Journal: Journal of cellular biochemistry

Article Title: MicroRNA-145-5p suppresses cell proliferation, migration, and invasion in upper tract urothelial carcinoma by targeting 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase.

doi: 10.1002/jcb.30449

Figure Lengend Snippet: FIGURE 6 5‐aminoimidazole‐4‐carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC) reversed the inhibitory effects of miR‐145‐5p in upper tract urothelial carcinoma (UTUC) cells. (A) miR‐145‐5p inhibited proliferation in BFTC909 and UM‐UC‐14 cells but was restored by ATIC overexpression (N = 3). (B) miR‐145‐5p repressed the migration abilities of BFTC909 and UM‐UC‐14 cells but they were restored by ATIC overexpression (N = 3). (C) miR‐145‐5p suppressed the invasion abilities of BFTC909, UM‐UC‐14, and KMPC3 cells but they were restored by ATIC overexpression (N = 3). (D) Western blot analysis revealed that ATIC, FN1, Slug, cyclin A2, cyclin B1, P57, and IFITM1 levels were restored after the cotransfection of miR‐145‐5p mimics and CMV‐ATIC compared with the levels in BFTC909 cells transfected with miR‐145‐5p alone with α‐tubulin as a reference. (E) Quantification of the protein levels of ATIC, FN1, Slug, cyclin A2, cyclin B1, P57, and IFITM1 from (D) (N = 3). (F) Analysis of miR‐145‐5p, ATIC, IFITM1, CDKN1C, FN1, SNAI2, CCNA2, and CCNB1 expression in UTUC tissues compared with that in adjacent normal tissues using GSE159824 database (N = 2). Data are shown as the mean ± standard deviation; *p < 0.05, **p < 0.01, and ***p < 0.001.

Article Snippet: See the T erm s and C onditions (https://onlinelibrary.w iley.com /term s-and-conditions) on W iley O nline L ibrary for rules of use; O A articles are governed by the applicable C reative C om m ons L icense Technology), fibronectin 1 (FN1) (GTX112794, GeneTex), Slug (NBP2‐52570, Novus), cyclin A2 (#4656, Cell Signaling Technology), cyclin B1 (#4138, Cell Signaling Technology), p57 (NBP1‐89917, Novus), and interferon‐ induced transmembrane 1 (IFITM1) (NBP1‐89345, Novus).

Techniques: Over Expression, Migration, Western Blot, Cotransfection, Transfection, Expressing, Standard Deviation

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: Vero (A), 293 (B), and HeLa (C) cells were incubated with the indicated concentrations of IFN-α for 24 h and then transfected with AiV replicon RNA. At the indicated time points after transfection, cell lysates were harvested, and luciferase activity was measured. Each right panel represents the protein levels of IFITM1, IFITM2, or IFITM3 by immunoblotting for cell lysates harvested from experiments shown in the left panels. Data are the means ±SD of at least three independent experiments. *, P < 0.05; **, P < 0.001; ***, P < 0.0001.

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Incubation, Transfection, Luciferase, Activity Assay, Western Blot

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: (A–C) 293 (A), HeLa (B), or Vero (C) cell lines stably expressing tetracycline (Tet)-inducible IFITM1, IFITM2 or IFITM3 were cultured with or without Tet for 72 h and then transfected with AiV replicon RNA. Cell lysates were harvested at the indicated time points after transfection and then assayed for luciferase activity. The peak activity obtained for cells in the absence of Tet was taken as 100%. Tet-induced IFITM1, IFITM2, or IFITM3 protein expression in each cell line was detected by Western blotting (bottom panels of (A) and right panel of (B) or right panel of (C)). (D) HeLa cells were transfected with the control, IFITM1, IFITM2, or IFITM3 siRNA for 72 h and then transfected with replicon RNA. At the indicated time points after replicon RNA transfection, cell lysates were harvested and subjected to the luciferase assay. The maximum value obtained for cells treated with control siRNA was taken as 100%. (E) IFITM1, IFITM2, or IFITM3 knockdown was confirmed by Western blotting. (F) Cell viability was determined by the CellTiter-Glo assay. Data are the means ± SD of at least three independent experiments. *, P < 0.05; **, P < 0.001; ***, P < 0.0001.

