| Starr, T. N |
Measuring the effect of RBD amino acid mutations on the expression of folded protein and its affinity for ACE2 |
Yeast cells |
Deep mutational scanning, FACS, Illumina sequencing, Pseudotypedlentiviral assay, binding assays |
ACE2 |
Most mutations were deleterious for RBD expression and ACE2 binding, while other mutations that had not been selected in the current SARS-CoV-2 pandemic isolates were well tolerated or even enhanced ACE2 binding |
(19) |
| Chan, K. K |
Analyzing the effect of ACE2 mutations on spike binding using deep mutagenesis |
Fetal kidney (293T cells) |
Flow cytometry, ACE2 catalytic activity assay, ELISA, virus microneutralization assay, VSV-G pseudovirus assay, spectroscopy |
ACE2 |
Mutations in the asparagine 90-glycosylation motif and at buried sites increased affinity. The HR1 sequence in the S2 possessed markedly increased alpha-helicity, thermostability, and a better binding affinity with its corresponding HR2 site |
(20) |
| Zhou, L |
Analyzing human post-mortem eyes of COVID-19 patients and surgical specimens for the expression of ACE2 and TMPRSS2 |
Eye tissue |
Immunohistochemistry, Western blot |
ACE2, TMPRSS2 |
IHC and WB Identified the expression of ACE2 and TMPRSS2 in conjunctiva, limbus, and cornea of the eye tissue |
(21) |
| Zhang, Q |
Exploring the binding possibility of sEVs and exomeres containing ACE2 to SARS-CoV-2 |
Colorectal cancer (LS174T, LIM1215, DiFi, DLD-1, Caco-2, DKO-1 cell lines) |
IB, FAVS, IP, Lectin Precipitation Assays |
ACE2, DPP4, TACE, TMPRSS2,4 |
TACE, TMPRSS2, TMPRSS4, ACE2, and DPP4 were secreted in all or two/three sEVs cells. ACE2-positive DiFisEVs and exomeres could bind to the RBD, whereas sEVs and exomeres from the ACE2-negative LS174T cell line could not |
(22) |
| L. Torre-Fuentes |
Analyzing the frequency of exonic variants of the ACE2, TMPRSS2, and Furin genes in relation to the presence or absence of SARS-CoV-2 infection in a familial multiple sclerosis cohort including 120 individuals from Madrid |
CNS & myelin (nerve fibers in the brain and spinal cord) |
Whole-exome sequencing |
ACE2, TMPRSS2, furin |
ACE2 presented a low level of polymorphism, with only two variants, while TMPRSS2 was highly polymorphic and its two variants showed a significant association with SARS-CoV-2 infection. Several variants were found in the furin, none of which was associated with COVID-19 |
(23) |
| Yamamoto, M |
Establishing a quantitative fusion assay dependent on SARS-CoV-2 S protein, ACE2, and TMPRSS2 |
Fetal kidney (Calu-3, HEK293FT, and Vero cell lines) |
DSP Assay, WB, and SARS-CoV-2 infection assay |
ACE2, TMPRSS2 |
Nafamostatmesylate significantly inhibited the fusion, while camostatmesylate was about 10-fold less active. A significantly higher dose was required for VeroE6/TMPRSS2 cells where the TMPRSS2-independent but cathepsin-dependent endosomal infection pathway likely predominated |
(24) |
| Shang, J |
Investigation of receptor binding and protease activation of the spike protein of SARS-CoV-2 by biochemical and pseudovirus entry assays |
Human embryonic kidney (HEK293T cell line) |
Pseudovirus entry assay, siRNA assay, WB, protein pull-down assay |
hACE2, TMPRSS2, furin |
The binding affinity of SARS-CoV-2 RBD and entire spike to hACE2 was higher and comparable or lower than that of SARS-CoV, respectively. This indicated that SARS-CoV-2 RBD, albeit more potent, was less exposed than SARS-CoV RBD and led to an efficient entry to the cell while evading immune surveillance. Unlike SARS-CoV, cell entry of SARS-CoV-2 was preactivated by furin |
(25) |
| Ou, X. Y |
Identification of SARS-CoV-2 entry to host cells and spike interactions |
Fetal kidney (293/hACE2 cells) |
SARS-CoV-2 pseudovirus system, WB, flow cytometry, MTT, fusion assay |
ACE2, PIKfyve, two-pore segment channel 2 (TPC2), CTSL |
SARS-CoV-2 used ACE2 as its receptor and entered cells mainly through endocytosis. PIKfyve, TPC2, and CTSL were vital for entry. SARS-CoV-2 S protein was less stable than the S protein of SARS-CoV |
(26) |
| Wrapp, D |
Determining the cryo-EM structure of the SARS-CoV-2 S trimer in the prefusion conformation and its binding kinetics to ACE2 |
Kidney (FreeStyle 293 cells) |
Cryo-EM, negative-stain EM, SPR sensorgram, affinity chromatography |
ACE2 |
The spike bound ACE2 with higher affinity than did SARS-CoV S. The predominant state of the trimer had one of the three RBDs rotated up in a receptor-accessible conformation |
(27) |
| Zhu, Y |
Verification of SARS-CoV-2 interaction with human ACE2 as a cell receptor |
Fetal kidney (293T cells) |
Single-cycle infection assay, DSP fusion assay, N-PAGE |
ACE2 |
Spike protein mediated high membrane fusion activity. SARS-CoV-2 might possess an enhanced HR1-HR2 interaction in comparison to SARS-CoV |
(28) |
| Sakaguchi, W |
Determining the presence of ACE2, TMPRSS2, and furin molecules in the oral cavity by examination of dorsal tongue and gingiva, saliva, and tongue coating samples |
Tongue and gingival squamous epithelium, submandibular glands |
IHC, RT-PCR, WB, immunofluorescence staining |
ACE2, TMPRSS2, furin, N-EL |
ACE2 was expressed in the stratified squamous epithelium of the dorsal tongue and gingiva. TMPRSS2 was highly expressed in the stratified squamous epithelium in the keratinized surface layer and the saliva and tongue coating samples. Furin was mainly located in the lower layer of stratified squamous epithelium and was detected in the saliva but not tongue coating samples. ACE2, TMPRSS2, and furin mRNA expressions were observed in taste bud-derived cultured cells |
(29) |
| Qiao, J |
Investigating the expression levels of receptors and related proteases for better apprehension of the neuropathy in COVID-19 |
Lung cancer cell line (Calu3), brain cell lines (SY5Y, HMC3, U87) |
qRT-PCR, WB |
ACE2, TMPRSS2, and CD147 |
ACE2, CD147, and TMPRSS2 were expressed in both human and mouse brain cell lines, but the expression pattern was very distinct from that of the lung. ACE2 was expressed less, but CD147 was highly expressed in brain cell lines. Mouse brain tissues were mostly compared with lung cell lines and tissues, and TMPRSS2 had a consistent expression in brain cell lines and mouse lung tissues |
(30) |
| Aguiar, J. A |
To determine the expression and in situ localization of candidate receptors in the respiratory mucosa for SARS-CoV-2 |
Lung (Calu-3, HBEC-6KT cells) |
scRNAseq, microarray, IB, IHC, gene promoter activity analysis |
ACE2, GRP78, TMPRSS2, ADAM17, CTSL-1, CD147 |
Low expression of ACE2 gene and absent to low promoter activity of ACE2 in a variety of lung epithelial cells. Rare ACE2 protein expression was detectable in the airway epithelium and alveoli of the human lung. TMPRSS2, CD147, and GRP78 proteins were expressed in vitro in airway epithelial cells. Broad in situ protein expression of CD147 and GRP78 in the respiratory mucosa was examined |
(31) |
| Maya, E. A. L |
Investigating the roles of ACE2 and TMPRSS2 genes (1273 genetic variants) in human phenotypes of general population (n=36339) by the 178 PheWAS technique |
. . . |
PheWAS, genotyping |
ACE2, TMPRSS2 |
None of the gene variants reached the threshold for significance. SNPs near the TMPRSS2 genes were associated with thrombocytes count. SNPs of the ACE2 gene were associated with the use of Angiotensin II Receptor Blockers (ARBs) combination therapies as well as with the use of non-steroid anti-inflammatory and antirheumatic products |
(32) |
| Leonardi, A |
Examination of SARS-CoV-2 receptors gene expression in conjunctival epithelial cell samples and ex-vivo cornea samples using microarray or transcriptome sequencing |
Cells from 18 healthy conjunctival imprints and 6 ex-vivo cornea |
RNA extraction and PCR, microarray, or transcriptome sequencing |
ACE2, CD147, TMPRSS2, BSG, ANPEP, AGTR2, APOBEC3A, ADAR-1 |
ACE2 was slightly expressed in conjunctival samples, while BSG and TMPRSS2 were expressed at intermediate levels in both the conjunctiva and cornea.
