Probing effects of the SARS-CoV-2 E protein on membrane curvature and intracellular calcium
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SciScore for 10.1101/2021.05.28.446179: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Experimental Models: Cell Lines Sentences Resources The gene was later modified for expression in HEK293 cells by adding a fluorescent EGFP tag, in frame, to the C-terminus of the E-protein sequence via overlap extension PCR and subsequently subcloning the product into a pcDNA3.1 vector; generating the EcGFP construct. HEK293suggested: CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045)Microscopy: HEK293T cells were seeded on an 18 mm diameter coverslip and transiently transfected with the constructs mentioned above (Results and Fig. 2). HEK293Tsuggested: NoneRecombinant DNA Sentences Resources The gene was later modified for expression in … SciScore for 10.1101/2021.05.28.446179: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Experimental Models: Cell Lines Sentences Resources The gene was later modified for expression in HEK293 cells by adding a fluorescent EGFP tag, in frame, to the C-terminus of the E-protein sequence via overlap extension PCR and subsequently subcloning the product into a pcDNA3.1 vector; generating the EcGFP construct. HEK293suggested: CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045)Microscopy: HEK293T cells were seeded on an 18 mm diameter coverslip and transiently transfected with the constructs mentioned above (Results and Fig. 2). HEK293Tsuggested: NoneRecombinant DNA Sentences Resources The gene was later modified for expression in HEK293 cells by adding a fluorescent EGFP tag, in frame, to the C-terminus of the E-protein sequence via overlap extension PCR and subsequently subcloning the product into a pcDNA3.1 vector; generating the EcGFP construct. pcDNA3.1suggested: RRID:Addgene_79663)The gene was subcloned in the pFastBac1 (for Sf9 expression) and pEG-BM (for HEK293 expression) vectors and baculovirus was generated according to the bac-to-bac baculovirus expression system. pEG-BMsuggested: NoneSoftware and Algorithms Sentences Resources Colors were added to signals post-acquisition using default look-up tables in Fiji software. Fijisuggested: (Fiji, RRID:SCR_002285)Electrophysiology: Whole-cell patch clamp recordings were performed using an EPC-10 amplifier and Patchmaster software (HEKA Elektronik; Lambrecht/Pfalz, Germany). Patchmastersuggested: (Patchmaster, RRID:SCR_000034)Results from OddPub: Thank you for sharing your data.
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:Although several articles and preprints in the past year74,79–84 have reported simulations of this structure, a review of their results shows similar limitations, with abbreviated timescales,80–82 reliance on secondary-structure restraints,74 elevated RMSDs,74,80 and/or dewetting in the absence of electric fields.74 Instability of the open structure may reflect underdetermination of the starting protein or membrane models, unresolved interactions with the C-terminal or other domains, or other factors yet to be identified. Taken together, we have shown that recombinantly expressed full-length E protein from SARS-CoV-2 is capable of independent multimerization, possibly as a tetrameric or smaller species. We also confirmed that the protein localizes intracellularly, similar to its predecessor from SARS-CoV, with no evidence of ion channel properties at the cell surface. Our coarse-grained simulations further support a role for the E protein in viral budding, as the presence of the protein bends the surrounding membrane. Reduction of intracellular Ca2+ in E-protein-transfected cells may further promote viral replication. However, our atomistic simulations and permeation calculations based on previously reported NMR structures resulted in unstable proteins incapable of Ca2+ conduction. We emphasize the importance of using high-resolution structural data obtained from a full-length protein to gain detailed molecular insight of the E protein, and enable future drug-screening effort...
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.
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Results from scite Reference Check: We found no unreliable references.
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