Journal: The Journal of Nutrition, Health & Aging

Article Title: Adipokine pathways are altered in hippocampus of an experimental mouse model of Alzheimer's disease

doi: 10.1007/s12603-014-0574-5

Figure Lengend Snippet: mRNA expression profile (n=5-8) of stat1, stat3, stat5a, stat5b, K-ras, sos1, socs1, socs2 and socs3 in hippocampal extracts of 3 and 6 months-old wild-type and APP/PS1 mice. (Statistical analysis was performed with one-way ANOVA, with Tukey’s post-hoc test, where * denotes p < 0.05, and with the student’s t-test, where $ denotes p < 0.05)

Article Snippet: Socs2 , suppressor of cytokine signaling 2 , 216233 , Mm00850544_g1 , .

Techniques: Expressing

Journal: British journal of cancer

Article Title: Identification of SOCS2 and SOCS6 as biomarkers in human colorectal cancer.

doi: 10.1038/bjc.2014.377

Figure Lengend Snippet: Figure 2. SOCS2 and SOCS6 are potential biomarkers in human CRC. ROC curves with corresponding AUC values for (A) SOCS2 and (B) SOCS6 when classifying CRC patients and healthy donors. All grades of CRC were pooled. Distributions of gene expression values for healthy and CRC patients are shown in the insets.

Article Snippet: Membranes were blocked with 10% milk for 1 h at room temperature and were then incubated overnight at 41C with antibodies directed against SOCS2 (1 : 1000, OriGene Technologies, Uden, Netherlands; TA307336), SOCS6 (1 : 1000, Santa Cruz Biotechnology, Heidelberg, Germany; sc-133058) and GAPDH (1 : 3000, Sigma-Aldrich, G9545).

Techniques: Gene Expression

Journal: British journal of cancer

Article Title: Identification of SOCS2 and SOCS6 as biomarkers in human colorectal cancer.

doi: 10.1038/bjc.2014.377

Figure Lengend Snippet: Figure 1. Bioinformatic analysis reveals SOCS2 and SOCS6 downregulation in CRC. (A) Bar plot showing the log2 FC values of the SOCS family genes and marker genes in adenoma and CRC samples compared to normal colorectal mucosa. Error bars correspond to 95% confidence intervals for mean log FC; *FDRo0.05, **FDRo0.01 and ***FDRo0.001. (B) Heatmap representation of SOCS (highlighted in yellow) and STAT (highlighted in grey) family genes as well as marker genes in normal tissues, IBD, adenomas, and CRC stages (A–D).

Article Snippet: Membranes were blocked with 10% milk for 1 h at room temperature and were then incubated overnight at 41C with antibodies directed against SOCS2 (1 : 1000, OriGene Technologies, Uden, Netherlands; TA307336), SOCS6 (1 : 1000, Santa Cruz Biotechnology, Heidelberg, Germany; sc-133058) and GAPDH (1 : 3000, Sigma-Aldrich, G9545).

Techniques: Marker

Journal: British journal of cancer

Article Title: Identification of SOCS2 and SOCS6 as biomarkers in human colorectal cancer.

doi: 10.1038/bjc.2014.377

Figure Lengend Snippet: Figure 3. Expression of SOCS family members in primary human CRC samples. (A) Marker gene expression levels in CRC vs normal tissue of two patients. (B) SOCS1, SOCS2, SOCS3 and SOCS6 mRNA levels in CRC compared to normal mucosal tissue. Data are presented as mean±s.e.m.; *Po0.05, **Po0.001 and ***Po0.0001.

Article Snippet: Membranes were blocked with 10% milk for 1 h at room temperature and were then incubated overnight at 41C with antibodies directed against SOCS2 (1 : 1000, OriGene Technologies, Uden, Netherlands; TA307336), SOCS6 (1 : 1000, Santa Cruz Biotechnology, Heidelberg, Germany; sc-133058) and GAPDH (1 : 3000, Sigma-Aldrich, G9545).

Techniques: Expressing, Marker, Gene Expression

Journal: British journal of cancer

Article Title: Identification of SOCS2 and SOCS6 as biomarkers in human colorectal cancer.

doi: 10.1038/bjc.2014.377

Figure Lengend Snippet: Figure 4. Protein levels of SOCS2 and SOCS6 in human CRC. (A) Western blot detection of SOCS2 and SOCS6 in CRC and distant normal counterpart tissues. Caco-2 and Hela cells served as positive control for SOCS2 and SOCS6, respectively. The samples were run on the same gel but the membrane was cut for representative purposes (dotted line). (B) Immunohistochemical staining for SOCS2 and SOCS6 in CRC and normal counterpart for two different patients, magnification 200.

Article Snippet: Membranes were blocked with 10% milk for 1 h at room temperature and were then incubated overnight at 41C with antibodies directed against SOCS2 (1 : 1000, OriGene Technologies, Uden, Netherlands; TA307336), SOCS6 (1 : 1000, Santa Cruz Biotechnology, Heidelberg, Germany; sc-133058) and GAPDH (1 : 3000, Sigma-Aldrich, G9545).

Techniques: Western Blot, Positive Control, Membrane, Immunohistochemical staining, Staining

Journal: British journal of cancer

Article Title: Identification of SOCS2 and SOCS6 as biomarkers in human colorectal cancer.

doi: 10.1038/bjc.2014.377

Figure Lengend Snippet: Figure 5. Methylation analysis of the SOCS2 promoter in primary human CRC samples. (A) Treatment with 5-aza-dC/DAC increases basal SOCS2 expression in LS174t and HT-29 colon cancer cell lines whereas SOCS6 levels are unchanged. Data are representative of four independent experiments and presented as mean±s.d. (B) Representative histogram of the methylation pattern of four primary human CRC samples compared to their respective normal counterpart in a sequence 158 to þ 334 relative to the start codon. Each CpG site analysed is represented by a dot. (C) Representation of the SOCS2 promoter region that shows differential methylation between tumour and normal control samples. The sequence analysed by mass array is underlined and the CpG sites are highlighted in red. Analysis of the methylated sequence shows the presence of 2 STAT GAS motifs (TTCnnnGAA) highlighted in yellow. The start codon is indicated in green. The primers used in other SOCS2 methylation studies are indicated (Liu et al: pink arrows; Fiegl et al: green arrows; Sutherland et al: blue arrows).

Article Snippet: Membranes were blocked with 10% milk for 1 h at room temperature and were then incubated overnight at 41C with antibodies directed against SOCS2 (1 : 1000, OriGene Technologies, Uden, Netherlands; TA307336), SOCS6 (1 : 1000, Santa Cruz Biotechnology, Heidelberg, Germany; sc-133058) and GAPDH (1 : 3000, Sigma-Aldrich, G9545).

Techniques: Methylation, Expressing, Sequencing, Control

Journal: British journal of cancer

Article Title: Identification of SOCS2 and SOCS6 as biomarkers in human colorectal cancer.

doi: 10.1038/bjc.2014.377

Figure Lengend Snippet: Figure 6. Prognostic value of SOCS2 and SOCS6 in CRC. Disease-free survival curves for patients with high and low expression of SOCS2 or SOCS6 in primary CRC including (A) all stages or (B) stages I and II only. The number of patients included in each group is mentioned within brackets. Significant P-values are indicated.

Article Snippet: Membranes were blocked with 10% milk for 1 h at room temperature and were then incubated overnight at 41C with antibodies directed against SOCS2 (1 : 1000, OriGene Technologies, Uden, Netherlands; TA307336), SOCS6 (1 : 1000, Santa Cruz Biotechnology, Heidelberg, Germany; sc-133058) and GAPDH (1 : 3000, Sigma-Aldrich, G9545).