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Stable Transfection, Expressing, Cell Culture, Transfection, Luciferase, Activity Assay, Western Blot, Control, Knockdown, Glo Assay

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: (A) A mammalian two-hybrid assay was performed by transfecting Vero cells with the indicated combinations of pACT and pBIND plasmids along with the pG5luc firefly luciferase reporter plasmid. At 48 h after transfection, the normalized firefly luciferase activity (firefly luciferase activity/Renilla luciferase activity) was measured and represented as fold of the control activity, which was obtained by combination with the empty pACT or empty pBIND plasmid. Data are the mean ± SD of at least three independent experiments. (B) 293T cells were transfected for 24 h with FLAG-tagged IFITM1 and HA-tagged 3Cm, 2B, 2BC, 2C, 3A, or 3AB, or HA-tagged IFITM1 and FLAG-tagged TGN46 expression plasmids, as indicated, followed by coimmunoprecipitation (IP) with an anti-FLAG, -HA or control IgG antibody. The resulting immunoprecipitates and whole-cell lysates were subjected to immunoblotting (IB) with anti-FLAG and anti-HA antibodies. (C) Vero cells were transfected with FLAG-IFITM1. At 24 h after post-transfection, the cells were fixed and double stained with anti-IFITM and anti-EEA1, anti-CD63, or anti-LBPA antibodies, as indicated. Pearson correlation coefficient analyses for data were obtained from ≥10 cells. Correlation coefficients are presented as the mean and standard deviation. (D) Vero cells was transfected with FLAG-IFITM1 and HA-2B, HA-2BC, HA-2C, HA-3A, or HA-3AB. At 24 h after transfection, cells were fixed and stained with anti-FLAG and anti-HA antibodies. Bars, 4 μm. Pearson correlation coefficient analyses for data were obtained from 4–8 cells.

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Two Hybrid Assay, Luciferase, Plasmid Preparation, Transfection, Activity Assay, Control, Expressing, Western Blot, Staining, Standard Deviation

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: (A) A mammalian two-hybrid analysis was carried out to examine interactions between IFITM1 and ACBD3, PI4KB, OSBP, or CERT, and the results are shown as described in . Data are the mean ± SD of at least three independent experiments. (B) HA-tagged IFITM1 and FLAG-tagged ACBD3, PI4KB, OSBP, or CERT were cotransfected into 293T cells, and the cell lysates were subjected to immunoprecipitation with anti-HA antibody or control IgG. The resulting immunocomplexes and whole-cell lysates were detected by anti-FLAG and anti-HA antibodies. (C) Vero cells were transfected with FLAG-IFITM1. At 24 h, the cells were labeled with anti-FLAG and anti-ACBD3 (top), anti-PI4KB (middle), or anti-OSBP (bottom) antibodies. (D) Vero IFITM1 cells were incubated with Tet (−) or Tet (+) for 72 h, and the cells were fixed and double stained with the indicated antibodies. (E) Vero cells were transfected with HA-IFITM1. At 24 h, the cells were labeled using anti-HA, anti-OSBP and anti-EEA1, or anti-CD63 antibodies. Bars, 4 μm. Pearson correlation coefficient analyses for data were obtained from ≥10 cells. Correlation coefficients are presented as the mean and standard deviation (C-E).

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Immunoprecipitation, Control, Transfection, Labeling, Incubation, Staining, Standard Deviation

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: (A–F) Vero cells were pretreated with DMSO (as the control) or the indicated concentrations of 25-HC (A), U18666A (B), or BF738735 (C) for 24 h and then transfected with AiV replicon RNA. At 10 h after transfection, the cells were analyzed for luciferase activity. Data were normalized to the DMSO treatment, and cell viability was scored. (D–F) Vero IFITM1 cells were incubated with or without Tet for 48 h and then treated with mock (–) or 1 μM of 25-HC (D), 1.5 μM of U18666A (E), or 150 nM of BF738735 (F). After 24 h, the cells were transfected with AiV replicon RNA, and luciferase activity was determined at 10 h after transfection. (G) Cell viability was measured in parallel. The maximum value obtained for Tet (–) drug-untreated cells was taken as 100%. Data are the mean ± SD of at least three independent experiments. *, P < 0.05; **, P < 0.001; ***, P < 0.0001.

