Journal: Journal of Virology

Article Title: NFAT4 Is Required for JC Virus Infection of Glial Cells

doi: 10.1128/jvi.01456-06

Figure Lengend Snippet: FIG. 2. NFAT4 is functionally active in glial cells. (a) NFAT expression in SVG-A and U-87MG whole-cell lysates. NFATs 1 to 4 were detected using antibodies (NFATc1 7A6, NFATc2 4G6-G5, NFATc3 F-1, and NFATc4 H-74; Santa Cruz Biotech Inc.) diluted 1:200. Ramos cell lysates were used as a positive control. (b) Luciferase reporter gene assays using an NFAT-responsive reporter construct were used to compare NFAT activity in SVG-A cells and U-87MG cells in the presence or absence of 292 mg/liter glutamate. A control construct that lacked the NFAT binding site was used to measure basal transcriptional activity. (c) The NFAT reporter construct was cotransfected with either a control construct ( SV40 T-Ag) or a construct expressing the SV40 large T-Ag ( SV40 T-Ag).

Article Snippet: Ramos cell lysates were purchased from Santa Cruz Biotech Inc. Whole-cell extracts were separated on Tris-HCl-ready gels (Bio-Rad).

Techniques: Expressing, Positive Control, Luciferase, Construct, Activity Assay, Control, Binding Assay

Journal: Cancer Research

Article Title: Nuclear Factor of Activated T-cell Activity Is Associated with Metastatic Capacity in Colon Cancer

doi: 10.1158/0008-5472.can-14-1592

Figure Lengend Snippet: Figure 2. Inhibition of NFATc1 in MC38Met cells. Effect of control RNAi (siScr) or NFATc1-specific RNAi on specific NFAT mRNA species: NFATc1 (A), NFATc2 (B), NFATc3 (C). D, the effect of NFATc1 siRNA versus scrambled control si-Scr on NFAT family protein levels in MC38Met cells, and steady state levels in both MC38Par and MC38Met cells with Ramos cell lysate positive control in the right hand lane. E, relative rates of invasion for cells shown in A–D . , P < 0.0005; , P < 0.00005.

Article Snippet: Ramos cell (Burkitt lymphoma, B lymphocytes) lysate (Santa Cruz Biotechnology) was used as positive control.

Techniques: Inhibition, Control, Positive Control

Journal: Cancer Research

Article Title: Nuclear Factor of Activated T-cell Activity Is Associated with Metastatic Capacity in Colon Cancer

doi: 10.1158/0008-5472.can-14-1592

Figure Lengend Snippet: Figure 7. Knockdown of NFATc1 in MC38Met cells decreases tumor incidence and liver metastases. A, Western blot analysis showing NFAT proteins in MC38MetþshCtrl and MC38MetþshNFATc1 cell lines. b-actin was used as a loading control and Ramos cell extract as positive control. B, analysis of NFATc1 specific mRNA in cells shown in A. C, rates of trans-endothelial invasion for cells are shown in A and B. Individual replicate wells from a representative experiment are plotted with the mean and the SEM (bars and whiskers). D, representative (n ¼ 14-15 mice/group) bioluminescence image corresponding day 21. E, summary- of bioluminescence signal quantified at day 21. F, summary incidence of liver metastases. G, liver metastases measured by liver weight to body weight ratio at 21 days postinjection. , P < 0.03; , P < 0.001; , P < 0.0001.

Article Snippet: Ramos cell (Burkitt lymphoma, B lymphocytes) lysate (Santa Cruz Biotechnology) was used as positive control.

