Journal: International Journal of Molecular Sciences

Article Title: SARS-CoV-2 ORF7a Protein Impedes Type I Interferon-Activated JAK/STAT Signaling by Interacting with HNRNPA2B1

doi: 10.3390/ijms26125536

Figure Lengend Snippet: ORF7a has no effect on Janus kinases (JAKs). Cells were transfected with EV or ORF7a plasmid for 24 h, then stimulated with IFNα. After 24 h IFN-stimulation, the mRNA ( A ) and protein ( B ) levels of JAK1, JAK2, and TYK2 were evaluated. ( C ) Densitometry analysis showing the relative fold changes of JAK1, JAK2, and TYK2 total protein levels. ( D ) The phosphorylation levels of JAK1 (JAK1-P), JAK2 (JAK2-P), and TYK2 (TYK2-P) after 0.5 h of IFN-stimulation. ( E ) Densitometry analysis showing the relative fold changes of JAK1-P, JAK2-P, and TYK2-P. Relative fold changes are shown after normalization to the corresponding tubulin level. Error bars indicate the standard errors of the results obtained from three independent experiments. Student’s t -test was used for statistical analyses.

Article Snippet: The following primary antibodies were used: STAT1 (Cat# 14994T, CST, Danver, MA, USA, 1:1000), phospho-STAT1 (STAT1-y701) (Cat# ab109457, Abcam, Cambridge, UK, 1:1000), STAT2 (Cat# ab32367, Abcam, Cambridge, UK, 1:5000), phospho-STAT2 (STAT2-y690) (Cat# ab191601, Abcam, Cambridge, UK, 1:1000), β-tubulin (Cat# HC101, TransGen Biotech, Beijing, China, 1:1000), Histone H2A (Cat# 7631, CST, Danver, MA, USA, 1:1000), JAK1 (Cat# ab133666, Abcam, Cambridge, UK, 1:1000), phosphorylated JAK1 (JAK1-P) (Cat# ab138005, Abcam, Cambridge, UK, 1:1000), JAK2 (Cat# ab108596, Abcam, Cambridge, UK, 1:5000), phosphorylated JAK2 (JAK2-P) (Cat# WL02997, Wanlei bio, Shenyang, China, 1:500), TYK2 (Cat# A2128, Abclonal, Wuhan, China, 1:500), phosphorylated TYK2 (TYK2-P) (Cat# bs-3437R, Bioss Inc., Woburn, MA, USA, 1:1000), HNRNPA2B1 (Cat# AY2564, Abways, Beijing, China, 1:500), and Flag (Cat# 66008, Proteintech, Chicago, IL, USA, 1:5000).

Techniques: Transfection, Plasmid Preparation, Phospho-proteomics

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Phosphorylation of IRF3, Tyk2 and STAT1 were assessed by Western blot. The expression of IFN-β and IP-10 mRNA were detected by real-time RT-PCR. n = 6−8 . *P <0.05 compared with control group; # P <0.05 compared with LPS group. $ P <0.05 compared with Ber+LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Phospho-proteomics, Western Blot, Expressing, Quantitative RT-PCR, Control

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Phosphorylation of IRF3, Tyk2 and STAT1 were assessed by Western blot. The expression of IFN-β and IP-10 mRNA were detected by real-time RT-PCR. n = 6−8 . *P <0.05 compared with control group; # P <0.05 compared with LPS group. $ P <0.05 compared with Ber+LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Phospho-proteomics, Western Blot, Expressing, Quantitative RT-PCR, Control

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Spleen was removed 1 h after LPS or normal saline injection, extraction of total protein was analyzed for the presence of total and phosphorylated forms of IκBα, ERK, JNK and p38 MAPK using Western blot, cytoplasmic extracts were subjected to Western blot analysis for determining NF-κB p65 level, nuclear extracts were used to measure the NF-κB activity using NF-κB p65 transcription factor assay kit, and NF-κB activity was normalized to control. n = 5−6 . * P <0.05 compared with control group; # P <0.05 compared with LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Saline, Injection, Extraction, Western Blot, Activity Assay, Transcription Factor Assay, Control