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Control, Transfection, Luciferase, Activity Assay, Incubation

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: (A–E) Vero-IFITM1 cells were incubated with or without Tet for 48 h and then treated with mock (A and D), 1 μM 25-HC (B), 1.5 μM U18666A (C), or 150 nM BF738735 (E), followed by pCMV-polyprotein transfection. At 24 h after transfection, the cells were fixed and stained with filipin III (A–C) or anti-PI4P (D and E), anti-IFITM1 and anti-2B antibodies. Bars, 4 μm. Pearson’s correlation coefficient was measured for individual cells.

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Incubation, Transfection, Staining

Journal: PLOS Pathogens

Article Title: IFITM1 enhances nonenveloped viral RNA replication by facilitating cholesterol transport to the Golgi

doi: 10.1371/journal.ppat.1011383

Figure Lengend Snippet: (A and B) Vero-IFITM1 cells were incubated with or without Tet for 48 h, and then treated with 3 μM AY9944. After 24 h, the cells were fixed and stained with filipin III and the indicated antibodies. Bars, 4 μm. Pearson correlation coefficient analyses for data were obtained from ≥10 cells. Correlation coefficients are presented as the mean and standard deviation.

Article Snippet: The coding regions of IFITM1, IFITM2, and IFITM3 were amplified by PCR using primer pairs containing MluI – EcoRV sites from cDNA clones (OriGene; IFITM1: RC201617, IFITM2: RC202067, IFITM3: RC201635), respectively, and then inserted into the same sites of pACT and pBIND, resulting in pACT-IFITM1, pACT-IFITM2, pACT-IFITM3, pBIND-IFITM1, pBIND-IFITM2, and pBIND-IFITM3.

Techniques: Incubation, Staining, Standard Deviation

Journal: bioRxiv

Article Title: The zinc metalloprotease ZMPSTE24 binds a distinct topological isoform of the tail-anchored protein IFITM3

doi: 10.64898/2026.02.27.708584

Figure Lengend Snippet: (A) Assessing loss of catalytic activity for ZMPSTE24 HExxH…E mutants. Flag-tagged wild-type and mutant ZMPSTE24 plasmids were co-transfected with a myc-tagged prelamin A plasmid (myc-LMNA (431-664) ) in ZMPSTE24 knockout HEK293 cells to evaluate ZMPSTE24 proteolytic activity. Flag-ZMPSTE24, prelamin A and its cleaved product, mature lamin A, were detected by Western blotting and are indicated. All active site mutants block cleavage of the prelamin A construct. (B) Co-IP’s of endogenous IFITMs by ZMPSTE24 and ZMPSTE24 E336A . HEK293 cells transfected with vector, Flag-ZMPSTE24 or Flag-ZMPSTE24 E336A were induced with 150 units of interferon β for 18 hours prior to immunoprecipitation with anti-Flag agarose. Whole cell lysate (WCL) and IP’d proteins were resolved by SDS-PAGE analyzed by Western blotting with anti-Flag (ZMPSTE24), anti-IFITM1, IFITM2 or IFITM3 antibodies. (C) Comparing co-IPs of IFITMs by Flag-ZMPSTE24 E336A and Flag-IFITM3. Cells transfected with vector, Flag-ZMPSTE24, Flag-ZMPSTE24 E336A or Flag-IFITM3 and the indicated myc-tagged IFITM proteins were subjected to co-IP as described. Positions of ZMPSTE24 and IFITM proteins are indicated. Lanes 9-12 were used to make . (D) Flag-tagged IFITM3 co-IPs myc-ZMPSTE24 E336A more efficiently than wild-type myc-ZMPSTE24. HEK293 cells were transfected with Flag-IFITM3 and myc-ZMPSTE24 or myc-ZMPSTE24 E336A . Proteins were co-IP’d with anti-Flag agarose prior to SDS-PAGE and Western blotting. Positions of Flag-IFITM3 and myc-ZMPSTE24 proteins are indicated.