ANPEP and AGTR2 were expressed at a low level in the conjunctiva.
APOBEC3A and ADAR-1, two RNA editing enzymes, were involved in antiviral responses and were highly expressed |
(33) |
| Benton, D. J |
Investigating the binding of ACE2 to the furin-cleaved form of SARS-CoV-2 S by cryoEM. Classifying 10 different molecular species including the unbound, closed spike trimer, the fully open ACE2-bound trimer, and dissociated monomeric S1 bound to ACE2 |
Expi293F cells |
Affinity chromatography, EM |
ACE2, furin |
The 10 structures described ACE2 binding events, which destabilized the spike trimer, progressively opening up and out the individual S1 components. The opening process reduced S1 contacts and un-shielded the trimeric S2 core, priming fusion activation and dissociation of ACE2-bound S1 monomers. The structures also revealed refolding of an S1 subdomain following ACE2 binding that disrupted interactions with S2, notably involving Asp61413–15, leading to destabilization of the structure of S2 proximal to the secondary (S2′) cleavage site |
(34) |
| Wrobel, A. G |
Investigating the relationship between SARS-CoV-2 S and the RaTG13 S using cryo-EM. Investigating and comparing both furin-cleaved and uncleaved SARS-CoV-2 S.
Characterizing the relative stabilities and affinities for the SARS-CoV-2 receptor, ACE2, biochemically |
Expi293F |
Cryo-EM, furin treatment |
ACE2 |
Although the overall structures of human and bat virus S proteins were similar, there were key differences in their properties including a more stable pre-cleavage form of human S and about 1000-fold tighter binding of SARS-CoV-2 to the human receptor.
Cleavage at the furin-cleavage site decreased the overall stability of SARS-CoV-2 S and facilitated the adoption of the open conformation, which is required for S to bind to the ACE2 receptor. Cryo-EM and functional analyses of the furin-cleaved spike from SARS-CoV-2 and the closely related spike from bat virus RaTG13 revealed differences in protein stability and binding to human receptor ACE2 |
(35) |
| Zhang, S |
Evaluating the changes in platelet and coagulation parameters in COVID-19 patients.
Investigation of ACE2 expression and direct effect of SARS-CoV-2 virus on platelets |
201 healthy volunteers and 589 patients suspected of having COVID-19 |
RNA extraction, ELISA, RT-PCR, flow cytometry, WB, EM, IF, platelet functional studies, FeCl3-induced thrombus formation, thrombus formation under flow conditions ex vivo, and co-immunoprecipitation |
ACE2, TMPRSS2, integrin PAC-1 binding, and CD62P (P-selectin) |
Platelets expressed ACE2 and TMPRSS2.
Detectable SARS-CoV-2 RNA in the bloodstream was related to platelet hyperactivity in critically ill patients.
SARS-CoV-2 and its spike directly increased platelet activation such as platelet aggregation, PAC-1 binding, CD62P expression, alpha granule secretion, dense granule release, platelet spreading, and clot retraction in vitro.
Spike protein enhanced the formation of thrombosis in wild-type mice transfused with hACE2 transgenic platelets |
(36) |
| Hoffmann, M |
Evaluating the SARS-CoV-2 spike interactions with ACE2 as the entry molecule and the TMPRSS2 as the priming protease |
Liver (Huh-7), kidney (293T, Vero, BHK-21, MDCKII), lung and bronchus (Calu-3, MRC-5, A549, BEAS-2B, NCI-H1299), colon (Caco-2) |
Pseudotyping of VSV and transduction experiments |
ACE2, TMPRSS2 |
SARS-CoV-2 used the SARS-CoV receptor ACE2 to enter the host cell.
The spike protein of SARS-CoV-2 was primed by TMPRSS2.
Antibodies against SARS-CoV spike might have the ability to protect against SARS-CoV-2.
A TMPRSS2 inhibitor blocked entry and might constitute a treatment option.
Sera from convalescent SARS patients cross-neutralized SARS-2-S-driven entry |
(37) |
| Ratajczak, M. Z |
Investigating the expressions of ACE2 and TMPRSS2 on HSCs and EPCs cells in human umbilical cord blood |
HSCs, EPCs, VSELs, MNC cells |
Real-Time qPCR, FACS |
ACE2, CMA1, AGTR1&2, MAS1, RENIN, TMPRSS2, angiotensin 1–7, Nlrp3 |
ACE2 and TMPRSS2 were expressed on very small CD133 (+) CD34 (+)/Lin (−) CD45 (−) cells in human umbilical cord blood.
The interaction of the ACE2 with the spike in VSELs and HSCs cells activated the Nlrp3 inflammasome, which might lead to cell death by pyroptosis, if hyperactivated.
ACE2 was expressed on the surface of HSCs and VSELs of murine bone marrow, although it is known that murine cells are not infected by SARS-CoV-2 |
(38) |
| Clausen, T. M |
Clarifying the interaction of SARS-CoV-2 spike protein via RBD segment with both cellular heparan sulfate and ACE2 |
Vero E6 or Hep3B cells, ExpiHEK cells, H1299 cells, A549, Hep3B cells, and A375 cells |
TEM and NS microscopy, flow cytometry, LC-MS, IB, qPCR, pseudotyped VSV, virus plaque assays |
Heparan sulfate, ACE2 |
Both ACE2 and heparin could bind independently to the spike in vitro, and a ternary complex could be generated using heparin as a scaffold.
Heparin increased the open conformation of the RBD, which bound ACE2.
Unfractionated heparin, non-anticoagulant heparin, heparin lyases, and lung heparan sulfate potently blocked spike protein binding and/or infection |
(39) |
| Barnes, C. O |
Determining 8 hNAbs structures in complex with SARS-CoV-2 spike trimer or RBD. |
Expi293F cells |
X-Ray crystallography, cryo-EM, polyreactivity assays, SPR binding experiments |
ACE2 |
Structural comparisons allowed classification into four categories: (1) VH3-53 hNAbs with short CDRH3s that blocked ACE2 and bound only to “up” RBDs, (2) ACE2- blocking hNAbs that bound both “up” and “down” RBDs and could contact adjacent RBDs, (3) hNAbs that bounded outside the ACE2 site and recognized “up” and “down” RBDs, and (4) previously-described antibodies that did not block ACE2 and bound only “up” RBDs |
(40) |
| Lukassen, S |
Investigating the ACE2 and TMPRSS2 expression levels and their distribution across cell types in the lung tissue |
Lung tissue (12 donors and 39,778 cells), bronchus (4 donors and 17,521 cells) |
Single nuclei isolation and single RNA sequencing library preparation |
ACE2, TMPRSS2, furin |
TMPRSS2 was highly expressed in both tissues in the subsegmental bronchial branches, but ACE2 was predominantly expressed in a transient secretory cell type.
Interestingly, these transiently differentiating cells showed enrichment for pathways related to Rho GTPase function and viral processes, suggesting increased vulnerability for SARS-CoV-2 infection |
(41) |
| Zang, R. C |
Analyzing the productive infection of SARS-CoV-2 in ACE2-positive mature enterocytes in human small intestinal enteroids |
Enteroids from the duodenum, ileum, and colon |
African green monkey kidney epithelial cell line MA104, human embryonic kidney cell line HEK293 |
ACE2, DPP4, ANPEP, ST14, TMPRSS2&4 |
The expressions of two TMPRSS2 and TMPRSS4 proteases facilitated SARS-CoV-2 spike fusogenic activity and promoted virus entry into host cells.
Simulated human colonic fluid inactivated the viruses that were released into the intestinal lumen |
(42) |
| Sacconi, A |
To investigate the differences in the expressions of both ACE2 and TMPRSS2 in normal tissues from the oral cavity, pharynx, larynx, and lung tissues as well as neoplastic tissues from the same areas |
Two HNSCC cell lines (Cal-27 and Detroit-562 cell lines) |
RNA extraction, qRT-PCR, SARS-CoV-2 detection, NGS, IHC, TCGA, microRNA. |
ACE2, TMPRSS2 |
A significant reduction of TMPRSS2 expression in HNSCC compared to the normal tissues was observed. It was more evident in women than in men, in TP53 mutated than in wild TP53 tumors, and in HPV negative patients compared to HPV positive counterparts.
Functionally, the multivariate effects of TP53, HPV, and other inherent variables on TMPRSS2 were modeled.
