Journal: The Journal of Cell Biology

Article Title: Epigenetic deprogramming by disruption of CIZ1-RNA nuclear assemblies in early-stage breast cancers

doi: 10.1083/jcb.202409123

Figure Lengend Snippet: Resources

Article Snippet: Primary murine embryonic fibroblasts 13.31, 13.32 (WT female), and 13.33 (WT male) were transduced with virus bearing either the empty pLVX-EF1a-IRES-ZsGreen1 vector (Takara) or the same plasmid expressing the coding sequence of C181 as described in lentivirus transduction.

Techniques: Virus, Recombinant, Transfection, Protease Inhibitor, Mutagenesis, Plasmid Preparation, Generated, Sequencing, Software, Gene Expression, Functional Assay, Inverted Microscopy

Journal: STAR Protocols

Article Title: FlipGFP protease assay for evaluating in vitro inhibitory activity against SARS-CoV-2 M pro and PL pro

doi: 10.1016/j.xpro.2023.102323

Figure Lengend Snippet:

Article Snippet: pLVX-EF1alpha-SARS-CoV-2-nsp5-2×Strep-IRES-Puro (M pro expression plasmid) , Addgene , Cat#141370.

Techniques: Recombinant, Modification, Plasmid Preparation, Transfection, Sequencing, Expressing, Software, Cell Culture, Blocking Assay, Electrophoresis, Inverted Microscopy, Sterility, Transferring, Binding Assay

Journal: bioRxiv

Article Title: The SARS-CoV-2 Spike protein induces long-term transcriptional perturbations of mitochondrial metabolic genes, causes cardiac fibrosis, and reduces myocardial contractile in obese mice

doi: 10.1101/2023.01.05.522853

Figure Lengend Snippet: LDL-c augments Spp viral entry which can be blocked by a SR-B1 inhibitor BLT-1. (A) Spp virus shows a higher maximal binding affinity to LDL-c than HDL-c. The biotinylated LDL-c and HDL-c were incubated with various quantities of Spp virus. The lipoprotein cholesterol bound virus was determined by mCherry RNA copies. (B-E) LDL-c significantly increases cellular uptake of Spp virus in HDMVEC (B), MØ (C), LDLr −/− MØ (D), and H9C2 at 2 hpi (E) ( p <0.001 or 0.005). The addition of BLT-1 significantly blocks the LDL-c augmented Spp viral uptake in HDMVEC (B), MØ (C), LDLr −/− MØ (D), and H9C2 (E) at 2 hpi ( p <0.02 or 0.001).

Article Snippet: Lentiviral vector pLV-mCherry were obtained from Addgene (Watertown, MA, USA).

Techniques: Virus, Binding Assay, Incubation

Journal: bioRxiv

Article Title: The SARS-CoV-2 Spike protein induces long-term transcriptional perturbations of mitochondrial metabolic genes, causes cardiac fibrosis, and reduces myocardial contractile in obese mice

doi: 10.1101/2023.01.05.522853

Figure Lengend Snippet: Cardiac Spp viral accumulation in obese mice. (A) Outline of the animal treatment protocol. The C57BL/6J mice (2 months old) were fed normal chow or HFD. The viral administration was given at 3 months later (5 months old). The animals were remained on the same types of diets until being sacrificed at 2 hpi, 24 hpi, 3 wpi, 6 wpi, and 25 wpi. (B) Quantification of serum levels of total cholesterol, LDL/VLDL, and HDL in HFD mice and the NCF group. # indicates p < 0.01 for intergroup comparison. (C) The relative protein levels of SR-B1 in hearts and adipose tissues in HFD mice and the NCF group by Western blot. Blots from two animals per group were shown. # indicates p < 0.01 in HFD vs NCF group for SR-B1 with the 75Kd MW band (**). (D) Systemic dissemination of Spp virus in HFD mice. Viral load was determined by ratios of viral mCherry levels to housekeeping gene RPS18 levels in each tissue using a real time RT-PCR (Log 10 scale in Y-axis). & indicates statistical significance in HFD group as compared with normal mice group 2 hpi or 24 hpi. Dashed line: the average Spp level in the lungs at 24 hpi. *, indicates statistical significance at 24 hpi as compared with 2 hpi in adipose tissues from normal chow-fed mice. The datasets of various tissues (except adipose tissues) at 2 and 24 hpi from normal-chow-fed mice were adapted from our previous report for a direct comparison to the datasets from HFD mice. (E) Cellular colocalizations of Spike protein of Spp virus in the HFD hearts at 2 and 24 hpi. An anti-Spike S1 subunit antibody (red) is used to recognize Spike protein in Spp. An anti-CD31 or anti-MRC1 antibody (green) is used to stain blood vessels or MØ in heart, respectively. Yellow arrowhead indicates cells positive for both markers. Blue: DAPI staining for nuclei. Scale bar: 20 μm.