Article Snippet: Anti-IFITM1 (Proteintech, 11727-3-AP) was used at 1:1000.

Techniques: Activity Assay, Mutagenesis, Transfection, Plasmid Preparation, Knock-Out, Western Blot, Blocking Assay, Construct, Immunoprecipitation, SDS Page, Co-Immunoprecipitation Assay

Journal: Journal of Virology

Article Title: ISG56 and IFITM1 Proteins Inhibit Hepatitis C Virus Replication

doi: 10.1128/jvi.05633-11

Figure Lengend Snippet: FIG. 1. HCV infection inhibits IFN--induced ISG56 and IFITM1 expression. IHH were treated with IFN alone, infected with HCV, or pretreated with IFN and infected with HCV. Total RNA was isolated 96 h postinfection, and ISG56 (A), IFITM1 (B), or IFITM2 (C) mRNA was measured by real-time RT-PCR. GAPDH was used as an internal control for normalization. Huh7 cells were similarly treated, and the mRNA expression status of ISG56 (D), IFITM1 (E), or IFITM2 (F) is shown. The fold changes of mRNA are presented after normalizing data to a mock-infected control and arbitrarily set as 1. The results are presented as means from four independent experiments with standard errors.

Article Snippet: Cells were transfected with 0.6 g of a pCDNA3-Myc-tagged open reading frame (ORF) clone of human ISG56 and a pCMV6-Myc-DDKtagged ORF clone of human IFITM1 (Origene) using Lipofectamine (Invitrogen, CA).

Techniques: Infection, Expressing, Isolation, Quantitative RT-PCR, Control

Journal: Journal of Virology

Article Title: ISG56 and IFITM1 Proteins Inhibit Hepatitis C Virus Replication

doi: 10.1128/jvi.05633-11

Figure Lengend Snippet: FIG. 2. Inhibition of IFN--induced IFITM1 expression by HCV. IHH were mock treated, treated with IFN as a positive control, infected with HCV genotype 2a, or pretreated with IFN and infected with HCV. (A) Cells were fixed after 96 h of infection and stained with DAPI (blue), IFITM1 antibody (red), and HCV NS3 antibody (green). Merged photographs are shown. (B) Huh7 cells were treated and stained as described for panel A. A merged photograph from a randomly selected field is shown.

Article Snippet: Cells were transfected with 0.6 g of a pCDNA3-Myc-tagged open reading frame (ORF) clone of human ISG56 and a pCMV6-Myc-DDKtagged ORF clone of human IFITM1 (Origene) using Lipofectamine (Invitrogen, CA).

Techniques: Inhibition, Expressing, Positive Control, Infection, Staining

Journal: Journal of Virology

Article Title: ISG56 and IFITM1 Proteins Inhibit Hepatitis C Virus Replication

doi: 10.1128/jvi.05633-11

Figure Lengend Snippet: FIG. 3. Transient expression of ISG56 and IFITM1 inhibits HCV growth. IHH were infected with HCV genotype 2a and transfected with vector DNA, ISG56, or IFITM1 2 days postinfection. (A) HCV replication was measured after 48 h of transfection from intracellular RNA by RT-qPCR. The results are shown as means from four independent experiments with standard errors (P 0.006 for ISG56 and P 0.001 for IFITM1). (B) HCV infectivity was measured from culture supernatants of virus-infected IHH at serial dilutions and are expressed as a percentage of that in vector-transfected control cells (P 0.0001). Western blot analysis was performed for the expression of ISG56 (C) or IFITM1 (D) in IHH transfected with the respective plasmid DNAs, empty vector, or mock-treated cells using specific antibodies. The blot was reprobed with an antibody to actin for comparisons of protein loads. (E) The transient expression of ISG56 or IFITM1 also suppresses HCV replication in Huh7 cells. The results are shown as means from three independent experiments with standard errors (P 0.001 for ISG56 or IFITM1). Huh7 cells were stably transfected with vector, ISG56, and IFITM1. (F) Pooled stable cell lines were incubated with VSV/HCV pseudotype virus for 48 h, and plaque assay was performed to examine HCV entry. The results are shown as means from three independent experiments with standard errors.

Article Snippet: Cells were transfected with 0.6 g of a pCDNA3-Myc-tagged open reading frame (ORF) clone of human ISG56 and a pCMV6-Myc-DDKtagged ORF clone of human IFITM1 (Origene) using Lipofectamine (Invitrogen, CA).

Techniques: Expressing, Infection, Transfection, Plasmid Preparation, Quantitative RT-PCR, Virus, Control, Western Blot, Stable Transfection, Incubation, Plaque Assay

Journal: Journal of Virology

Article Title: ISG56 and IFITM1 Proteins Inhibit Hepatitis C Virus Replication

doi: 10.1128/jvi.05633-11

Figure Lengend Snippet: FIG. 4. Transient expression of ISG56 or IFITM1 suppresses subgenomic HCV RNA replication. Huh7 cells harboring subgenomic replicon Con1 (A) or JFH1 (B) were transfected with vector DNA, ISG56, or IFITM1. Cellular RNA was extracted 48 h posttransfection, and viral RNA was analyzed by RT-qPCR. The results are shown as means from three independent experiments with standard errors (P 0.003 for ISG56 and P 0.001 for IFITM1). Huh7 cells harboring subgenomic replicon Con1 (C) or JFH1 (D) also were transfected with plasmid DNA containing CMV promoter-driven luciferase or IFITM2 as a control. Cellular RNA was extracted 48 h posttransfection, and viral RNA was analyzed by real-time RT-PCR. The results shown are from three independent experiments.

Article Snippet: Cells were transfected with 0.6 g of a pCDNA3-Myc-tagged open reading frame (ORF) clone of human ISG56 and a pCMV6-Myc-DDKtagged ORF clone of human IFITM1 (Origene) using Lipofectamine (Invitrogen, CA).

Techniques: Expressing, Transfection, Plasmid Preparation, Quantitative RT-PCR, Luciferase, Control

Journal: Journal of Virology

Article Title: ISG56 and IFITM1 Proteins Inhibit Hepatitis C Virus Replication

doi: 10.1128/jvi.05633-11

Figure Lengend Snippet: FIG. 5. Knockdown of ISG56 or IFITM1 enhances HCV RNA replication. IHH were transfected with control siRNA, ISG56 siRNA, or IFITM1 siRNA and subsequently infected with HCV genotype 2a. Total cellular RNA was extracted 96 h postinfection, and ISG56 (A), IFITM1 (C), and intracellular HCV RNA (B and D) were analyzed by RT-qPCR. The results are shown as means from three independent experiments with standard errors (P 0.001). The introduction of plasmid DNA encoding ISG56 or IFITM1 reduced the siRNA-mediated interference of HCV replication (B and D). IHH also were transfected with control siRNA or were mock treated and then infected with HCV genotype 2a to ascertain nonspecific inhibitory effects. (E) Total RNA was extracted 96 h postinfection, and HCV RNA was analyzed by RT-qPCR. The results shown are from two independent experiments with three replicates.

Article Snippet: Cells were transfected with 0.6 g of a pCDNA3-Myc-tagged open reading frame (ORF) clone of human ISG56 and a pCMV6-Myc-DDKtagged ORF clone of human IFITM1 (Origene) using Lipofectamine (Invitrogen, CA).