In tumor tissues, HPV negative, TP53 mutated status, and elevated TP53-dependent Myc-target genes were associated with low TMPRSS2 expression.
There was an anti-correlation between microRNAs and TMPRSS2 expression in SARS-CoV-2 positive HNSCC tissues. There was no TMPRSS2 promoter methylation. Tumoral tissues might be more resistant to SARS-CoV-2 infection due to the reduced expression of TMPRSS2 |
(43) |
| Allen, J. D |
Impact of ACE2 glycosylation on virus binding |
HEK 293F cell |
Mass spectrometry, purification, SPR |
ACE2 |
ACE2 glycan processing had a subtle Influence on SARS-CoV-2 receptor recognition |
(44) |
| Bojkova, D |
Detection of viral RNA in supernatants of infected cardiomyocytes |
iPS-CMs, Caco-2 |
RNA sequencing |
ACE2- and cathepsin |
Infection of cardiomyocytes in vitro in cathepsin and ACE2-dependent manner |
(45) |
| Hörnich, B. F |
SARS-CoV-2 and SARS-CoV spike-mediated cell-cell fusion |
293T, Calu-3 |
Western blotting, flow cytometry, RT-qPCR |
S and ACE2 |
SARS-CoV-2 spike-mediated cell fusion differed in the expression of its receptor and proteolytic activation |
(46) |
| Kuzmina, A |
SARS-CoV-2 spike variants and convalescent or post-vaccination sera |
10 human, HEK-ACE2 |
Neutralization assays |
S and ACE2 |
Spike variants of SARS-CoV-2 exhibited differential neutralization resistance and infectivity to post-vaccination or convalescent sera |
(47) |
| Müller, J. A |
Infection of endocrine and exocrine pancreas cells by SARS-CoV-2 |
Vero E6, Calu-3, EndoC-βH1, hESC |
RT-qPCR, TCID50, RNA-seq, immunoblotting, immunofluorescence |
ACE2 and TMPRSS2 |
The pancreas as a target of SARS-CoV-2 infection |
(48) |
| Sanders, D. W |
Cholesterol function in SARS-CoV-2 entry and pathological syncytia formation |
VeroE6, HEK293T, U2OS, Beas2B, Calu3, A549 |
TIRF imaging, heterokaryon co-culture assay, FRAP, GPMVs, pseudovirus blocking assay, pseudovirus luciferase assay, titration, SARS-CoV-2 cholesterol depletion assay, RT-qPCR, immunofluorescence |
ACE2, S |
SARS-CoV-2 used cholesterol for entry to cell and pathological syncytia formation |
(49) |
| Schmitz, A |
RBD-independent mechanism of SARS-CoV-2 Spike |
Single-stranded DNA aptamer |
DNA interaction analysis, NGS, SPR, ELONA, protein expression and purification, pulldown assays, chromatography, pseudovirus generation |
ACE2 |
The aptamer did not block the interaction of CoV2-S with ACE2, but studies showed potent and specific pseudo viral infection inhibition by the aptamer |
(50) |
| Suprewicz, Ł |
Vimentin and SARS-CoV-2 entry |
HEK293 |
Tissue staining, dynamic light scattering, atomic force microscopy |
ACE2, S |
Focusing on targeting cell host surface vimentin as a new therapeutic strategy for preventing SARS-CoV-2 infection |
(51) |
| Yeung, M. L |
Investigation of soluble ACE2 role in the entry of SARS-CoV-2 |
HK-2, Caco-2, A549, Calu-3, Huh7, HepG2, PLC/PRF/5, RD, HeLa, NT2, and 293T |
Knockdown of gene expressions by siRNAs, high-throughput sequencing, TCID50 assay, confocal and immunofluorescence microscopic analysis, qRT-PCR, Western blot, co-immunoprecipitation assay |
ACE2 |
Angiotensin system proteins induced cell entry of SARS-CoV-2 |
(52) |
| Zeng, C |
SARS-CoV-2 and cell-to-cell transmission |
293T, Vero-E6, Vero-ACE2, Caco-2, Calu-3, NCI-H520, 293T/ACE2 |
Plaque assay, flow cytometry, Western blotting, neutralization assays |
ACE2 |
SARS-CoV-2 could spread by cell-to-cell transmission |
(53) |
| J. Kruger |
Drug inhibition of SARS-CoV-2 replication in human pluripotent stem cell-derived intestinal organoids |
Human intestinal organoids derived from pluripotent stem cells (PSC-HIOs) |
Immunostaining |
ACE2, TMPRSS2 |
Low doses of remdesivir efficiently inhibited SARS-CoV-2 infection and rescued PSC-HIO morphology |
(54) |
| M. R. Bristow |
Dynamic regulation of SARS-Cov-2 binding and cell entry mechanisms in remodeled human ventricular myocardium |
Remodeled human ventricular myocardium |
RNA-seq, microarray |
ACE2 |
Up-regulation in ACE2 in remodeled left ventricles might explain worse outcomes in patients with COVID-19 with underlying myocardial disorders and minimize cardiac damage by counteracting ACE2 up-regulation as a possible therapeutic approach |
(55) |
| R. Tandon |
Using heparin and enoxaparin derivatives to inhibit the entry of SARS-CoV-2 |
HEK293T cells |
Inhibitor screening, surface plasmon resonance |
ACE2 |
Concentration-response curves proved that unfractionated heparin (UFH), enoxaparin, 6-O-desulfated UFH, and 6-O-desulfated enoxaparin could efficiently neutralize the spike expressing lentiviral vector (pLV-S) particles with different concentrations. The low serum bioavailability of intranasally administered UFH along with the data suggesting that the nasal epithelium is a portal for initial infection and transmission indicated that the intranasal administration of UFH might be an effective and safe prophylactic treatment |
(56) |
| T. L. Pham |
Elovanoid-N32 or RvD6-isomer decreased ACE2 and binding of S protein RBD after injury or INFγ in the eye |
Human corneal epithelial cells |
RNA-sequencing, immunohistochemistry, droplet digital PCR (dd-PCR), capillary-based Western blot, high-throughput qPCR |
ACE2 |
The expression of the ACE2 receptor, furin, and integrins in damaged corneas or IFNγ-stimulated human corneal epithelial cells (HCEC) was consistently decreased by Elovanoid (ELV)-N32 or Resolvin D6-isomer (RvD6i). There was also a concomitant decrease in the binding of spike RBD with the lipid treatments |
(57) |
| J. M. Calandria |
Downregulation of SARS-CoV-2 cell entry, canonical mediators, and enhancement of protective signaling in human alveolar cells by elovanoids |
Human alveolar cells |
Western blot, real-time PCR, eSight assay |
ACE2 |
Elovanoids downregulated ACE2 and enhanced the expression of a set of protective proteins hindering cell surface virus binding and upregulating defensive proteins against lung damage. In addition, the signal of spike (S) protein found in SARS-CoV-2 infected cells was decreased by Elovanoids and their precursors |
(58) |
| S. A. Sheehan |
Inhibition of SARS-CoV-2 infection and COVID-19 disease progression by Maackia amurensis seed lectin (MASL), which exerts pleiotropic actions on oral squamous cells |
HSC-2 cells |
RNA sequence analysis, Western blotting, transcriptional reporter assays |
ACE2 |
MASL targeted the ACE2 receptor, decreased ACE2 expression and glycosylation, suppressed binding of the SARS-CoV-2 spike protein, and decreased the expression of inflammatory mediators by oral epithelial cells that caused ARDS in COVID-19 patients |
(59) |
| J. Taneera |
Expression profile of SARS-CoV-2 host receptors in human pancreatic islets revealed the upregulation of ACE2 in diabetic donors |
Human pancreatic islets |
Microarray and RNA-sequencing |
ACE2, TMPRSS2, and ADAM17 |
Pancreatic islets expressed ACE2, TMPRSS2, and ADAM17 receptors irrespective of diabetes status. ACE2 expression was significantly increased in diabetic/hyperglycemic islets compared to non-diabetic ones. Islets from female donors had higher ACE2 expression than those from males. The expressions of ADAM17 and TMPRSS2 were not affected by gender. The expressions of the three receptors were similar in young (⩽40 years old) and old (⩾60 years old) donors. Obese (BMI >30) donors had higher expression levels of ADAM17 and TMPRSS2 compared to non-obese donors (BMI <25). TMPRSS2 expression was positively correlated to HbA1c and negatively to age, while ADAM17 and TMPRSS2 were positively correlated to BMI. The expressions of the three receptors were similar in muscles and subcutaneous adipose tissues obtained from diabetic and nondiabetic donors. ACE2 expression was higher in sorted pancreatic β-cells compared to other endocrine cells |
(60) |
| C. Y. Wu |
Evaluation of GB-2 ability in the inhibition of ACE2 and TMPRSS2 expression by in vivo and in vitro studies |