Techniques: Knockdown, Western Blot, Control, Positive Control

Journal: Biology of reproduction

Article Title: Members of the Toll-like receptor family of innate immunity pattern-recognition receptors are abundant in the male rat reproductive tract.

doi: 10.1095/biolreprod.106.059410

Figure Lengend Snippet: FIG. 2. Immunoblot analysis of TLR proteins in male reproductive organs. Aliquots (100 lg) of cytoplasmic protein extracts were separated by PAGE, electroblotted, and blots probed with anti-TLR antibodies followed by enhanced chemiluminescence detection using Pierce Super- Signal West Pico or Pierce SuperSignal West Femto (*F) substrate. Representative results are shown (n ¼ 3–5 rats). þ, control rat tissue extracts from spleen, TLRs 1–7; lung, TLR8; and small intestine, TLR9. þ Ext, positive-control whole cell lysates used were Raw 264 Abelson transformed macrophages (TLRs 1, 2–6, and 8–10); SW480 colorectal adenocarcinoma (TLR2); Daudi cell extract (TLR7); Ramos cell lysate (TLR10) and mouse heart whole cell lysate (TLR11). Blots were stripped and reprobed with anti-actin monoclonal antibody to detect actin as a loading control. Representative results are shown (n ¼ 3–5 rats) for blots exposed to film for the same length of time when using equivalent chemiluminescent substrate.

Article Snippet: Mouse heart tissue lysate (40102) and Ramos cell lysate (40175) was from Imgenex.

Techniques: Western Blot, Control, Positive Control, Transformation Assay

Journal: Nature Communications

Article Title: O -GlcNAcylation is required for B cell homeostasis and antibody responses

doi: 10.1038/s41467-017-01677-z

Figure Lengend Snippet: O -GlcNAcylation of Lyn at S19 is required for the interaction of Syk. a Lysates prepared from Ctrl and B-KO splenic B cells stimulated with anti-IgM for 5 min were subjected to immunoprecipitation (IP) with anti-Syk or control rabbit IgG (rIgG) antibody, followed by immunoblotting (IB) with the indicated antibodies. b Protein lysates from membrane fractions of anti-IgM stimulated Ctrl and B-KO splenic B cells were subjected to IP with anti-Syk or rIgG antibody, followed by IB with indicated antibodies. Calnexin served as a loading control for membrane fraction. c Mapping the O -GlcNAc site on S19 of mouse Lyn by using ETD fragmentation. The ions, z 6 + ( m/z 712.4188), z 7 + ( m/z 1002.5387), c 11 + ( m/z 1331.6101), and c 12 + ( m/z 1622.7351) indicated that S19 carried a GlcNAc moiety. d Lysates from 293 T cells overexpressing the vector control (Ve), Flag-EGFP-tagged WT, SA or SE Lyn were subjected to a pull-down assay using sWGA agarose beads, followed by IB with an anti-Flag antibody. e Lysates from 293T cells transfected with Flag-tagged WT, SA, or SE Lyn along with myc-tagged Syk were subjected to IP with anti-Flag antibody, followed by IB with the indicated antibodies. f Anti-Flag immunoprecipitates from Ramos B cells transfected with vector expressing Flag-EGFP-WT, -SA or -SE Lyn and stimulated with anti-IgM (25 µg ml −1 ) for 5 min were subjected to IB with indicated antibodies. In a , b , d , e and f , the representative data from one of at least 2–3 experimental repeats are shown. Quantification of band intensity is indicated in a , b , d , e and f

Article Snippet: Human Ramos B cells were purchased from Bioresource Collection and Research Center, Taiwan (BCRC) (BC-60252) and were cultured in RPMI 1640 medium (Life Technologies) containing 10% FBS, 100 U ml −1 penicillin and 100 μg ml −1 streptomycin (Life Technologies) at 37 °C with 5% CO 2 .

