes in DNA methylation throughout the genome of exhausted monocytes, including at several genes previously implicated as major drivers of immune dysregulation during sepsis and Covid-19 infection (e.g. Plac8). Methylome alterations are driven in part by Wnt signaling inhibition in exhausted monocytes, and can be reversed through treatment with DNA methyltransferase inhibitors, Wnt agonists, or immune training molecules. Importantly, these changes are recapitulated in septic mice following cecal slurry injection, resulting in stable changes at critical immune genes that support the involvement of DNA methylation in acute and long-term monocyte dysregulation during sepsis.

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“…Trypsin may act directly on spike protein or indirectly through other cryptic host receptors, but based on biolayer interferometry studies, BtCoV-422 does not bind the hACE2 receptor ( 21 ). Alternatively, a recently identified host protein, placenta-associated 8, functions as an early replication factor for SADS-CoV infection ( 52 ), and several cofactors have been identified that enhance SARS-CoV-2, MERS-CoV, and seasonal hCoV entry ( 53 , 54 ). Such an alternative entry pathway might reflect “in transition viral adaptation” and could correlate with zoonotic transmission.…”

Section: Discussionmentioning

confidence: 99%

“…Neutralization on Huh7.5 cell was performed as described previously ( 52 ). Briefly, Huh7.5 cells were seeded in at day −1 on a black-well, black-wall, tissue culture–treated, 96-well plate (Corning, catalog no.…”

Section: Methodsmentioning

confidence: 99%

A MERS-CoV antibody neutralizes a pre-emerging group 2c bat coronavirus

Tse,

Hou,

McFadden

et al. 2023

Sci. Transl. Med.

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The repeated emergence of zoonotic human betacoronaviruses (β-CoVs) dictates the need for broad therapeutics and conserved epitope targets for countermeasure design. Middle East respiratory syndrome (MERS)–related coronaviruses (CoVs) remain a pressing concern for global health preparedness. Using metagenomic sequence data and CoV reverse genetics, we recovered a full-length wild-type MERS-like BtCoV/ li /GD/2014-422 (BtCoV-422) recombinant virus, as well as two reporter viruses, and evaluated their human emergence potential and susceptibility to currently available countermeasures. Similar to MERS-CoV, BtCoV-422 efficiently used human and other mammalian dipeptidyl peptidase protein 4 (DPP4) proteins as entry receptors and an alternative DPP4-independent infection route in the presence of exogenous proteases. BtCoV-422 also replicated efficiently in primary human airway, lung endothelial, and fibroblast cells, although less efficiently than MERS-CoV. However, BtCoV-422 shows minor signs of infection in 288/330 human DPP4 transgenic mice. Several broad CoV antivirals, including nucleoside analogs and 3C-like/M pro protease inhibitors, demonstrated potent inhibition against BtCoV-422 in vitro. Serum from mice that received a MERS-CoV mRNA vaccine showed reduced neutralizing activity against BtCoV-422. Although most MERS-CoV–neutralizing monoclonal antibodies (mAbs) had limited activity, one anti-MERS receptor binding domain mAb, JC57-11, neutralized BtCoV-422 potently. A cryo–electron microscopy structure of JC57-11 in complex with BtCoV-422 spike protein revealed the mechanism of cross-neutralization involving occlusion of the DPP4 binding site, highlighting its potential as a broadly neutralizing mAb for group 2c CoVs that use DPP4 as a receptor. These studies provide critical insights into MERS-like CoVs and provide candidates for countermeasure development.

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Genomewide CRISPR knockout screen identified PLAC8 as an essential factor for SADS-CoVs infection

Cited by 18 publications

References 69 publications

Autophagy‐linked plasma and lysosomal membrane protein PLAC8 is a key host factor for SARS‐CoV‐2 entry into human cells

Autophagy‐linked plasma and lysosomal membrane protein PLAC8 is a key host factor for SARS‐CoV‐2 entry into human cells

Altered DNA methylation underlies monocyte dysregulation and innate exhaustion memory in sepsis

A MERS-CoV antibody neutralizes a pre-emerging group 2c bat coronavirus

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