Techniques: Expressing

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Inhibition of cell proliferation in METTL3-KO SW480 cells. (A) METTL3-KO SW480 cells showed dysregulation of several proliferation-related genes (*P<0.05, compared with the SW480 WT cells). (B) MTS assay results validated the decreased proliferation of METTL3-KO cells (*P<0.05, compared with the SW480 WT cells). (C) METTL3-KO cells showed a reduced proliferation rate, as measured by cell counting (*P<0.05, compared with the SW480 WT cells). (D) METTL3-KO cells showed a lower colony formation rate than control cells at 7 days after seeding. (E) Tumorigenic ability of METTL3-KO SW480 cells was reduced (*P<0.05, compared with the SW480 WT cells). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5; SOX2, SRY-box transcription factor 2; PCNA, proliferating cell nuclear antigen; KO, knockout; SW480 A7 and SW480 A12, METTL3-KO SW480 cells.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Inhibition, MTS Assay, Cell Counting, Control, Knock-Out

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: SOCS2 is upregulated in colon cancer and downregulated in METTL3-KO SW480 cells. (A) TCGA database analysis revealed a lower expression level of SOCS2 in colon cancer tissues than in the paired normal tissues (*P<0.05). (B) Expression levels of SOCS2 and METTL3 were negatively correlated in colon cancer tissues. (C-E) METTL3-KO upregulated SOCS2 expression at both the (C) mRNA and (D and E) protein levels in SW480 cells (*P<0.05, compared with the SW480 WT cells). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; KO, knockout; SW480 A7 and SW480 A12, METTL3-KO SW480 cells.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Expressing, Knock-Out

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Methylation status of SOCS2 mRNA in SW480 cells. (A) Comparison of the SOCS2 mRNA methylation status between SW480 WT and METTL3-KO SW480 cells via MeRIP-qPCR (*P<0.05, compared with the SW480 WT cells). (B) Dose-dependent increase in SOCS2 mRNA levels in SW480 cells in response to demethylation treatment with DAA (*P<0.05, compared with non-treated group). (C and D) SOCS2 protein expression was upregulated after demethylation treatment with DAA (*P<0.05, compared with the SW380 WT DAA-group). (E) DAA appreciably decreased the m 6 A level in SOCS2 mRNA (*P<0.05, compared with the SW380 WT DAA-group). (F) METTL3-KO increased SOCS2 mRNA stability, showing a decreased mRNA decay rate (*P<0.05, compared with the SW480 WT cells). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; KO, knockout; SW480 A7 and SW480 A12, METTL3-KO SW480 cells; DAA, 3-deazaadenosine.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Methylation, Comparison, Expressing, Knock-Out

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Modulation of SOCS2 expression alters the proliferation ability of SW480 cells. (A-C) siRNA (si-SOCS2) was used to successfully knock down SOCS2 expression in SW480 cells. Knockdown of SOCS2 had minor impact on METTL3 expression but significantly increased LGR5 expression in SW480 cells (*P<0.05, compared with the si-Ctrl group). (D) Knockdown of SOCS2 enhanced SW480 cell proliferation, as determined by an MTS assay (*P<0.05, compared with the si-Ctrl group). (E-G) Transfection of the pCMV6-SOCS2 plasmid upregulated SOCS2 expression and decreased LGR5 expression in SW480 cells (*P<0.05, compared with the pCMV6-Entry group). SOCS2 overexpression had minor impact on METTL3 expression (P>0.05, compared with the pCMV6-Entry group). (H) SOCS2 overexpression decreased SW480 cell proliferation, as determined by an MTS assay (*P<0.05, compared with the pCMV6-Entry group). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Expressing, Knockdown, MTS Assay, Transfection, Plasmid Preparation, Over Expression

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: METTL3 modulates LGR5 transcription through m 6 A-mediated changes in SOCS2 expression rather than through direct regulatory mechanisms. (A) TCGA database analysis revealed an induction of LGR5 transcription in colon cancer tissues when compared with normal tissues (**P<0.01, compared with the normal tissues). (B) Expression levels of SOCS2 and LGR5 were negatively correlated in colon cancer tissues. (C) METTL3 knockout (KO) did not obviously impact LGR5 mRNA methylation (as measured by MeRIP-qPCR). (D) METTL3 KO had a minor effect on the LGR5 mRNA decay rate. (E) DAA treatment did not impact LGR5 expression in SW480 cells. (F) Luciferase reporter assay results confirmed that SOCS2 negatively controls LGR5 promoter-driven luciferase activity (*P<0.05, pGL3 basic-LGR5 promoter group vs. FULL SOCS2-LGR5 promoter group). METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5; SOCS2, suppressor of cytokine signaling 2; TCGA, The Cancer Genome Atlas; SW480 A7 and SW480 A12, METTL3-KO SW480 cells; DAA, 3-deazaadenosine.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Expressing, Knock-Out, Methylation, Luciferase, Reporter Assay, Activity Assay