Techniques: Immunoprecipitation, Western Blot, Plasmid Preparation, Pull Down Assay, Transfection, Expressing

Journal: Leukemia

Article Title: Transient stabilization, rather than inhibition of MYC amplifies extrinsic apoptosis and therapeutic responses in refractory B-cell lymphoma

doi: 10.1038/s41375-019-0454-4

Figure Lengend Snippet: A) Model of Myc protein regulation by GSK-3β. B) Myc wild type (left) & Thr58 mutant (right) Burkitt lymphoma cell lines were treated with 3 μM CHIR99021 for a 6 hour time course. Western blotting was performed for β-catenin, Myc, MycThr58 phosphorylation and actin control, with Myc quantification using ImageJ below. C) Ramos cells were treated with DMSO or 3 μM CHIR99021 and increasing concentrations of doxorubicin for 72 hours. Cell survival was assessed as in Fig. 1B).

Article Snippet: P493-6 cells and Burkitt lymphoma cell lines Ramos, Daudi, Raji, and Mutul were acquired from Drs. Chi Dang and Riccardo Dalla-Favera.

Techniques: Mutagenesis, Western Blot, Phospho-proteomics, Control

Journal: Leukemia

Article Title: Transient stabilization, rather than inhibition of MYC amplifies extrinsic apoptosis and therapeutic responses in refractory B-cell lymphoma

doi: 10.1038/s41375-019-0454-4

Figure Lengend Snippet: A) Volcano plot of KEGG apoptosis genes derived from RNA-Seq data on Ramos cells treated for 3 hours with 3 μM CHIR99021 in biological triplicates. Key apoptotic genes are labeled, with genes in blue being down-regulated, genes in red being up-regulated, and genes in green having no significant changes. B) qRT-PCR expression analysis for known Myc-dependent intrinsic/mitochondrial apoptotic factors was performed on Ramos cells treated for 2 hours with DMSO (blue bars) or 3 μM CHIR99021 (red bars) followed by a 6 hour time course of .25 μM doxorubicin. C) qRT-PCR expression analysis for extrinsic apoptosis factors was performed on Ramos cells treated as in B). Notable changes are indicated with yellow arrows.

Article Snippet: P493-6 cells and Burkitt lymphoma cell lines Ramos, Daudi, Raji, and Mutul were acquired from Drs. Chi Dang and Riccardo Dalla-Favera.

Techniques: Derivative Assay, RNA Sequencing, Labeling, Quantitative RT-PCR, Expressing

Journal: Leukemia

Article Title: Transient stabilization, rather than inhibition of MYC amplifies extrinsic apoptosis and therapeutic responses in refractory B-cell lymphoma

doi: 10.1038/s41375-019-0454-4

Figure Lengend Snippet: A) Ramos cells expressing an empty vector construct or the Bcl-2 construct were treated for 2 hours with DMSO or 3 μM CHIR99021 followed by a 6.5 hour time course of .25 μM doxorubicin. Western blotting was performed for Myc, MycThr58 phosphorylation, cleaved PARP, and Bcl-2. B) Ramos cells expressing an empty vector construct or FLIP construct were treated as in A). Western blotting was performed for markers of GSK3-β inhibition, cell death, and FLIP expression.

Article Snippet: P493-6 cells and Burkitt lymphoma cell lines Ramos, Daudi, Raji, and Mutul were acquired from Drs. Chi Dang and Riccardo Dalla-Favera.

Techniques: Expressing, Plasmid Preparation, Construct, Western Blot, Phospho-proteomics, Inhibition

Journal: Leukemia

Article Title: Transient stabilization, rather than inhibition of MYC amplifies extrinsic apoptosis and therapeutic responses in refractory B-cell lymphoma

doi: 10.1038/s41375-019-0454-4

Figure Lengend Snippet: A) Ramos derivative cell lines with scrambled gRNA, DR4 gRNA, and DR5 gRNA were treated and cell survival assessed as in Fig. 2C. B) and C) Ramos cells were treated with DMSO or 3 μM CHIR99021 and increasing concentrations of TRAIL (B) or mapatumumab (C) for 72 hours and analyzed as in A).

Article Snippet: P493-6 cells and Burkitt lymphoma cell lines Ramos, Daudi, Raji, and Mutul were acquired from Drs. Chi Dang and Riccardo Dalla-Favera.

Techniques: