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Authors: Zhang Y et al.

Link to paper:

Journal/ Pre-Print: Biorxiv

Tags: Autophagy/Lysosome, Cell Biology, Clinical, Immunology/Immunity, Inflammation, Molecular biology, Therapeutics, Virology

Research Highlights 

1. The protein encoding Open Reading Frame 8 (ORF8) in SARS-CoV2 shares the lowest sequence homology to SARS-CoV (26%), and may explain notable differences in pathogenesis between the two.

2. In SARS-CoV2 infected cells, ORF8 abrogates trafficking of MHC-I from the ER to the cell surface. Instead, MHC-I molecules bind directly to ORF8, and directed to lysosomal degradation though an autophagy-dependant pathway.

3. Reduced levels of MHC-I in SARS-CoV2-infected cells impairs CTL-mediated killing, thus allowing the virus to evade immune surveillance and increase in viral load.


The authors focus upon a protein with the lowest sequence homology between SARS-CoV2 and SARS-CoV: Open Reading Frame 8 (ORF8), an accessory protein. Direct binding of ORF8 significantly downregulates both surface and total levels of MHC-I, while isoforms of ORF8 in SARS-CoV have no effect. ORF8-mediated MHC-I degradation is lysosomal, which is confirmed as autophagy-dependant by confocal microscopy, proposing but not proving a role for ER-Phagy. It prevents effective viral antigen presentation and CTL-mediated killing of virally infected cells. The authors propose this can, to an extent, explain why SARS-CoV2 is able to evade immune surveillance and may replicate undetected in vivo.

Impact for SARS-CoV2/COVID19 research efforts

Understand the virology and/or cell biology of SARS-CoV2/COVID19

Ø Unique role of 3’ accessory protein ORF8 in SARS-CoV2 allows trafficking of MHC-I from ER to lysosome (instead of to the plasma membrane), ultimately directing it for degradation and vastly reducing both cell surface and total levels of MHC-I in virally infected cells.

Ø Reduced MHC-I levels result in ineffective antigen presentation, and this results in impaired CTL-mediated killing of virally infected cells, thus allowing the virus to go largely undetected and evade an appropriate anti-viral response.

Ø Not a result of broadly abrogated endocytosis process, but instead as a result of the direct binding of MHC-I to ORF8, and the purposeful misdirection of the receptor.

Ø Similar principle to other viral infections able to evade immune surveillance, such as HIV-I protein Nef, KSHV proteins K3&K5, and E3/E19 protein encoded by adenovirus, all impairing the function of MHC-I. Mechanistically different, principally similar

Ø Implies role of ER-Phagy to control process but have tried to knock-down several proteins involving classical ER-phagy but seems not the case.

Treat of SARS-CoV2/COVID19 positive individuals

Ø Target ORF8 to prevent SARS-CoV2 immune surveillance evasion and promote self-CTL-mediated killing of virally infected cells.

o NB L84S mutant in SARS-CoV2 ORF8 protein was key for genotyping but did not affect the ability of ORF8 to downregulate MHC-I

Study Type

· In vitro study

· Patient Case study (very small!)

Strengths and limitations of the paper

Novelty: > ORF8 highlighted to interact with human proteins in recent nature paper doi:10.1038/s41586-020-2286-9 (2020)

Standing in the field: > Current drugs focus largely on targeting enzymes and structural proteins essential for viral replication. This study looks more at the ability to ‘remove the cloak’ hiding the virus, thus allow the immune system to detect and orchestrate the killing of these infected cells.

> Some viruses (e.g. HIV and influenza) show ability to downregulate MHC-I molecule to evade immune surveillance. It may explain why some COVID-19 patients are asymptomatic or fail to completely clear the virus after recovery.

Appropriate statistics: > Yes, mainly using 1-Way-ANOVA and T-Test, although in places not entirely clear which test was used where/specifics of test.

Viral model used: > Large portion of ORF8 exp. focus on overexpression in HEK293T cells. Validation of ORF8’s effect in MHC-I downregulation also involves human fetal colon cell line FHC, human bronchial epithelial cell line HBE, and human liver cell line Huh7.

> Progressed to looking at expression of ORF8/MHC-I in SARS-CoV-2 strain named hCoV-19/CHN/SYSU-IHV/2020

> Final Exp looked at SARS-CoV-2-specific CD8+ T cells from 5 patients recovered from SARS-CoV2 to evaluate whether exposure to SARS-CoV2 can alter the biology of your CTLs.

Translatability: Strong fundamentally, needs testing in vivo and lacking a molecule to target ORF8 to abrogate its action/block its function

Main limitations: >Final experiment looking at patient specific SARS-CoV-2-specific CD8+ T cells feels very weak and read into. Only 5 patients, HC not numerically matched, nor are any details concerning the patient matching to HC’s.

>Lack of direct molecular mechanism of how ORF8 hijacks MHC-I into autophagy degradation. (e.g. ORF8 recruits MHC-1 containing vesicles released from ER to autophagosomes? Or it integrates the vesicles into the double-membrane structure of the autophagosomes?)