Intracellular Ionic Strength Sensing Using NanoLuc

et al. 2022

Preprint

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Major histocompatibility complex class I (MHC-I) molecules, which are dimers of a glycosylated polymorphic transmembrane heavy chain and the small protein β2-microglobulin (β2m), bind peptides in the endoplasmic reticulum that are generated by the cytosolic turnover of cellular proteins. In virus-infected cells these peptides may include those derived from viral proteins. Peptide-MHC-I complexes then traffic through the secretory pathway and are displayed at the cell surface where those containing viral peptides can be detected by CD8+ T lymphocytes that kill infected cells. Many viruses enhance their in vivo survival by encoding genes that downregulate MHC-I expression to avoid CD8+ T cell recognition. Here we report that two accessory proteins encoded by SARS-CoV-2, the causative agent of the ongoing COVID-19 pandemic, downregulate MHC-I expression using distinct mechanisms. One, ORF3a, a viroporin, reduces global trafficking of proteins, including MHC-I, through the secretory pathway. The second, ORF7a, interacts specifically with the MHC-I heavy chain, acting as a molecular mimic of β2m to inhibit its association. This slows the exit of properly assembled MHC-I molecules from the endoplasmic reticulum. We demonstrate that ORF7a reduces antigen presentation by the human MHC-I allele HLA-A*02:01. Thus, both ORF3a and ORF7a act post-translationally in the secretory pathway to lower surface MHC-I expression, with ORF7a exhibiting a novel and specific mechanism that allows immune evasion by SARS-CoV-2.Significance StatementViruses may down-regulate MHC class I expression on infected cells to avoid elimination by cytotoxic T cells. We report that the accessory proteins ORF7a and ORF3a of SARS-CoV-2 mediate this function and delineate the two distinct mechanisms involved. While ORF3a inhibits global protein trafficking to the cell surface, ORF7a acts specifically on MHC-I by competing with β2m for binding to the MHC-I heavy chain. This is the first account of molecular mimicry of β2m as a viral mechanism of MHC-I down-regulation to facilitate immune evasion.

Section: Methodsmentioning

confidence: 99%

SARS-CoV-2 accessory proteins ORF7a and ORF3a use distinct mechanisms to downregulate MHC-I surface expression

Arshad

Laurent-Rolle

et al. 2022

Preprint

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“…Molecular dynamics simulations were performed using NAMD V2.13 software [25] and CHARMM36 force field [26], as described previously [27]. The simulation system consisting of the biomolecular complex formed by the ACE2-S1-RBD was generated from the previously prepared PDB files using the QwikMD Toolkit [28] available as a plugin in Visual Molecular Dynamics (VMD) [29] software V1.9.3.…”

Section: Methodsmentioning

confidence: 99%

Decreased Interfacial Dynamics Caused by the N501Y Mutation in the SARS-CoV-2 S1 Spike:ACE2 Complex

Philip

Biswas

2021

Preprint

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COVID-19 caused by SARS-CoV-2 has caused a massive health crisis across the world, and genetic variants such as the D614G gaining enhanced infectivity and competitive fitness have significantly aggravated the global concern. In this regard, the latest SARS-CoV-2 variant, B.1.1.7 lineage, reported from the United Kingdom (UK) is of great significance, in that it contains several mutations that increases its infection and transmission rates as evidenced by the increased number of clinical reports. Specifically, the N501Y mutation in the SARS-CoV-2 S1 receptor binding domain (RBD) domain has been shown to possess increased affinity for ACE2, although the basis for this not yet clear. Here, we dissect the mechanism underlying the increased affinity using molecular dynamics (MD) simulations of the available ACE2-S1-RBD complex structure (6M0J) and show a prolonged and stable interaction of the Y501 residue in the N501Y mutant S1-RBD with interfacial residues, Y41 and K353, in ACE2 as compared to the wild type S1-RBD. Additionally, we find that the N501Y mutant S1-RBD displays altered dynamics that likely aids in its enhanced interaction with ACE2. By elucidating a mechanistic basis for the increased affinity of the N501Y mutation in S1-RBD for ACE2, we believe that the results presented here will aid in developing therapeutic strategies against SARS-CoV-2 including designing drugs targeting the ACE2-S1-RBD interaction.

“…Molecular dynamics simulations were performed using NAMD version 2.13 software (Phillips et al, 2005) and CHARMM36 force field (Best et al, 2012), as described previously (Altamash et al, 2021). The simulation system consisting of the biomolecular complex formed by the ACE2-S1-RBD was generated from the previously prepared PDB files using the QwikMD Toolkit (Ribeiro et al, 2016) available as a plugin in the Visual Molecular Dynamics (VMD) (Humphrey et al, 1996) software V1.9.3.…”

Section: Ace2-s1-rbd Molecular Dynamics Simulationsmentioning

confidence: 99%

Decreased Interfacial Dynamics Caused by the N501Y Mutation in the SARS-CoV-2 S1 Spike:ACE2 Complex

Philip

Biswas

2022

Front. Mol. Biosci.

Self Cite

Coronavirus Disease of 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has resulted in a massive health crisis across the globe, with some genetic variants gaining enhanced infectivity and competitive fitness, and thus significantly aggravating the global health concern. In this regard, the recent SARS-CoV-2 alpha, beta, and gamma variants (B.1.1.7, B.1.351, and P.1 lineages, respectively) are of great significance in that they contain several mutations that increase their transmission rates as evident from clinical reports. By the end of March 2021, these variants were accounting for about two-thirds of SARS-CoV-2 variants circulating worldwide. Specifically, the N501Y mutation in the S1 spike receptor binding domain (S1-RBD) of these variants have been reported to increase its affinity for ACE2, although the basis for this is not entirely clear yet. Here, we dissect the mechanism underlying the increased binding affinity of the N501Y mutant for ACE2 using molecular dynamics (MD) simulations of the available ACE2-S1-RBD complex structure (6M0J) and show a prolonged and stable interfacial interaction of the N501Y mutant S1-RBD with ACE2 compared to the wild type S1-RBD. Additionally, we find that the N501Y mutant S1-RBD displays altered dynamics that likely aids in its enhanced interaction with ACE2. By elucidating a mechanistic basis for the increased affinity of the N501Y mutant S1-RBD for ACE2, we believe that the results presented here will aid in developing therapeutic strategies against SARS-CoV-2 including designing of therapeutic agents targeting the ACE2-S1-RBD interaction.