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Comment
. 2022 Nov 14;13(1):6551.
doi: 10.1038/s41467-022-34065-3.

The stress-inducible ER chaperone GRP78/BiP is upregulated during SARS-CoV-2 infection and acts as a pro-viral protein

Woo-Jin Shin #  1 , Dat P Ha #  2   3 , Keigo Machida  4 , Amy S Lee  5   6
Affiliations
Comment

The stress-inducible ER chaperone GRP78/BiP is upregulated during SARS-CoV-2 infection and acts as a pro-viral protein

Woo-Jin Shin et al. Nat Commun. .
No abstract available

PubMed Disclaimer

Conflict of interest statement

A.S.L. is a scientific advisory board member of BiPER Therapeutics. The other co-authors declare no competing interests.

Figures

Fig. 1
Fig. 1. SARS-CoV-2 upregulated GRP78 protein and mRNA levels in infected cells.
a Vero E6-ACE2 cells were mock-infected or infected with SARS-CoV-2 at an MOI of 0.5 in triplicate. The cells were collected at 12, 24, 36, and 48 hr post infection (hpi) and cell lysates were analyzed by western blot for GRP78 protein level with GAPDH serving as loading control. Quantitation of relative protein level of GRP78 normalized against GAPDH was shown in the graph below (n = 3). b Same as in a except the human lung epithelial cell line H1299 was used (n = 3). c Same as in a and b but 36 hpi cell lysates were analyzed by Western blot for SARS-CoV-2 Spike protein level with GAPDH serving as loading control (n = 3). d Vero E6-ACE2 (upper) or H1299 (lower) cells were transiently transfected with control siRNA or siRNAs against GRP78 (left panel) or empty vector pcDNA3 or vector expressing full-length Flag-tagged GRP78 protein (F-GRP78) (right panel) for 48 hr. Cell lysates were analyzed by Western blot for GRP78 protein level with GAPDH serving as loading control. e H1299 cells were mock-infected or infected with SARS-CoV-2 at an MOI of 0.5 or 3. The cells were collected at 24 hpi and cell lysates were analyzed by western blot (n = 4) or RT-qPCR (n = 3) for GRP78 protein or mRNA levels, respectively, with GAPDH serving as an internal control. Quantitation of relative protein (left) or mRNA (right) levels of GRP78 normalized against GAPDH was shown in the graphs below. Data are means ± S.E.M. of three repeats. *P < 0.05; **P < 0.01; ***P < 0.001; ns denotes not significant (Student’s t test). Source data are provided as a Source Data file.
Fig. 2
Fig. 2. GRP78 knockdown by siRNA inhibited SARS-CoV-2 replication in vitro and GRP78 inhibitor HA15 blocked SARS-CoV-2 infection in vitro and in vivo.
a H1299 cells were transiently transfected with control siRNA (siCtrl) or siRNA against GRP78 (si78#1) for 24 hr before infection with SARS-CoV-2 at an MOI of 3 for 24 hr. The cell lysates were analyzed by western blot for GRP78 and Spike protein levels with GAPDH serving as a loading control (n = 3). b The supernatant containing the newly released virions from a was collected and virus titer was determined by plaque assay (n = 3). c Quantitation of the relative protein levels of GRP78 and Spike normalized against GAPDH was shown in the graphs on the left and virus titer (pfu/ml) was shown in the graph on the right. d Confluent monolayers of Vero E6-ACE2 cells in six-well plates were infected with SARS-CoV-2 virus and treated with different concentrations of HA15 as indicated for 72 hr. At the end of treatment, the cells were stained with 0.2% crystal violet. The images are representatives of three repeats. Plaques were counted and plotted in the graph on the left (n = 3). Plaque size was measured and expressed relative to DMSO-treated control in the graph on the right (n = 10). e Vero E6-ACE2 cells cultured under identical conditions as in d were treated with DMSO or increasing concentration of HA15 from 0.625 μM to 5 μM and cell viability was measured by WST-1 assay at 24, 48, and 72 hr after drug treatment (n = 4). f K18-hACE2 transgenic mice infected with SARS-CoV-2 were treated with vehicle control or HA15. Three days post infection, lung tissue RNA was isolated, and RT-qPCR was performed in triplicate reactions to detect SARS-CoV-2 N protein sequence (n = 3). Data are means ± S.E.M. of three repeats. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 (Student’s t test). Source data are provided as a Source Data file.
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  • Multi-level inhibition of coronavirus replication by chemical ER stress.
    Shaban MS, Müller C, Mayr-Buro C, Weiser H, Meier-Soelch J, Albert BV, Weber A, Linne U, Hain T, Babayev I, Karl N, Hofmann N, Becker S, Herold S, Schmitz ML, Ziebuhr J, Kracht M. Shaban MS, et al. Nat Commun. 2021 Sep 20;12(1):5536. doi: 10.1038/s41467-021-25551-1. Nat Commun. 2021. PMID: 34545074 Free PMC article.

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