Techniques: Knockdown, Transfection, Control, Infection, Quantitative RT-PCR, Plasmid Preparation

Journal: Virology journal

Article Title: Investigation of antiviral state mediated by interferon-inducible transmembrane protein 1 induced by H9N2 virus and inactivated viral particle in human endothelial cells.

doi: 10.1186/s12985-017-0875-5

Figure Lengend Snippet: Fig. 1 Quantitation of IFITM1 induced by H9N2 virus or inactivated viral particle in HUVECs and BEAS-2Bs. HUVECs and BEAS-2Bs were treated with H9N2 virus (i.e., Virus) or viral particle (i.e., Particle) at MOI of 5. Cells used for RT-PCR and western blot analysis were collected at different time points. a The expression of IFITM1 at mRNA level in HUVECs. b, c The expression of IFITM1 at protein level in HUVECs, and the relative protein level to β-actin. d The expression of IFITM1 at mRNA level in BEAS-2Bs. e, f The expression of IFITM1 at protein level in BEAS-2Bs, and the relative protein level to β-actin. * means particle group and virus group compared with control group (*, P < 0.05. **, P < 0.01, ANOVA). # means particle group compared with H9N2 virus group (#, P < 0.05. ##, P < 0.01, ANOVA)

Article Snippet: IFITM1 specific and control siRNA used in the study were purchased from Santa Cruz Biotechnology, and the siRNA for human cells is a pool of 3 target-specific 19–25 nt siRNAs designed to knock down gene expression.

Techniques: Quantitation Assay, Virus, Reverse Transcription Polymerase Chain Reaction, Western Blot, Expressing, Control

Journal: Virology journal

Article Title: Investigation of antiviral state mediated by interferon-inducible transmembrane protein 1 induced by H9N2 virus and inactivated viral particle in human endothelial cells.

doi: 10.1186/s12985-017-0875-5

Figure Lengend Snippet: Fig. 4 Expression of IFITM1 at mRNA level in HUVECs and BEAS-2Bs treated with hemagglutinin (HA) and neuraminidase (NA). HUVECs and BEAS- 2Bs were incubated with HA or NA at concentrations of 0.1, 1 μg/mL. Cells used for RT-PCR analysis were collected at 6, 12, 24 h. a The mRNA levels of IFITM1 induced by HA in HUVECs. b The mRNA levels of IFITM1 induced by NA in HUVECs. c The mRNA levels of IFITM1 induced by HA in BEAS-2Bs. d The mRNA levels of IFITM1 induced by NA in BEAS-2Bs

Article Snippet: IFITM1 specific and control siRNA used in the study were purchased from Santa Cruz Biotechnology, and the siRNA for human cells is a pool of 3 target-specific 19–25 nt siRNAs designed to knock down gene expression.

Techniques: Expressing, Incubation, Reverse Transcription Polymerase Chain Reaction

Journal: Virology journal

Article Title: Investigation of antiviral state mediated by interferon-inducible transmembrane protein 1 induced by H9N2 virus and inactivated viral particle in human endothelial cells.

doi: 10.1186/s12985-017-0875-5

Figure Lengend Snippet: Fig. 6 Antiviral activity of IFITM1 induced by H9N2 virus infection in HUVECs and BEAS-2Bs. HUVECs and BEAS-2Bs were infected with H9N2 virus at MOI of 5 and incubated for 1 h, then cells were transfected with control siRNA or IFITM1 specific siRNA for 36 h. Virus titer in each group was detected using plaque assay at 36 h postinfection. The effect of IFIMT1 specific siRNA on IFITM1 expression was detected using western blot. a IFITM1 protein level after transfected with siRNA in HUVECs. b IFITM1 protein level after transfected with siRNA in BEAS-2Bs. c Virus titers in HUVECs transfected with control siRNA or IFITM1 specific siRNA. Compared to virus group (control siRNA), the virus titer in virus + siRNA group (IFITM1 specific siRNA) was increased by 13.1 ± 2.4% (P > 0.05, t-test). d Virus titers in BEAS-2Bs transfected with control siRNA or IFITM1 specific siRNA. Compared to virus group (control siRNA), the virus titer in virus + siRNA group (specific siRNA) was increased by 9.7% ± 3.8% (P > 0.05, t-test)

Article Snippet: IFITM1 specific and control siRNA used in the study were purchased from Santa Cruz Biotechnology, and the siRNA for human cells is a pool of 3 target-specific 19–25 nt siRNAs designed to knock down gene expression.

