Dataset Information

Targeting conserved N-glycosylation blocks SARS-CoV-2 variant infection in vitro.

ABSTRACT:

Background

Despite clinical success with anti-spike vaccines, the effectiveness of neutralizing antibodies and vaccines has been compromised by rapidly spreading SARS-CoV-2 variants. Viruses can hijack the glycosylation machinery of host cells to shield themselves from the host's immune response and attenuate antibody efficiency. However, it remains unclear if targeting glycosylation on viral spike protein can impair infectivity of SARS-CoV-2 and its variants.

Methods

We adopted flow cytometry, ELISA, and BioLayer interferometry approaches to assess binding of glycosylated or deglycosylated spike with ACE2. Viral entry was determined by luciferase, immunoblotting, and immunofluorescence assays. Genome-wide association study (GWAS) revealed a significant relationship between STT3A and COVID-19 severity. NF-κB/STT3A-regulated N-glycosylation was investigated by gene knockdown, chromatin immunoprecipitation, and promoter assay. We developed an antibody-drug conjugate (ADC) that couples non-neutralization anti-spike antibody with NGI-1 (4G10-ADC) to specifically target SARS-CoV-2-infected cells.

Findings

The receptor binding domain and three distinct SARS-CoV-2 surface N-glycosylation sites among 57,311 spike proteins retrieved from the NCBI-Virus-database are highly evolutionarily conserved (99.67%) and are involved in ACE2 interaction. STT3A is a key glycosyltransferase catalyzing spike glycosylation and is positively correlated with COVID-19 severity. We found that inhibiting STT3A using N-linked glycosylation inhibitor-1 (NGI-1) impaired SARS-CoV-2 infectivity and that of its variants [Alpha (B.1.1.7) and Beta (B.1.351)]. Most importantly, 4G10-ADC enters SARS-CoV-2-infected cells and NGI-1 is subsequently released to deglycosylate spike protein, thereby reinforcing the neutralizing abilities of antibodies, vaccines, or convalescent sera and reducing SARS-CoV-2 variant infectivity.

Interpretation

Our results indicate that targeting evolutionarily-conserved STT3A-mediated glycosylation via an ADC can exert profound impacts on SARS-CoV-2 variant infectivity. Thus, we have identified a novel deglycosylation method suitable for eradicating SARS-CoV-2 variant infection in vitro.

Funding

A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

SUBMITTER: Huang HC

PROVIDER: S-EPMC8613501 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Despite the clinical success of anti-spike vaccines, the effectiveness of neutralizing antibodies and vaccines by rapidly spreading SARS-CoV-2 variants has been compromised. Viruses can hijack the glycosylation machinery of host cells to shield themselves from the host’s immune response and attenuate antibody efficiency. However, it still remains unclear whether targeting glycosylation on spike can impair SARS-CoV-2 and its variants infectivity. Methods: To assess the binding ability of glycosylated or deglycosylated spike with ACE2, we performed flow cytometry, ELISA, and BioLayer Interferometry methods. Viral entry ability was determined by luciferase intensity, immunoblotting, and immunofluorescence assay. A genome-wide association study (GWAS) was performed to identify the relationship of STT3A and COVID-19 severity. N-glycosylation regulated by NF-kB/STT3A axis was investigated by knockdown approach, chromatin immunoprecipitation, and promoter assay. To specifically target SARS-CoV-2 infected cells, we developed an antibody-drug conjugate coupling non-neutralization anti-spike antibody with NGI-1 (4G10-ADC) on inhibitory effects of SARS-CoV-2 infection. Results: We found receptor binding domain and three SARS-CoV-2 distinct surface Nglycosylation sites in 57,311 spikes retrieved from NCBI-Virus-database are highly evolutionarily conserved (99.67%) and involved in ACE2 interaction. We further identified STT3A as a key glycosyltransferase that catalyzed spike glycosylation and positively correlated with COVID-19 severity. Inhibition of STT3A by N-linked glycosylation inhibitor-1 (NGI-1) impaired SARS-CoV-2 and its variants (B.1.1.7, and B.1.351) infectivity. Most importantly, 4G10-ADC internalized SARS-CoV-2 infected cells and subsequently released NGI-1 to deglycosylate spike protein. Thereby, it reinforces the neutralizing abilities in antibodies, vaccines, or convalescent sera, inhibiting SARS-CoV-2 and its variants’ infectivity. Our results suggest targeting STT3A-mediated evolution conserved glycosylation via ADC can provide a widespread impact on SARS-CoV-2 variants infection. Together, we identified a novel deglycosylation method to eradicate SARS-CoV-2 variants infection.