Modulation of SARS-CoV-2 Spike-induced Unfolded Protein Response (UPR) in HEK293T cells by selected small chemical molecules
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Abstract
Coronaviruses (CoV) exploits the endoplasmic reticulum (ER) of the host cells for replication and in doing so, increases ER stress. evokes Unfolded Protein Response (UPR) and possibly autophagy, which could all attribute to the pathophysiology of the viral infections. To date, little is known about the roles of ER stress, UPR, and autophagy in SARS-CoV-2 infection. Here we over-expressed the viral Spike (S) protein in cultured HEK293T cells, as it has been shown that such protein is largely responsible for UPR activation in other CoV-infected cells. We noticed, in the transfected cells, heightened ER stress, activation of the PERK-eIF2α arm of the UPR, induction of autophagy and cell death. When we treated the transfected cells with Tauroursodeoxycholic acid (TUDCA), 4-phenyl butyric acid (PBA), Salubrinal, Trazadone hydrochloride, and Dibenzoylmethane (DBM), we saw reduced the BiP/GRP78 levels, but only PBA and TUDCA could significantly diminish the levels of peIF2α and autophagy expression.
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SciScore for 10.1101/2021.02.04.429769: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Anti-GRP78 (BiP), anti-peIF2α, anti-LC3B, and anti-GAPDH were purchased from Cell Signaling Technology Inc. ( anti-GAPDHsuggested: None2 Spike protein-expressing cells were washed with cold PBS, lysed with MPER buffer, and subjected to Western blot analysis with antibodies against LC3B. LC3Bsuggested: NoneExperimental Models: Cell Lines Sentences Resources 2.1. Cells, Antibodies, and Reagents: HEK293T cells obtained from ATCC (Manassas, VA) were cultured in a … SciScore for 10.1101/2021.02.04.429769: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Anti-GRP78 (BiP), anti-peIF2α, anti-LC3B, and anti-GAPDH were purchased from Cell Signaling Technology Inc. ( anti-GAPDHsuggested: None2 Spike protein-expressing cells were washed with cold PBS, lysed with MPER buffer, and subjected to Western blot analysis with antibodies against LC3B. LC3Bsuggested: NoneExperimental Models: Cell Lines Sentences Resources 2.1. Cells, Antibodies, and Reagents: HEK293T cells obtained from ATCC (Manassas, VA) were cultured in a 37°C, 5% CO2 incubator in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% (vol/vol HEK293Tsuggested: NoneSoftware and Algorithms Sentences Resources The signals were detected by Infrared detection using an Odyssey scanner and analyzed by Image Studio software (LI-COR). 2.5. Detection of Autophagy: For the microtubule-associated protein 1A/1B-light chain 3 (LC3) mobility shift assay, Image Studiosuggested: (Image Studio Lite, RRID:SCR_013715)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- No funding statement was detected.
- No protocol registration statement was detected.
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