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Mouse peritoneal macrophages were incubated with vehicle, Ber (2.0 µM), Ber (2.0 µM)+Y (5.0 µM) or Y (5.0 µM) for 2 h and then treated with LPS (100 ng/ml) for another 1 h. Whole cell lysates were examined for the presence of total and phosphorylated form of IκBα, ERK, JNK, p38 MAPK and IRF3 using Western blotting. Data in graph are presented as the mean of the ratio of phosphorylated protein to total protein. Nuclear extracts were also used to measure the NF-κB activity using NF-κB p65 transcription factor assay kit, and NF-κB activity was normalized to control macrophages. n = 3−4. *P <0.05 compared with control group; # P <0.05 compared with LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Incubation, Western Blot, Activity Assay, Transcription Factor Assay, Control

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Phosphorylation of IRF3, Tyk2 and STAT1 were assessed by Western blot. The expression of IFN-β and IP-10 mRNA were detected by real-time RT-PCR. n = 6−8 . *P <0.05 compared with control group; # P <0.05 compared with LPS group. $ P <0.05 compared with Ber+LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Phospho-proteomics, Western Blot, Expressing, Quantitative RT-PCR, Control

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Phosphorylation of IRF3, Tyk2 and STAT1 were assessed by Western blot. The expression of IFN-β and IP-10 mRNA were detected by real-time RT-PCR. n = 6−8 . *P <0.05 compared with control group; # P <0.05 compared with LPS group. $ P <0.05 compared with Ber+LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Phospho-proteomics, Western Blot, Expressing, Quantitative RT-PCR, Control

Journal: PLoS ONE

Article Title: Yohimbine Enhances Protection of Berberine against LPS-Induced Mouse Lethality through Multiple Mechanisms

doi: 10.1371/journal.pone.0052863

Figure Lengend Snippet: Mouse peritoneal macrophages were incubated with vehicle, Ber (2.0 µM), Ber (2.0 µM)+Y (5.0 µM) or Y (5.0 µM) for 2 h and then treated with LPS (100 ng/ml) for another 1 h. Whole cell lysates were examined for the presence of total and phosphorylated form of IκBα, ERK, JNK, p38 MAPK and IRF3 using Western blotting. Data in graph are presented as the mean of the ratio of phosphorylated protein to total protein. Nuclear extracts were also used to measure the NF-κB activity using NF-κB p65 transcription factor assay kit, and NF-κB activity was normalized to control macrophages. n = 3−4. *P <0.05 compared with control group; # P <0.05 compared with LPS group.

Article Snippet: Antibodies, including IκBα, ERK, JNK, p38MAPK, NF-κB, IRF3, Tyk2, STAT1, phosphorylated IκBα (p-IκBα), phosphorylated JNK (p-JNK), phosphorylated ERK(p-ERK), phosphorylated p38MAPK (p-p38 MAPK), phosphorylated IRF3(p-IRF3), phosphorylated Tyk2 (p-Tyk2), phosphorylated STAT1(p-STAT1) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and horseradish peroxidase -labeled secondary antibody were from Cell Signaling Technology, Inc. (Danvers, MA, USA).

Techniques: Incubation, Western Blot, Activity Assay, Transcription Factor Assay, Control

Journal: Toxicology and applied pharmacology

Article Title: Physapubescin B enhances the sensitivity of gastric cancer cells to trametinib by inhibiting the STAT3 signaling pathway.

doi: 10.1016/j.taap.2020.115273

Figure Lengend Snippet: Fig. 5. PB treatment stimulates the phosphorylation of EGFR and ERK in GC cells.

Article Snippet: Antibodies against PARP, Caspase 3, β-actin, p-STAT3-Y705, STAT3, XIAP, ERK), p-ERK, JAK1, p-JAK1, JAK2, p-JAK2, JAK3, p-JAK3, TYK2, p-TYK2, EGFR, p-EGFR, GAPDH, were purchased from Cell Signaling Technology Inc. (Boston, MA, USA), antibody against cyclin D1 and c-Myc were purchased from Abcam (Cambridge, UK).

Techniques: Phospho-proteomics