HepG2 cells and 293 T cells |
Western blot analysis, quantitative real-time PCR, immunohistochemistry |
ACE2, TMPRSS2 |
The results indicated that the expressions of ACE2 mRNA, ACE2, and TMPRSS2 protein in HepG2 and 293 T cells could be inhibited by GB-2 without cytotoxicity |
(61) |
| K. J. Senthil Kumar |
Downregulation of ACE2, a SARS-CoV-2 Spike receptor-binding domain, in epithelial cells by geranium and lemon essential oils and their active compounds |
Human colorectal adenocarcinoma cell line (HT-29) |
Immunoblotting, ELISA, and quantitative real-time PCR |
ACE2 |
Geranium and lemon oils indicated significant ACE2 inhibitory effects in epithelial cells. Citronellol, geraniol, and neryl acetate were the major compounds of geranium oil and limonene that represented the major compound of lemon oil. Moreover, treatment with citronellol and limonene showed the significant downregulation of ACE2 expression in epithelial cells |
(62) |
| P. Karoyan |
Blockage of SARS-CoV-2 pulmonary cells infection by human ACE2 peptide-mimics |
Calu-3 |
ELISA, biolayer interferometry experiments |
ACE2 |
Peptide-mimics blocked SARS-CoV-2 human pulmonary cell infection with an inhibitory concentration (IC50) in the nanomolar range upon binding to the virus spike protein with high affinity |
(63) |
| K. Y. Huang |
Effective blockage of viral entry and prevention of SARS-CoV-2 infection by humanized COVID-19 decoy antibody |
HEK293T and H1975 cells |
ELISA, Western blotting, flow cytometry, immunofluorescence staining, immunoprecipitation |
ACE2 |
ACE2-Fc showed specific abrogation of virus replication by blocking the entry of SARS-CoV-2 spike-expressing pseudotyped virus into both ACE2-expressing lung cells and lung organoids. Furthermore, this Fc domain of ACE2-Fc activated NK cell degranulation after co-incubation with Spike-expressing H1975 cells |
(64) |
| T. L. Ou |
Attenuation of the hydroxychloroquine-mediated inhibition of SARS-CoV-2 entry by TMPRSS2 |
HEK293T, H1299, and H1975 cell lines |
Luciferase assay |
ACE2 |
The entry of SARS-CoV-2 into cells was more dependent on TMPRSS2 than on SARS-CoV-1. This difference was obvious when the furin-cleavage site of the SARS-CoV-2 S protein was ablated or when it was introduced into the SARS-CoV-1 S protein. Hydroxychloroquine showed efficient blockage of viral entry mediated by cathepsin L, but not by TMPRSS2, and a combination of hydroxychloroquine and a clinically-tested TMPRSS2 inhibitor prevented SARS-CoV-2 infection more potently than either drug alone |
(65) |
| E. Prieto-Fernández |
Reduction of cell attachment of SARS-CoV-2 spike protein by hypoxia through modulating the expressions of ACE2, neuropilin-1, syndecan-1, and cellular heparan sulfate |
NCI-H460 human lung epithelial cells |
Lactoferrin assay, flow cytometry, quantitative PCR, Western blot |
ACE2 |
The binding of the spike to NCI-H460 human lung epithelial cells could be inhibited by hypoxia by decreasing the cell surface levels of heparan sulfate (HS), a known attachment receptor of SARS-CoV-2. This interaction was also reduced by lactoferrin, a glycoprotein that blocks HS moieties on the cell surface. The expression of syndecan-1, an HS-containing proteoglycan expressed in the lung, was inhibited by hypoxia in an HIF-1α-dependent manner. Hypoxia or deletion of syndecan-1 resulted in the reduced binding of the RBD to host cells |
(66) |
| P. A. C. Wing |
Inhibition of SARS-CoV-2 infection of lung epithelial cells by hypoxic and pharmacological activation of HIF |
Lung epithelial cells, RKO, U2-OS, Caco-2, Calu-3, U937, A549, SH-SY5Y cell lines |
Immunoblotting, RT-qPCR
|
ACE2 |
Hypoxia and the HIF prolyl hydroxylase inhibitor Roxadustat reduced the expression of ACE2 and inhibited the entry and replication of SARS-CoV-2 in lung epithelial cells via an HIF-1a-dependent pathway. Hypoxia and roxadustat inhibited SARS-CoV-2 RNA replication, showing that post-entry steps in the viral life cycle were oxygen sensitive |
(67) |
| M. R. Spalinger |
Identification of a novel susceptibility marker for SARS-CoV-2 infection in human subjects and risk mitigation with a clinically approved JAK inhibitor in human/mouse cells |
Samples from IBD patients |
Western blotting, RNA isolation and qPCR, ELISA, RNA sequencing |
ACE2 |
The autoimmune PTPN2 risk variant rs1893217 promoted ACE2 expression and increased cellular entry mediated by SARS-CoV-2 spike protein. Elevated ACE2 expression and viral entry were mediated by increased JAK-STAT signaling and were reversed by the JAK inhibitor, tofacitinib |
(68) |
| M. Ejemel |
IgA MAb blocked the interaction of SARS-CoV-2 spike and ACE2, which provided mucosal immunity |
Human epithelial cells |
ELISA, flow cytometry, virus neutralization assay
|
ACE2 |
MAb362, a cross-reactive human IgA monoclonal antibody that can bind to both SARS-CoV and SARS-CoV-2 spike proteins, competitively blocked hACE2 receptor binding by completely overlapping the hACE2 structural binding epitope. In addition, MAb362 IgA neutralized both pseudotyped SARS-CoV and SARS-CoV-2 in human epithelial cells expressing hACE2 |
(69) |
| Y. M. Hu |
The in vitro antiviral activity of lactoferrin against common human coronaviruses and SARS-CoV-2 was mediated by targeting the heparan sulfate co-receptor |
Human RD, Huh-7 cell, HEK293T cell, HCT-8 cell, Caco-2 cell, Calu-3 cell, and MRC-5 cell lines |
Immunofluorescence, differential scanning fluorimetry, real-time PCR |
ACE2 |
lactoferrin (LF) had broad-spectrum antiviral activity against SARS-CoV-2, HCoV-OC43, HCoV-NL63, and HCoV-229E in cell culture, and bovine lactoferrin (BLF) was more potent than human lactoferrin. BLF bound to heparan sulfate proteoglycans (HSPGs), thereby blocking viral attachment to the host cell. The antiviral activity of BLF could be antagonized by the HSPG mimetic heparin. The antiviral activity of LF was synergistic with remdesivir in cell culture. The N-terminal positively charged region in BLF (residues 17–41) conferred the binding to HSPGs |
(70) |
| V. K. Outlaw |
Ex vivo and in vitro inhibition of coronavirus entry by a lipid-conjugated peptide derived from the SARS-CoV-2 spike glycoprotein hrc domain |
HEK293T cell |
RT-qPCR, RNA sequencing |
ACE2 |
The SARS-CoV-2 C-terminal heptad repeat (HRC)-derived lipopeptide potently blocked the spread of SARS-CoV-2 in human airway epithelial (HAE) cultures, an ex vivo model designed to mimic respiratory viral propagation in humans. While the viral spread of SARS-CoV-2 infection was widespread in untreated airways, those treated with SARS-CoV-2 HRC lipopeptide showed no detectable evidence of viral spread |
(71) |
| Q. Yang |
Blockage of SARS-CoV-2 viral entry upon blocking N- and O-glycan elaboration |
HEK293T and Lenti-X 293 T cells |
Flow cytometry |
ACE2 |
N- and O-glycans had only minor contributions to Spike-ACE2 binding. However, these carbohydrates played important roles in regulating viral entry. Blocking N-glycan biosynthesis at the oligomannose stage using both genetic approaches and the small molecule kifunensine dramatically reduced viral entry into HEK293T cells. Blocking O-glycan elaboration also partially blocked viral entry. Mechanistic studies suggested multiple roles for glycans during viral entry. Among them, inhibition of N-glycan biosynthesis enhanced Spike-protein proteolysis |
(72) |
| J. S. Zhang |
Inhibition of SARS-CoV-2 replication by interferon-stimulated exosomal hACE2 through competitively blocking of the virus entry |
HEK293T, H1299 cells |
Western blot, immunofluorescence |
ACE2 |
hACE2 existed on the surface of exosomes released by different cell types, and the expression of exosomal hACE2 was increased by IFN alpha/beta treatment. In particular, exosomal hACE2 could specifically block the cell entry of SARS-CoV-2, which led to the inhibition of SARS-CoV-2 replication in vitro and ex vivo |