Techniques: Activity Assay, Virus, Infection, Incubation, Transfection, Control, Plaque Assay, Expressing, Western Blot

Journal: Virology journal

Article Title: Investigation of antiviral state mediated by interferon-inducible transmembrane protein 1 induced by H9N2 virus and inactivated viral particle in human endothelial cells.

doi: 10.1186/s12985-017-0875-5

Figure Lengend Snippet: Fig. 7 Antiviral activity of IFITM1 induced by viral particle inoculation in HUVECs and BEAS-2Bs. HUVECs and BEAS-2Bs were inoculated with viral particle at MOI of 5 and incubated for 1 h, then cells were transfected with control siRNA or IFITM1 specific siRNA for 36 h before infected with H9N2 virus at MOI of 5. Virus titer of each group was detected by plaque assay at 36 h postinfection. The effect of IFIMT1 specific siRNA on IFITM1 expression was detected using western blot. a IFITM1 protein level after transfected with siRNA in HUVECs. b IFITM1 protein level after transfected with siRNA in BEAS-2Bs. c Virus titers in HUVECs transfected with control siRNA or IFITM1 specific siRNA. Compared to particle group (control siRNA), the virus titer in particle + siRNA group (IFITM1 specific siRNA) was increased by 60.5 ± 10.7% (P < 0.05, t-test). d Virus titers in BEAS-2Bs transfected with control siRNA or IFITM1 specific siRNA. Compared to particle group (control siRNA), the virus titer in particle + siRNA group (specific siRNA) was increased by 12.9 ± 3.6% (P > 0.05, t-test). * means particle group compared with particle + siRNA group (*, P < 0.05, t-test)

Article Snippet: IFITM1 specific and control siRNA used in the study were purchased from Santa Cruz Biotechnology, and the siRNA for human cells is a pool of 3 target-specific 19–25 nt siRNAs designed to knock down gene expression.

Techniques: Activity Assay, Incubation, Transfection, Control, Infection, Virus, Plaque Assay, Expressing, Western Blot

Journal: Virology journal

Article Title: Investigation of antiviral state mediated by interferon-inducible transmembrane protein 1 induced by H9N2 virus and inactivated viral particle in human endothelial cells.

doi: 10.1186/s12985-017-0875-5

Figure Lengend Snippet: Fig. 8 Overexpression of IFITM1 significantly reduced virus titers in HUVECs and BEAS-2Bs. HUVECs and BEAS-2Bs were transfected with control plasmid (Control) or IFITM1 CRISPR activation plasmid (plasmid) for 36 h, then cells were infected with H9N2 virus at MOI of 5. Virus titer of each group was detected by plaque assay at 36 h postinfection. The overexpression of IFIMT1 was detected by western blot. a IFITM1 protein level after transfected with plasmid in BEAS-2Bs. b IFITM1 protein level after transfected with plasmid in HUVECs. c Virus titers in BEAS-2Bs transfected with control plasmid or IFITM1 CRISPR activation plasmid. Compared to control group (control plasmid), the virus titer in plasmid group (IFITM1 CRISPR activation plasmid) was decreased by 55.72.5 ± 7.53% (P < 0.01, t-test). d Virus titers in HUVECs transfected with control plasmid or IFITM1 CRISPR activation plasmid. Compared to control group (control plasmid), the virus titer in plasmid group (IFITM1 CRISPR activation plasmid) was decreased by 52.76 ± 1.02% (P < 0.01, t-test). * means control group compared with plasmid group (**, P < 0.01, t-test)

Article Snippet: IFITM1 specific and control siRNA used in the study were purchased from Santa Cruz Biotechnology, and the siRNA for human cells is a pool of 3 target-specific 19–25 nt siRNAs designed to knock down gene expression.

Techniques: Over Expression, Virus, Transfection, Control, Plasmid Preparation, CRISPR, Activation Assay, Infection, Plaque Assay, Western Blot