Article Snippet: Lentiviral vector pLV-mCherry were obtained from Addgene (Watertown, MA, USA).

Techniques: Comparison, Western Blot, Virus, Quantitative RT-PCR, Staining

Journal: Experimental and Therapeutic Medicine

Article Title: Severe acute respiratory syndrome coronavirus 2 ORF3a induces the expression of ACE2 in oral and pulmonary epithelial cells and the food supplement Vita Deyun ® diminishes this effect

doi: 10.3892/etm.2021.9916

Figure Lengend Snippet: Relative ACE2 expression in cells expressing SARS-CoV-2 ORF3a. (A) A549, DOK and HaCaT cells were transduced for 24 h with lentiviral particles carrying ORF3a from SARS-CoV-2 and were subsequently treated with Vita Deyun solution. The expression of ACE2 was determined by reverse transcription-quantitative PCR, with RPL32 and RPS18 serving as reference genes and the value of untreated cells as the control (2 -ΔΔCp algorithm). The values are presented as the mean ± SD from at least three different experiments. * P<0.05. (B) Data from project GSE152586 of the Gene Expression Omnibus were used to determine variations in ACE2 expression following 48 h of SARS-CoV-2 infection in alveolar spheroids. The values are expressed in fragments per kilobase of exon model per million reads mapped (FPKM), which is a normalized estimation of gene expression based on RNA-seq data. ACE2 , angiotensin-converting enzyme 2; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; ORF3a , open reading frame 3a; RP , ribosomal protein.

Article Snippet: For the transfection assays, Lipofectamine ® 3000 (Invitrogen; Thermo Fisher Scientific, Inc.) was used according to the manufacturer's protocol and utilizing a mix of Lenti-X HT packaging systems (4 µg) and 1 µg of one of the following lentiviral vectors, pLVX-Puro (Takara Bio, Inc.) or the pLVX-EF1alpha-SARS-CoV-2-ORF3a-2xStrep-IRES-Puro vector (cat no. 141383; Addgene, Inc.) ( ).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Infection, RNA Sequencing Assay

Journal: Experimental and Therapeutic Medicine

Article Title: Severe acute respiratory syndrome coronavirus 2 ORF3a induces the expression of ACE2 in oral and pulmonary epithelial cells and the food supplement Vita Deyun ® diminishes this effect

doi: 10.3892/etm.2021.9916

Figure Lengend Snippet: Cytokine mRNA levels in A549 cells constitutively expressing SARS-CoV-2 ORF3a. A model of A549 cells that constitutively expressed the ORF3a were developed and analyzed to determine variations in the expression of ACE2 , IL-6 , IL-6R , IL-18 and TNF-α by reverse transcription-quantitative PCR. The expression is presented as relative expression and was determined using the 2 -ΔΔCp algorithm with RPL32 and RPS18 serving as the reference genes. The values are presented as the mean ± SD from at least three different experiments. * P<0.05. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; ORF3a, open reading frame 3a; ACE2 , angiotensin-converting enzyme 2; IL , interleukin; TNF -α, tumor necrosis factor-α; RP , ribosomal protein.

Article Snippet: For the transfection assays, Lipofectamine ® 3000 (Invitrogen; Thermo Fisher Scientific, Inc.) was used according to the manufacturer's protocol and utilizing a mix of Lenti-X HT packaging systems (4 µg) and 1 µg of one of the following lentiviral vectors, pLVX-Puro (Takara Bio, Inc.) or the pLVX-EF1alpha-SARS-CoV-2-ORF3a-2xStrep-IRES-Puro vector (cat no. 141383; Addgene, Inc.) ( ).

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: Cell reports

Article Title: Activation of GPR44 decreases severity of myeloid leukemia via specific targeting of leukemia initiating stem cells

doi: 10.1016/j.celrep.2023.112794

Figure Lengend Snippet:

Article Snippet: Candidate oligomers for sgRNA were generated by adding respectively the overhang sequences “CACC” and “AAAC” to the 5′ end of forward and reverse oligos to facilitate the cloning of oligomers into the BsmBI sites in the lentiviral transfer vector, pLV hU6-sgRNA hUbC-dCas9-KRAB-T2a-GFP (Addgene plasmid # 71237, a gift from Charles Gersbach).

Techniques: Virus, Recombinant, Enzyme-linked Immunosorbent Assay, Binding Assay, Staining, Modification, Saline, Concentration Assay, Over Expression, Protein Extraction, Membrane, SYBR Green Assay, Bicinchoninic Acid Protein Assay, Protease Inhibitor, CCK-8 Assay, Reverse Transcription, Selection, Plasmid Preparation, Knock-Out, Software, Real-time Polymerase Chain Reaction, Flow Cytometry