(73) |
| M. Zhong |
Analyzing ACE2 and furin expression in human oral mucosa using the public single-cell sequence datasets and immunohistochemistry |
Mucosal tissue from different oral anatomical sites |
Single-cell sequencing analysis, immunohistochemical staining |
ACE2, furin |
mRNA level expression of furin enzyme and ACE2 receptor were reported in airway epithelia, cardiac tissue, and enteric canals. Immunohistochemistry results indicated that both ACE2-positive and furin-positive cells in the target tissues were mainly positioned in the epithelial layers, partly expressed in fibroblasts. Based on these findings, we speculated that SARS-CoV-2 could invade oral mucosal cells through two possible routes: First, binding to the ACE2 receptor and second, fusion with cell membrane activated by furin protease. The results showed that oral mucosa tissues were susceptible to SARS-CoV-2, which could facilitate COVID-19 infection via respiratory and fecal-oral routes |
(74) |
| S. Shiers |
The hypothesis that ACE2 is expressed in a subset of human dorsal root ganglion (DRG) neurons was tested by using tissues obtained from organ donors or vertebrectomy surgery based on RNAscope in situ hybridization and western blot |
Human dorsal root ganglion |
RNAscope in situ hybridization, imaging by a confocal microscope, immunohistochemistry, Western blot, RNA sequencing |
ACE2 |
It was shown that human DRG neurons expressed the ACE2 at RNA and protein levels. It was also demonstrated that SARS-CoV-2 and coronavirus-associated factors and receptors were broadly expressed in human DRG at lumbar and thoracic levels. ACE2 mRNA was expressed by a subset of nociceptors that expressed MRGPRD mRNA, suggesting that SARS-CoV-2 might gain access to the nervous system through entry into neurons that formed free nerve endings at the outermost layers of skin and luminal organs. Therefore, DRG sensory neurons were a potential target for SARS-CoV-2 invasion of the peripheral nervous system, and viral infection of human nociceptors might cause some of the persistent neurological effects seen in COVID-19 |
(75) |
| L. Chen |
ACE2 expression was investigated in adult human hearts from healthy and diseased individuals |
Human heart tissues |
In-house RNA sequencing, liquid chromatography-tandem mass spectrometry (LC-MS/MS) |
ACE2 |
Pericytes with a high expression of ACE2 might act as the target cardiac cell of SARS-CoV-2. The pericytes injury due to virus infection might result in capillary endothelial cells dysfunction inducing microvascular dysfunction. Besides, patients with basic heart failure showed increased ACE2 expression at both mRNA and protein levels, meaning that if infected by the virus, these patients might have a higher risk of heart attack and critical conditions |
(76) |
| S. Kala |
mRNA levels of ACE2, TMPRSS2, and L-SIGN were compared by qPCR in placentae from pregnant women with HIV exposed to protease inhibitor (PI)-based ART, pregnant women with HIV on non-PI-based ART, and HIV-uninfected women |
Placentae |
qPCR |
ACE2, TMPRSS2, L-SIGN |
Pregnant women of the Black race and WHIV who were on PI-based ART had a relatively lower expression of placental ACE2 compared to the White and HIV-uninfected women. This effect might potentially contribute to altered susceptibility to COVID-19 in these women |
(77) |
| W. P. Cao |
Quantification of the specific interactions between SARS-CoV-2 or SARS-CoV-1 RBD and ACE2 |
HEK293T |
Single-molecule force spectroscopy, steered molecular dynamics (SMD) |
N-linked glycan on Asn90 of ACE2 |
The unbinding forces between SARS-CoV-2 RBD and ACE2 range were 30–40% higher than those of SARS-CoV-1 RBD and ACE2. SARS-CoV-2 RBD interacted with the N-linked glycan on Asn90 of ACE2 |
(78) |
| E. E. F. Brown |
Identifying and characterizing the impact of mutating certain amino acid residues in the RBD of SARS-CoV-2 and ACE2 using NanoBiT technology-based biosensor as well as pseudotyped-virus infectivity assays |
HEK293T cells |
Western immunoblotting, transient transfection, in vitro NanoBit assay, lentiviral pseudovirus assay |
ACE2 |
ACE2 SNPs in the human population might account for the variability in infectivity and disease progression in the context of SARS-CoV-2 infection. Individuals harboring these SNPs (rs766996587, rs759134032, rs200909433, rs961360700) might be more resistant to the SARS-CoV-2 virus infection. |
(79) |
| B. F. Zheng |
Investigation of the potential tropism of SARS-CoV-2, with the recombinant receptor-binding subdomain 1 of the spike protein of SARS-CoV-2 (RBD-SD1) being used as a probe |
Bone marrow cells |
RT-qPCR, immunohistofluorescence |
ACE2 |
Human bone marrow cells could be strongly infected by SARS-CoV-2 |
(80) |
| X. Song |
Investigating the expression level of ACE2 |
Alveolar macrophages, Kupffer cells within liver, and microglial cells in the brain |
Flow cytometry, immunohistochemistry |
ACE2 |
Immunohistochemistry results showed high ACE2 expressions on human tissue macrophages such as alveolar macrophages, Kupffer cells within the liver, and microglial cells in the brain at a steady state. The data suggested that alveolar macrophages, as the frontline immune cells, might be directly targeted by the SARS-CoV-2 infection. Therefore, they have to be considered for the prevention and treatment of COVID-19 |
(81) |
| C. Blume |
cDNA library preparation and sequencing |
|
Mapping, RT-PCR, qPCR, WB |
Short transcript of ACE2 |
Short ACE2 was upregulated in response to IFN and RV16 infection, but not SARS-CoV-2 infection |
(82) |
| P. Varma |
|
Fetal brain |
RNA sequencing, scRNA sequencing |
ACE2, TMPRSS2, furin |
Novel S protein interactors were abundantly present and could play a direct or indirect role in SARS-CoV-2 fetal brain pathogenesis, especially during the 2nd and 3rd trimesters of pregnancy |
(83) |
| J. M. McCord |
Examining gene expression levels |
HepG2 and HPAEC |
GeneChip microarray, RNA-seq assays |
Nrf2, ACE2, TMPRSS2 |
Nrf2 activation might significantly decrease the intensity of the SARS-COV-2 cytokine storm |
(84) |
| M. C. Johnson |
Surface labeling |
293FT or 293FT/ACE2 cells |
Neutralization assay |
ACE2, TMPRSS2 |
Truncation of SARS-COV-2 spike enhanced viral pseudotyping. The D614G and R682Q modifications in SARS-COV-2 Δ19 Spike enhanced pseudotyping efficiency |
(85) |
| A. Goc |
Binding, fusion, and activity assay |
A549 |
WB |
ACE2, TMPRSS2, Catepsin L |
Polyunsaturated fatty acids most effectively interfered with binding to hACE2, linolenic acid, and eicosapentaenoic acid significantly blocked SARS-CoV-2 entry. Eicosapentaenoic acid showed higher efficacy than linolenic acid in reducing the activity of TMPRSS2 and cathepsin L proteases |
(86) |
| A. Tito |
Binding, fusion, and activity assay |
HK-2 |
High-resolution mass spectrometry, Microscale thermophoresis |
Pomegranate peel extract, ACE2, virus 3CL protease |
Pomegranate peel extract inhibited SARS-CoV-2 spike binding to human ACE2 receptors |
(87) |
| Y. Zhan |
Cytotoxicity assay, Ca2+ mobilization assay, surface plasmon resonance assay, virus entry detection |
HEK293 |
Chromatography |
Isorhamnetin, ACE2 |
Isorhamnetin inhibited SARS-CoV-2 spike pseudotyped virus entering ACE2(h) cells |
(88) |
| D. Narang |
|
|
Hydrogen–Deuterium Exchange Mass spectrometry (HDX-MS) |
ACE2 |
31–41 and 353–357 protein regions of hACE2 and hACE2 binding motif of spike protein (residues 437–508) were defined as important protein-protein interfaces |
(89) |
| K. Gorshkov |
Protein conjugation to NPs: QD-spike and AuNP-ACE2, NP-based energy transfer assay |
HEK293T |
Quantum dots |
ACE2 |
Endocytosis of spike bound to ACE2 was one potential mechanism for viral entry |
(90) |
| A. Pramanik |
|
HEK293T |
Raman spectroscopy |
ACE2 |
Antibody attached gold nanoparticles bound to SARS-CoV-2 spike protein, thereby inhibiting the virus from binding to cell receptors, which stopped virus infection and spread. It could also destroy the lipid membrane of the virus |
(91) |
| J. Kim |
mRNA transfection |
Bronchoalveolar lavage |
WB, immunoprecipitation |
ACE2 |
The soluble form of hACE2 bound to SARS-CoV-2 spike protein and prevented viral entry into target cells |
(92) |
| R. C. Larue |
Investigating the SARS-CoV-2 inhibition by ACE2-derived peptides |
HEK293T |
Affinity precipitation assay, plaque assay |
ACE2, ACE2 derived peptides |
(Glu37-Gln42) ACE2 motif that is important for SARS-CoV-2 inhibition, spike-targeting ACE2-derived peptide (SAP) with the amino acid sequence “27-TFLDKFNHEAEDLFYQ-42″, and other SAPs with the amino acid sequence"37-EDLFYQ-42″ inhibited SARS-CoV-2 infection |
(93) |
| M. Lu |
Observing conformational dynamics of SARS-COV2 spike on virus particles |
HEK293T |
Single-molecule fluorescence (Forster) resonance energy transfer (smFRET) imaging |
ACE2, TMPRSS2, trypsin |
Proteolytic processing by serine protease trypsin stimulated hACE2-dependent activation of S |
(94) |
| J. Patiño-Galindo |
Finding the evidence of a recombination event in the RBD involving ancestral lineages to both SARS-CoV and SARS-CoV-2 |
|
Sequence analyzing |
hACE2 |
H427N and F426Y mutations had a significant effect on improving the binding affinity of RaTG13 to hACE2 |
(95) |
| Hoffmann, M |
Investigating the spike cleavage site and the role of furin enzyme in S1/S2 cleavage site |
Lung (Calu-3)Kidney (Vero, 293T) |
Pseudotyped transduction assay, WB, syncytium formation assay |
Furin, TMPRSS2, trypsin |
Furin cleaved the spike at the multibasic S1/S2 site. This cleavage was essential for S-protein-mediated cell-cell fusion and entry into human lung cells.Optimizing the S1/S2 site increased cell-cell fusion, but not virus-cell fusion |
(96) |
| Bestle, D |
Analyzing the role of furin and TMPRSS2 enzymes in cleaving the spike protein |
Lung (Calu-3), kidney (Vero E6, HEK293) |
RNA isolation, RT-PCR, WB, IHC, microscopy |
TMPRSS2, furin |
Furin cleaved the spike at the S1/S2 site and the TMPRSS2 at the S29 site.TMPRSS2 was essential for the activation of spike in Calu-3 human airway epithelial cells.The furin inhibitor in human airway cells inhibited the replication of SARS-CoV-2 |
(97) |
| Kishimoto, M |
Activation of SARS-CoV-2 spike protein |
HEK293T, Vero E6, Vero-T2 |
Immunoblotting, virus entry assay, multi-cycle replication assay, indirect immunofluorescence assay |
TMPRSS11D, TMPRSS13, and S |
TMPRSS11D and TMPRSS13 activated the spike protein of SARS-CoV-2 |
(98) |
| Schönfelder, K |
Polymorphisms of transmembrane serine protease 2 and susceptibility to SARS-CoV-2 |
239 SARS-CoV-2-positive and 253 SARS-CoV-2-negative |
Genotyping |
TMPRSS2 |
TMPRSS2 rs383510 polymorphism (CC genotype) was associated with an increased risk of SARS-CoV-2 infection, but was not correlated to COVID-19 severity. Neither TMPRSS2 rs2070788 nor rs12329760 polymorphisms were related to the risk of SARS-CoV-2 infection or COVID-19 severity |
(99) |
| . B. A. Schuler |
TMPRSS2 expression across the human lifespan was examined by RNA-ISH. |
Human lung tissue |
RNA-ISH, immunofluorescence and image acquisition |
TMPRSS2 |
Expression of TMPRSS2 was highest in ciliated cells and type I alveolar epithelial cells (AT1), and TMPRSS2 expression was increased with aging in humans. SARS-CoV-2 RNA was highly colocalized in the cells expressing TMPRSS2 |
(100) |
| L. Fan |
Magnesium treatment on methylation changes of transmembrane serine protease 2 |
|
DNA methylation |
TMPRSS2 |
Mg treatment significantly increased 5-methylcytosine methylation at cg16371860 (TSS1500, promoter) and reduced 5-hydroxymethylcytosine levels at cg26337277 (proximity to TSS200 and the 5′ untranslated region promoter) |
(101) |
| A. Le Coupanec |
Protein assay, neutralization of endosomal acidification, internalization assay, evaluation of neuroinvasiveness |
CNS |
WB, immunofluorescence |
Cathepsins, furin, TMPRSS2, spinesin, S glycoprotein |
Differences in the cleavage of the S protein and type 1 interferon controlled human coronavirus infection, propagation, and neuropathology within the central nervous system |
(102) |
| Kim, S. Y |
Characterization of the spike of SARS-CoV-2and heparin-binding interactions |
Porcine intestinal HP |
SPR direct binding assay |
Heparin GAGs |
Both monomeric and trimeric forms of SARS-CoV-2 SGP bound more tightly to heparin compared to SARS-CoV and MERS-CoV SGPs. The degree and position of sulfation within HP were important for its successful binding to monomeric SARS-CoV-2 SGP. |
(103) |
| Wang, S. B |
Description of the anti-SARS-CoV-2 function of CH25H and the related mechanisms |
Lung cell lines (Calu-3 cells) |
Pseudovirus system, WB, RNA sequencing, binding, and fusion assay |
CH25H |
CH25H showed extensive anti-coronavirus activity by blocking membrane fusion. 25HC inhibited SARS-CoV-2 viral entry in human lung organoids by activating the ER-localized ACAT that consequently led to the depletion of accessible cholesterol from the plasma membrane |
(104) |
| Farrera-Soler, L |
Applying peptide array to map the antibody response of plasma from healing patients (n=12) and healthy patients (n=6) |
Sf9 insect cells |
RT-PCR, SARS-CoV-2 IgG serology, microarray epitope mapping, ELISA |
Immunodominant linear epitopes |
Three immunodominant linear epitopes were identified and two of them corresponded to key proteolytic sites on the spike protein (S1/S2 and S2’), which is critical for cellular entry. Plasma positive for the epitope adjacent to the S1/S2 cleavage site inhibited the furin-mediated proteolysis of the spike |
(105) |
| Edwards, M. J |
Analyzing the regulatory role of sphingosine in the infection of human epithelial cells by applying pseudoviral particles expressing SARS-CoV-2 spike |
Nasal epithelial cells from healthy donors, Vero, HEK-293T cells |
Pseudotype VSV–SARS–CoV-2, FITC–annexin V binding, TUNEL studies, flow cytometry, kinase assay |
Sphingosine |
Sphingosine bound to ACE2 and prevented the interaction of RBD with ACE2. Thus, exogenously applied sphingosine prevented cellular infection with pp-SARS-CoV-2 spike |
(106) |
| Bayati, A |
Sing lentivirus pseudotyped with spike glycoprotein and purified spike glycoprotein |
HEK-293T and A549 cell lines |
Immunoblot, confocal microscopy, endocytosis assay, purified SARS-Cov-2 spike, and pseudovirus production |
Clathrin |
Viral entry via clathrin-mediated endocytosis |
(107) |
| S. Wang |
Analyzing protein complexes interacting with SARS-CoV-2 S in pulmonary and bronchial cells by tandem affinity purification (TAP)-mass spectrometry (MS) |
BEAS-2B, H1299, human lung cancer tissues, bronchoalveolar lavage fluid cells from COVID-19 patients |
Western blotting, immunoprecipitation, immunofluorescence, LC and MS, tandem affinity purification, cell culture, plasmid construction, transfection, biolayer interferometry quantification assay, quantitative real-time PCR, SARS-CoV-2 virus pseudotype production, and infection |
Tyrosine-protein kinase receptor UFO (AXL) |
AXL was a novel candidate receptor for SARS-CoV-2, which might play an important role in promoting the viral infection of the human respiratory system. The AXL specifically interacted with the N-terminal domain of SARS-CoV-2 S. The AXL expression level was well correlated to SARS-CoV-2 S level in bronchoalveolar lavage fluid cells from COVID-19 patients. Overexpression of AXL promoted SARS-CoV-2 entry as efficiently as did the overexpression of ACE2, while knocking out AXL significantly reduced SARS-CoV-2 infection in vitro |
(108) |
| D. P. Collins |
Evaluation of specific binding of biotinylated spike and spike 1 proteins to different human cells by confocal microscopy |
Multi-lineage progenitor cells (MLPC), alveolar type 2 (AT2) cells, primary human hepatocytes (PHH), and hepatocyte-like cells (HLC) |
Confocal microscopy |
Hepatocyte asialoglycoprotein receptor 1 (ASGr1) |
The absence of ACE-2 receptors and inhibition of spike binding by an antibody to the ASGr1 on both PHH and HLC suggested that the spike protein interacted with the ASGr1. The differential antibody blocking of spike binding to AT2, PHH, and HLC indicated that the neutralizing activity of SARS-CoV-2 binding might involve additional mechanisms beyond RBD binding to ACE-2 |
(109) |
| A. Hudák |
Investigation of the interaction of both SARS-CoV-2 and its spike protein S1 subunit with syndecans isoforms |
K562 cells, A549 |
Cell culture, pseudovirus, flow cytometry, lentiviral vector system to knockdown gens, EZ4U cell proliferation assay, microscopic visualization, co-immunoprecipitation experiments |
Syndecans |
Syndecans, the evolutionarily conserved family of transmembrane proteoglycans, facilitated the cellular entry of SARS-CoV-2. Among syndecans, the lung abundant syndecan-4 was the most efficient in mediating SARS-CoV-2 uptake |
(110) |
| T. Ichimura |
Biopsy and autopsy kidney proximal tubules of COVID-19 patients |
Lung, kidney |
Immunofluorescence staining, immunohistochemistry staining, PCR and cloning, virosome assembly, WB |
KIM-1/TIM-1 |
KIM-1 positive cells expressed less ACE2. KIM-1 was an alternative receptor to ACE2 for SARS-CoV-2. KIM-1 targeted therapeutics might prevent and/or treat COVID-19 |
(111) |
| S. Pfaender |
Immune control of viral disease |
Lymphocytes |
Flow cytometry, WB |
Lymphocyte antigen 6 complex, locus E (LY6E) |
LY6E potently restricted infection by multiple CoVs including SARS-CoV and SARS-CoV-2 |
(112) |
| T. Azad |
Developing a bioluminescence-based bioreporter to interrogate the interaction between the SARS-CoV-2 viral spike (S) protein and ACE2 |
HEK293T |
Bioluminescence |
Asparagine residues |
Glycosylation of asparagine residues within the RBD could mediate successful viral entry/importance of N-linked glycosylation to the RBD’s antigenicity and immunogenicity |
(113) |
| H. Winstone |
Investigating the relevance of the polybasic cleavage site in SARS-CoV-2 route of entry |
Vero E6, A549 |
RT-qPCR, interferon assay |
IFN-induced transmembrane (IFITM), furin, Cathepsin |
SARS-CoV-2 was inhibited by antiviral membrane protein IFITM2 and the sensitivity was exacerbated by deleting the furin cleavage site, which restricted viral entry to low pH compartments |
(114) |
| S. Kim |
|
Upper respiratory system |
Cloning, RT-PCR |
Furin |
Furin was indicated as an S cleavage site on the N-terminal of Arg685 |
(116) |
| T. Tang |
|
Vero E6 cells |
WB, fluorogenic peptide assay |
Furin, S1/S2 |
Furin itself might not be the only protease processing the S1/S2 site. Other proteases might also be involved in this process |
(117) |
| Bayarri-Olmos, R |
ELISA-based angiotensin-converting enzyme-2/RBD inhibition assay |
141 serum samples |
ELISA |
S |
Y453F mutation increased the ACE-2 affinity and did not challenge antibody neutralization |
(118) |
| Collier, D. A |
Neutralizing antibody responses to B.1.1.7 variant following mRNA vaccination |
Serum sample |
Neutralizing test |
S |
E484K mutation in the B.1.1.7 led to a more substantial loss of neutralizing activity |
(119) |
| Daniloski, Z |
Evaluation of spike D614G mutation in SARS-CoV-2 in different human cell types |
A549, HEK293FT, Huh-7.5, Calu-3, Caco-2 |
qPCR, flow cytometry, bio-layer interferometry, trans-complementation assay |
S |
The mutation of spike D614G increased SARS-CoV-2 infection |
(120) |
| Hoffmann, M |
Variants B.1.351 and P.1 of SARS-Cov2 and neutralizing antibodies |
293T, 293T-ACE2, A549, A549-ACE2, A549-ACE2/TMPRSS2, Caco-2, Calu-3, Vero, Vero-TMPRSS2 |
Fluorescence microscopy, qualitative and quantitative cell-cell fusion assay |
S |
B.1.351 and P.1 variants escaped from neutralizing antibodies |
(121) |
| Hou, Y. J |
Ex vivo replication and in vivo transmission of D614G variant |
Vero-81, Vero-E6, Huh7 and A549-ACE2, HNE, LAE, SAE |
Neutralization assay, immunostaining, Western blot, EM imaging, spike quantification, pathological examination |
S |
SARS-CoV-2 D614G variant had efficient transmission in vivo and replication ex vivo |
(122) |
| Li, Q |
Infectivity and immune escape status of SARS-CoV-2 501Y.V2 variants |
Huh-7, Vero, LLC-MK2, HEK293T |
RT-PCR, infection assays, neutralization assays |
S |
SARS-CoV-2 501Y.V2 variants had immune escape but lacked higher infectivity |
(123) |
| Mykytyn, A. Z |
SARS-CoV-2 and entry into human airway organoids |
VeroE6, Vero, Calu-3 |
Immunofluorescence microscopy, immunohistochemistry, fusion assay, entry route assay, entry speed assay, Western blot, silver staining |
S |
Serine proteases helped SARS-CoV-2 to enter airway cells and the multibasic cleavage site was an adaptation to this entry strategy |
(124) |
| Ozono, S |
D614G spike mutation and entry efficiency |
293T, HepG2 |
Cell entry assays, spike incorporation assays, biolayer interferometry, neutralization assays |
S |
D614G spike mutation with enhanced ACE2-binding affinity increased entry efficiency |
(125) |
| Park, E. J |
The spike of SARS-CoV-2 and β1 integrins on the lung epithelial cells |
C57BL/6J mice, MLO-A5, THP-1, 11-18, QG-56, MDA-MB-231, primary lung epithelial cells |
Cell adhesion assay, flow cytometry, RT-qPCR |
S |
SARS-CoV-2 spike bound to β1 integrins on lung epithelial cells |
(126) |
| Sasaki, M |
Generation of SARS-CoV-2 variants with mutations at the S1/S2 cleavage site |
Calu-3, Vero E6, 293T |
Sequencing, plaque assay, immunoblotting, indirect immunofluorescence assay, virus infection assay, deep sequencing |
S |
SARS-CoV-2 during propagation in TMPRSS2-deficient cells generated variants with mutations at the S1/S2 cleavage site |
(127) |
| Zhang, L |
Role of SARS-CoV-2 D614G mutation in infectivity |
HEK293T, NCI-H1299, Calu-3, NCI-H1975 |
Purification, RT-qPCR, entry assay, SPR analyses, neutralization assay |
S |
D614G mutation in S protein increased virion spike density and infectivity |
(128) |
| Zhou, B |
Effect of SARS-CoV-2 spike D614G mutation on replication and transmissibility |
Hamster, mouse, Vero E6, primary human nasal epithelial |
BLI assay, flow cytometry, plaque assay, RT-PCR, sequencing |
S |
D614G variant of SARS-CoV-2 S protein conferred increased transmissibility and replication |
(129) |
| V. Tchesnokova |
Investigation of genetic variations in a 414-583 amino acid region of the spike protein, partially encompassing the ACE2 receptor-binding domain |
Nasopharyngeal |
PCR, RT-PCR, whole-genome sequencing |
S |
Increased infectivity of the L452R variants was due to leucine-452 replacement with arginine, which led to a much stronger binding to the receptor and escape from neutralizing antibodies |
(130) |
| S. M. C. Gobeil |
Exploration of S conformational changes and the effects of the D614G mutation on a soluble S ectodomain construct |
293-F cells |
Protein production and purification, negative-stain electron microscopy, differential scanning fluorimetry, ELISA, cryoelectron microscopy |
S |
D614G mutation altered SARS-CoV-2 spike conformation and enhanced furin cleavage at the S1/S2 Junction.Despite a low mutation rate, isolates with the D614G substitution in the S protein appeared early during the pandemic and are now the dominant form worldwide. Cryoelectron microscopy (cryo-EM) altered RBD disposition. Antigenicity and proteolysis experiments revealed structural changes and enhanced furin cleavage efficiency of the G614 variant. Furthermore, furin cleavage altered the up/down ratio of the RBDs in the G614 S ectodomain, demonstrating an allosteric effect on RBD positioning triggered by changes in the SD2 region, which harbors residue 614 and the furin cleavage site |
(131) |
| M. F. L. Mendoza |
