Journal: Nature Communications

Article Title: Mechanisms of SARS-CoV-2 neutralization by shark variable new antigen receptors elucidated through X-ray crystallography

doi: 10.1038/s41467-021-27611-y

Figure Lengend Snippet: a ELISA screen for identification of potential SARS-CoV-2 RBD binders. Negative control wells containing expression media, and positive control wells containing VNAR E06 (anti-serum albumin) and rabbit monoclonal CR3022-RB (anti-SARS-CoV-2 Spike) are indicated. b Primary screen for identification of neutralizing VNAR domains. Concentration-dependent neutralization of pseudotyped SARS-CoV-2 (black) or SARS-CoV-1 (red) in HEK293T cells transiently expressing ACE2. Data represents mean ± s.e.m. relative luminescence units (RLUs) from n = 3 independent biological experiments. c Left , rank-ordered IC 50 values for neutralizing VNARs from panel ( b ). VNARs with IC 50 < 10 nM (dashed line) were selected for further characterization. Upper right, depiction of primary sequences of selected VNARs, relative length of complementarity determining regions (CDR1, CDR3) and location of cysteine residues (teal) are shown. d Phylogenetic tree of selected virus taxa, divergent lineages of betacoronaviruses are shown. Glycoproteins encoded by the indicated viruses (*) were used to generate pseudoviruses. e Heatmap summarizing IC 50 values for neutralization of the indicated pseudovirus with the indicated VNAR antibody. Values are derived from experiments described in ( f ). f Secondary validation of selected neutralizing VNAR domains. Concentration-dependent neutralization of viral particles pseudotyped with glycoproteins natively encoded by either SARS-CoV-2 (black), SARS-CoV-1 (red), WIV1-CoV (blue). MERS-CoV (green), or VSV (purple) in Calu-3 cells. Cell viability was also assessed in the presence of increasing concentrations of VNARs (yellow). Data represents mean ± s.e.m. RLU values from n = 3 independent biological experiments. g Concentration-dependent neutralization of replication-competent authentic SARS-CoV-2, strain USA_WA1/2020 in Vero E6 cells. Data represents mean ± s.e.m. RFU values from n = 3 independent biological experiments.

Article Snippet: Plasmids used for generating pseudovirus stocks were sourced as follows: plasmid encoding an Env -defective, luciferase-expressing HIV-1 genome (pNL4-3.luc.R-E − ) , was obtained through the NIH AIDS Reagent Program; plasmids encoding SARS-CoV-1 and SARS-CoV-2 Spike were from Fang Li (Addgene plasmid #145031 and #145032, respectively) ; WIV1-CoV Spike was from Alejandro Balazs (Addgene plasmid #164439) ; MERS-CoV Spike was from Sino Biological (VG40069-NF); and VSV-G was from Bob Weinberg (Addgene plasmid #8454).

Techniques: Enzyme-linked Immunosorbent Assay, Negative Control, Expressing, Positive Control, Concentration Assay, Neutralization, Derivative Assay

Journal: Nature Communications

Article Title: Mechanisms of SARS-CoV-2 neutralization by shark variable new antigen receptors elucidated through X-ray crystallography

doi: 10.1038/s41467-021-27611-y

Figure Lengend Snippet: a Sequence alignment between SARS-CoV-1, SARS-CoV-2, and the MERS RBDs. Residues different from SARS-CoV-2 are highlighted in red and the interaction interface for VNAR-3B4 is marked with a blue box. Bolded letters indicate residues critical for the interaction between the RBD and 3B4 with arrows indicating the residues that form a backbone beta-sheet. The sequence alignment is numbered above according to SARS-CoV-2. b Surface representations of SARS-CoV-1 (pink, above) and MERS (green, below) with variant residues colored red. The ACE2 binding interface is highlighted in purple for SARS-CoV-1 and the DPP4 binding interface in orange for MERS. The homology-modeled interaction interface for 3B4 is colored blue for both structures. c Zoomed in view of the modeled interaction interface between 3B4 (blue) and SARS-CoV-1 (pink, above) and MERS (green, below), with 3B4 colored blue in both pictures. Interacting residues are highlighted as in Fig. , showing the backbone interactions in black dashes and sidechain to backbone or sidechain to sidechain interactions shown as yellow dashes. Insets show the orientation of the zoomed in view. d Overlays of the 3B4 interface from modeled RBDs and their matching RBDs obtained by x-ray crystallography. The panel above shows the modeled SARS-CoV-1 RBD, colored pink, aligned with the crystal structure of SARS-CoV-1 RBD bound to ACE2 (PDB id: 2AJF), colored magenta. The panel below shows the modeled MERS RBD, colored light green, aligned with the crystal structure of MERS RBD bound to DPP4 (PDB id: 4L72), colored dark green.

Article Snippet: Plasmids used for generating pseudovirus stocks were sourced as follows: plasmid encoding an Env -defective, luciferase-expressing HIV-1 genome (pNL4-3.luc.R-E − ) , was obtained through the NIH AIDS Reagent Program; plasmids encoding SARS-CoV-1 and SARS-CoV-2 Spike were from Fang Li (Addgene plasmid #145031 and #145032, respectively) ; WIV1-CoV Spike was from Alejandro Balazs (Addgene plasmid #164439) ; MERS-CoV Spike was from Sino Biological (VG40069-NF); and VSV-G was from Bob Weinberg (Addgene plasmid #8454).

Techniques: Sequencing, Variant Assay, Binding Assay

Journal: iScience

Article Title: Gene repression through epigenetic modulation by PPARA enhances hepatocellular proliferation

doi: 10.1016/j.isci.2022.104196

Figure Lengend Snippet:

Article Snippet: C57BL/6J mice were tail-vein injected with pCMV3- CDH1 (Sino Biological, mouse/MG50671-CF, ) or EGFP (Sino Biological, Aequorea Victoria/AG13105-CF) using PEI Max transfection reagent (PEI, Polysciences) according to a method modified from .

Techniques: Recombinant, Chromatin Immunoprecipitation, AST Assay, Immunoprecipitation, shRNA, Luciferase, Software