Monitoring viral entry using luciferase vesicular stomatitis virus producing glycoproteins |
HEK293T, BHK-T7, BHK21, Vero s |
Real-time PCR |
S Glycoproteins |
SARS-CoV-2 spike-containing rVSV particles spread poorly in multi-step replication curves and produced significantly lower titers compared to other chimeric viruses. Surprisingly, the first round of entry was comparably efficient and produced luciferase as quickly as rVSV/VSV. |
(132) |
| M. M. Zhao |
Analyzing the circulating levels of CTSL in patients with COVID-19 and systematic study of the therapeutic effects of CTSL on this disease for the first time |
Huh7, A549 cells, human HEK293T cells, plasma samples |
Cell culture, luciferase assay, syncytium-formation assay, real-time PCR, MTT, ELISA, pseudovirus |
Cathepsin L (CTSL) |
The circulating level of cathepsin L elevated after SARS-CoV-2 infection and CTSL functionally cleaved the SARS-CoV-2 spike protein and enhanced virus entry |
(133) |
| R. Amraie |
Investigation of CD209L and CD209 roles in SARS-CoV-2 entry and infection |
Human lung, renal, and temporal arteriole tissues, HUVEC-TERT cells, HEK-293 cells, A549, and H1299 cells |
Immunofluorescence staining, Western blot, shRNA strategy, in vitro angiogenesis assay, neutralization assays, viral entry assay, immunoprecipitation assay, Far-Western blot analysis, dot blot assay |
DC-SIGN (CD209), L-SIGN (CD209L) |
CD209L and CD209 served as alternative receptors for SARS-CoV-2 in disease-relevant cell types including the vascular system.Immunofluorescence staining of human tissues revealed the prominent expression of CD209L in the lung and kidney epithelium and endothelium.CD209L and CD209 interacted with S-RBD.CD209L contained two N-glycosylation sequons at sites N92 and N361, but only site N92 was occupied |
(134) |
| M. Thépaut |
Investigation of the potential interaction of C-type lectin receptors, notably DC/L-SIGN with SARS-CoV-2, through glycan recognition of the spike envelope glycoprotein, as well as their potential role in SARS-CoV-2 transmission |
Calu-3, Jurkat, blood samples |
Protein production and purification, negative staining electron microscopy, surface plasmon resonance binding experiments, pseudovirus |
DC/L-SIGN |
CLRs, particularly DC/L-SIGN, have been associated with important steps of viral entry and infection of different viruses. The results indicated that DC/L-SIGN was an important factor contributing to additional routes of infection mediated by the S protein of SARS-CoV-2. This trans-infection process greatly facilitated viral transmission to susceptible cells |
(135) |
| S. Kamle |
Becausechitinase 3-like-1 induced during aging and comorbid diseases. The relationships between CHI3L1 and SC2 were investigated |
Human plasma samples |
Western blot analysis, real-time PCR, immunohistochemistry, double label immunohistochemistry, SARS-CoV-2 pseudovirus infection, generation of monoclonal antibodies against CHI3L1, co-immunoprecipitation and immunoblot assays, generation of WT and mutant forms of recombinant CHI3L1 |
Chitinase 3-like-1 (CHI3L1) |
CHI3L1 was a potent stimulator of ACE2 and viral spike protein priming proteases (SPP). This induction was a major mechanism contributing to the effects of aging during SARS-CoV-2 infection. CHI3L1 played a critical role in the pathogenesis of COVID-19 and was an attractive therapeutic target. |
(136) |
| E. R. Kastenhuber |
By using biochemical and cell-based assays, it was demonstrated that factor Xa and thrombin could directly cleave SARS-CoV-2 spike, enhancing viral entry |
Calu3, A549, Caco2 |
Enzymatic assay, cell culture, plasmids and lentivirus infection, syncytia assay, Incucyte, luciferase assay, pseudovirus |
Factor Xa, thrombin |
Factor Xa and thrombin could directly cleave SARS-CoV-2 spike and enhance the viral entry.Anticoagulation was critical in the management of COVID-19, and early intervention could provide collateral benefit by suppressing SARS-CoV-2 viral entry |
(137) |
| J. A. Jaimes |
|
|
Fluorogenic peptide assay |
Furin-like, trypsin-like, and cathepsin proteases |
The S1/S2 site could be cleaved by furin-like, trypsin-like, and cathepsin proteases |
(138) |
| K. Wang |
Investigation of the possible function of CD147 in SARS-CoV-2 infection |
BEAS-2B and HEK293T cell lines |
Cell culture, real-time PCR, in vitro SARS-CoV-2 pseudovirus infection test, SPR assay, Co-IP assay, ELISA, electron microscopy imaging, immuno-electron microscope, fluorescence resonance energy transfer assay (FRET), hematoxylin & eosin (HE) staining, multicolor immunofluorescence staining |
CD147 |
Although ACE2 was expressed in the lung, kidney, and intestine, its expression levels were rather low, especially in the lung. The interaction between host cell receptor CD147 and SARS-CoV-2 spike protein was discovered. The loss of CD147 or blocking CD147 in BEAS-2B cell lines by anti-CD147 antibody inhibited SARS-CoV-2. Interestingly, virions were observed in the lymphocytes of the lung tissue from a COVID-19 patient. Human T cells with a property of ACE2 natural deficiency could be infected with SARS-CoV-2 pseudovirus in a dose-dependent manner, which was specifically inhibited by the anti-CD147 antibody. Furthermore, CD147 mediated virus entry into host cells by endocytosis |
(139) |
| J. Shilts |
Removing basigin from the surface of human lung epithelial cells by CRISPR/Cas9 |
Lung epithelial cells |
CRISPR/Cas9 |
Basigin/CD147 |
No evidence for basigin/CD147 as a direct SARS-CoV-2 spike binding receptor |
(140) |
| C. Martino |
The relationship between SARS-CoV-2 susceptibility and the capacity of the human microbiome to catabolize heparan sulfate, a critical host factor involved in mediating SARS-CoV-2 infection, was investigated |
H1299 |
Heparan sulfate modification capability, abundance, and expression by log-ratios, cultivation of bacteroides strains, SARS-CoV-2 spike protein production, biotinylation, SARS-CoV-2 spike binding experiments |
Glycosaminoglycan heparan sulfate |
In vitro, bacterial glycosidases from unpurified culture media supernatants fully blocked SARS-CoV-2 spike binding to human H1299 protein lung adenocarcinoma cells probably by modification of the host glycosaminoglycan heparan sulfate. Thus, commensal host bacterial communities could modify HS, thereby modulating SARS-CoV-2 spike protein binding. These communities changed with age and sex of the host |
(141) |
| L. Cantuti-Castelvetri |
|
Olfactory epithelium |
Cell culture, pseudovirus, lentiviral vector system |
Neuropilin-1 (NRP1) |
NRP1, known to bind furin-cleaved substrates, significantly potentiated SARS-CoV-2 infectivity |
(142) |
| J. L. Daly |
Monitoring viral entry |
HEK293T, HeLa, Caco-2 |
X-Ray crystallography |
NRP1 |
Neuropilin-1 was a host factor for SARS-CoV-2 infection |
(143) |
| J. Davies |
|
Respiratory and olfactory epithelium, CNS |
RNA-Seq |
NRP1 |
Neuropilin-1 was an infection mediator for COVID-19 |
(144) |
| A. J. Carlos |
GRP78 interactions with SARS-2-S and ACE2 were established by utilizing biochemical and imaging approaches |
H1299 |
Flow cytometric analysis, immunofluorescent staining, confocal microscopy, Western blot, generation of VSV pseudotype and transduction experiments, Xtt assay, cell culture |
GRP78 (a stress-inducible chaperone) |
Treatment of lung epithelial cells with a humanized monoclonal antibody (hMAb159) depleted cell surface GRP78 and reduced cell surface ACE2 expression, SARS-2-S-driven viral entry, and SARS-CoV-2 infection in vitro. GRP78 directly bound to the RBD of SARS-2-S and ACE2. GRP78 was an important host auxiliary factor for SARS-CoV-2 entry and infection |
(145) |
| S. He |
Viral infectivity and attachment assay |
HEK293T, Vero E6 |
ELISA, viral infectivity assay |
P-selectin glycoprotein ligand-1 (PSGL-1) |
PSGL-1 might potentially inhibit coronavirus replication in PSGL-1+ cells |
(146) |