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Correlations among METTL3, SOCS2 and LGR5 expression levels in CRC. (A-C) Knockdown of METTL3 using siRNA (si-METTL3#1 and si-METTL3#2) significantly upregulated SOCS2 expression but downregulated LGR5 expression in SW480 cells (*P<0.05, compared with the si-Ctrl group). (D and E) Western blotting analysis of 24 paired tissues collected from CRC patients showed high protein levels of METTL3 and LGR5 and a relatively low protein level of SOCS2 in colon cancer (T, colon tumor samples; N, adjacent normal colon tissues; *P<0.05, compared to Normal tissues). (F) Visualization of expression profiles of METTL3, SOCS2 and LGR5 in CRC through the Oncomine database. METTL3 and LGR5 were upregulated in CRC datasets. In contrast, SOCS2 was downregulated in CRC datasets. CRC, colorectal cancer; SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Expressing, Knockdown, Western Blot

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Association between patient characteristics and gene expression in 24 paired CRC cases.

Article Snippet: The siRNA targeting SOCS2 (si-SOCS2) was obtained from Santa Cruz Biotechnology, Inc.

Techniques: Gene Expression, Expressing

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Inhibition of cell proliferation in METTL3-KO SW480 cells. (A) METTL3-KO SW480 cells showed dysregulation of several proliferation-related genes (*P<0.05, compared with the SW480 WT cells). (B) MTS assay results validated the decreased proliferation of METTL3-KO cells (*P<0.05, compared with the SW480 WT cells). (C) METTL3-KO cells showed a reduced proliferation rate, as measured by cell counting (*P<0.05, compared with the SW480 WT cells). (D) METTL3-KO cells showed a lower colony formation rate than control cells at 7 days after seeding. (E) Tumorigenic ability of METTL3-KO SW480 cells was reduced (*P<0.05, compared with the SW480 WT cells). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5; SOX2, SRY-box transcription factor 2; PCNA, proliferating cell nuclear antigen; KO, knockout; SW480 A7 and SW480 A12, METTL3-KO SW480 cells.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Inhibition, MTS Assay, Cell Counting, Control, Knock-Out

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: SOCS2 is upregulated in colon cancer and downregulated in METTL3-KO SW480 cells. (A) TCGA database analysis revealed a lower expression level of SOCS2 in colon cancer tissues than in the paired normal tissues (*P<0.05). (B) Expression levels of SOCS2 and METTL3 were negatively correlated in colon cancer tissues. (C-E) METTL3-KO upregulated SOCS2 expression at both the (C) mRNA and (D and E) protein levels in SW480 cells (*P<0.05, compared with the SW480 WT cells). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; KO, knockout; SW480 A7 and SW480 A12, METTL3-KO SW480 cells.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Expressing, Knock-Out

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Methylation status of SOCS2 mRNA in SW480 cells. (A) Comparison of the SOCS2 mRNA methylation status between SW480 WT and METTL3-KO SW480 cells via MeRIP-qPCR (*P<0.05, compared with the SW480 WT cells). (B) Dose-dependent increase in SOCS2 mRNA levels in SW480 cells in response to demethylation treatment with DAA (*P<0.05, compared with non-treated group). (C and D) SOCS2 protein expression was upregulated after demethylation treatment with DAA (*P<0.05, compared with the SW380 WT DAA-group). (E) DAA appreciably decreased the m 6 A level in SOCS2 mRNA (*P<0.05, compared with the SW380 WT DAA-group). (F) METTL3-KO increased SOCS2 mRNA stability, showing a decreased mRNA decay rate (*P<0.05, compared with the SW480 WT cells). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; KO, knockout; SW480 A7 and SW480 A12, METTL3-KO SW480 cells; DAA, 3-deazaadenosine.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Methylation, Comparison, Expressing, Knock-Out

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Modulation of SOCS2 expression alters the proliferation ability of SW480 cells. (A-C) siRNA (si-SOCS2) was used to successfully knock down SOCS2 expression in SW480 cells. Knockdown of SOCS2 had minor impact on METTL3 expression but significantly increased LGR5 expression in SW480 cells (*P<0.05, compared with the si-Ctrl group). (D) Knockdown of SOCS2 enhanced SW480 cell proliferation, as determined by an MTS assay (*P<0.05, compared with the si-Ctrl group). (E-G) Transfection of the pCMV6-SOCS2 plasmid upregulated SOCS2 expression and decreased LGR5 expression in SW480 cells (*P<0.05, compared with the pCMV6-Entry group). SOCS2 overexpression had minor impact on METTL3 expression (P>0.05, compared with the pCMV6-Entry group). (H) SOCS2 overexpression decreased SW480 cell proliferation, as determined by an MTS assay (*P<0.05, compared with the pCMV6-Entry group). SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Expressing, Knockdown, MTS Assay, Transfection, Plasmid Preparation, Over Expression

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: METTL3 modulates LGR5 transcription through m 6 A-mediated changes in SOCS2 expression rather than through direct regulatory mechanisms. (A) TCGA database analysis revealed an induction of LGR5 transcription in colon cancer tissues when compared with normal tissues (**P<0.01, compared with the normal tissues). (B) Expression levels of SOCS2 and LGR5 were negatively correlated in colon cancer tissues. (C) METTL3 knockout (KO) did not obviously impact LGR5 mRNA methylation (as measured by MeRIP-qPCR). (D) METTL3 KO had a minor effect on the LGR5 mRNA decay rate. (E) DAA treatment did not impact LGR5 expression in SW480 cells. (F) Luciferase reporter assay results confirmed that SOCS2 negatively controls LGR5 promoter-driven luciferase activity (*P<0.05, pGL3 basic-LGR5 promoter group vs. FULL SOCS2-LGR5 promoter group). METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5; SOCS2, suppressor of cytokine signaling 2; TCGA, The Cancer Genome Atlas; SW480 A7 and SW480 A12, METTL3-KO SW480 cells; DAA, 3-deazaadenosine.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Expressing, Knock-Out, Methylation, Luciferase, Reporter Assay, Activity Assay

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Correlations among METTL3, SOCS2 and LGR5 expression levels in CRC. (A-C) Knockdown of METTL3 using siRNA (si-METTL3#1 and si-METTL3#2) significantly upregulated SOCS2 expression but downregulated LGR5 expression in SW480 cells (*P<0.05, compared with the si-Ctrl group). (D and E) Western blotting analysis of 24 paired tissues collected from CRC patients showed high protein levels of METTL3 and LGR5 and a relatively low protein level of SOCS2 in colon cancer (T, colon tumor samples; N, adjacent normal colon tissues; *P<0.05, compared to Normal tissues). (F) Visualization of expression profiles of METTL3, SOCS2 and LGR5 in CRC through the Oncomine database. METTL3 and LGR5 were upregulated in CRC datasets. In contrast, SOCS2 was downregulated in CRC datasets. CRC, colorectal cancer; SOCS2, suppressor of cytokine signaling 2; METTL3, methyltransferase like 3; LGR5, leucine-rich repeat-containing G protein-coupled receptor 5.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Expressing, Knockdown, Western Blot

Journal: Oncology Reports

Article Title: m 6 A methyltransferase METTL3 maintains colon cancer tumorigenicity by suppressing SOCS2 to promote cell proliferation

doi: 10.3892/or.2020.7665

Figure Lengend Snippet: Association between patient characteristics and gene expression in 24 paired CRC cases.

Article Snippet: The pCMV6 plasmid containing the full-length SOCS2 sequence (FULL-SOCS2) was purchased from Origene (RC203163).

Techniques: Gene Expression, Expressing

Journal: European journal of immunology

Article Title: Deficient SOCS3 expression in CD4+CD25+FoxP3+ regulatory T cells and SOCS3-mediated suppression of Treg function.

doi: 10.1002/eji.200737193

Figure Lengend Snippet: Figure 2. SOCS3 expression is significantly lower in Treg as compared to that in Th cells. (A) Freshly isolated Th cells were stimulated under Th1 or Th2 biasing conditions and total cell extracts were made at t = 0, 1, 2, and 5 days. Extracts were analyzed by Western blotting techniques for SOCS3 expression, and then reprobed for b-actin to ensure equal protein loading in each lane. (B) Treg and Th cells were stimulated under neutral conditions and total cell extracts were made at t = 0, 3, and 5 days, and total cell extracts were made and analyzed as in A. (C) SOCS3 and FoxP3 mRNA expression was evaluated in unstimulated Treg and Th cells by quantitative RT-PCR techniques, and normalized to the housekeeping gene b-glu- nidase (GUS), which was equally expressed in Treg and Th cells. Graphs show the fold change of FoxP3 or SOCS3 in Treg compared to that in Th cells, which was arbitrarily set to one. (D) Cell extracts prepared from unstimulated Treg and Th cells were analyzed for expression of SOCS2 and SOCS1 proteins as in panel (A). Results shown are representative of two independent experiments.

Article Snippet: Membranes were probed with antimurine SOCS3mAb (MBL), or anti-murine SOCS1mAb (MBL) at 1:1000 dilutions, or anti-murine SOCS2 polyclonal antibody (Novus) at a 1:500 dilution.

Techniques: Expressing, Isolation, Western Blot, Quantitative RT-PCR

Journal: Journal of the American Chemical Society

Article Title: Rewiring the fusion oncoprotein EWSR1::FLI1 in Ewing sarcoma with bivalent small molecules

doi: 10.1021/jacs.5c05634

Figure Lengend Snippet: Volcano plots of differentially expressed RNA species in BCL6 KO (two guides averaged over three replicates per guide) EWS502 cells (A) or TC32 cells (B) vs Chr2.2 control cells. GSEA in EWS502 (C) or TC32 (D) BCL6 KO cells shows a positive correlation with a published BCL6 gene signature derived from BCL6 promoter binding data 25 . SOCS2 and CISH transcripts have increased expression by RT-qPCR in BCL6 KO EWS502 cells (E and F) or TC32 cells (G and H) vs control guides. Average expression from three independent cell transductions (performed in technical triplicate) is shown. RT-qPCR data was compared by one-way ANOVA; NS = not significant, **** p < 0.001. All error bars in the figure show mean ± SD. Immunoblotting shows BCL6 KO and corresponding increase in SOCS2 protein levels in EWS502 (I) and TC32 (J) cells. Each lane is from an independent transduction of cells. GAPDH serves as a loading control.

Article Snippet: Probes used in this study include: SOCS2 : Hs00919620_m1, CISH : Hs00367082_G1, BCL6 : Hs00153368_m1, and GAPDH : Hs02786624_G1 (60x primer limiting).

Techniques: Control, Derivative Assay, Binding Assay, Expressing, Quantitative RT-PCR, Western Blot, Transduction

Journal: Journal of the American Chemical Society

Article Title: Rewiring the fusion oncoprotein EWSR1::FLI1 in Ewing sarcoma with bivalent small molecules

doi: 10.1021/jacs.5c05634

Figure Lengend Snippet: (A) Schematic of EB-TCIP mechanism of action. EB-TCIP induces a ternary complex between FKBP F36V tagged EWSR1::FLI1 and BCL6, which leads to activation of BCL6 target gene transcription. Image made with Biorender. (B) Structures of compounds used in this work. (C) EB-TCIP increases the association of BCL6 with FKBP-E::F in a dose-dependent manner in EWS502 FKBP-E::F cell lysates while NEG-1 (D) does not induce a ternary complex. The association is reversible as excess BI3812 (C) and excess OAP (free acid) (D) abrogate ternary complex formation. GAPDH was probed to confirm that unbound proteins were removed by washing. EB-TCIP dose-dependently increases SOCS2 (E) and CISH (F) expression by RT-qPCR. (G) SOCS2 protein levels dose-dependently increase while BCL6 protein levels dose-dependently decrease after EB-TCIP treatment. EB-TCIP induces higher SOCS2 (H) and CISH (I) transcript levels than chemical inhibition with BI3812 or chemically induced degradation with BI3802 (DEG) . (J) SOCS2 protein levels are highest in EB-TCIP treated cells compared to BI3812 , BI3802 (DEG) , or negative control compounds that do not form ternary complexes. Immunoblotting is representative of three biological replicates and GAPDH serves as a loading control. All experiments were performed in FKBP-E::F expressing EWS502 cells. RT-qPCR experiments show one experiment with technical triplicate that is representative of three biological replicates. Means of SOCS2 and CISH expression were compared using one-way ANOVA with multiple comparisons; NS = not significant, *** p < 0.005, **** p < 0.001. All error bars in the figure indicate mean ± SD. Unless indicated with brackets, significance above each condition indicates comparison of that mean to the mean of DMSO.

Article Snippet: Probes used in this study include: SOCS2 : Hs00919620_m1, CISH : Hs00367082_G1, BCL6 : Hs00153368_m1, and GAPDH : Hs02786624_G1 (60x primer limiting).

Techniques: Expressing, Activation Assay, Quantitative RT-PCR, Inhibition, Negative Control, Western Blot, Control, Comparison

Journal: Journal of the American Chemical Society

Article Title: Rewiring the fusion oncoprotein EWSR1::FLI1 in Ewing sarcoma with bivalent small molecules

doi: 10.1021/jacs.5c05634

Figure Lengend Snippet: (A) Time course of SOCS2 and BCL6 protein levels. BCL6 degradation occurs within 1 h for both EB-TCIP and BI3802 (DEG) . EB-TCIP induces SOCS2 expression by 2 h and maintains higher expression levels than BI3812 or BI3802 (DEG) throughout the time course. SOCS2 (B) and CISH (C) transcripts reach a maximum between 2 and 4 h by RT-qPCR. (D) EB-TCIP induced SOCS2 protein expression can be reversed with 25-fold excess OAP (free acid). Co-treatment of 1 µM BI3812 and OAP do not increase SOCS2 protein expression more than 1 µM BI3812 alone. EB-TCIP induced SOCS2 (E) and CISH (F) transcript expression is reversed with excess OAP . BI3812 and OAP must be chemically linked to induce maximum transcript expression. (G) EB-TCIP induces the highest expression of SOCS2 protein in EWS502 FKBP-E::F cells compared to EWS502 parental cells or EWS502 cells expressing FKBP-GFP. Only treatment with EB-TCIP induces more expression of SOCS2 (H) and CISH (I) than BI3812 in EWS502 FKBP-E::F cells. EB-TCIP : BI3812 ratio was calculated by dividing the average expression of each transcript in EB-TCIP treated cells by the average expression of each transcript in BI3812 treated cells. Immunoblotting is representative of three biological replicates. RT-qPCR experiments show one experiment with technical triplicate or quadruplicate that is representative of three biological replicates. The means of SOCS2 and CISH expression were compared using one-way ANOVA with multiple comparisons; NS = not significant, * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.001. All error bars in the figure represent mean ± SD. Unless indicated with brackets, significance above each condition indicates comparison of that mean to the mean of DMSO. In (H) and (I), unless indicated with brackets, the means of BI3812 , EB-TCIP , and NEG-1 were compared to the DMSO sample for the corresponding cell line.

Article Snippet: Probes used in this study include: SOCS2 : Hs00919620_m1, CISH : Hs00367082_G1, BCL6 : Hs00153368_m1, and GAPDH : Hs02786624_G1 (60x primer limiting).

Techniques: Activity Assay, Expressing, Quantitative RT-PCR, Western Blot, Comparison

Journal: Journal of the American Chemical Society

Article Title: Rewiring the fusion oncoprotein EWSR1::FLI1 in Ewing sarcoma with bivalent small molecules

doi: 10.1021/jacs.5c05634

Figure Lengend Snippet: Volcano plots portraying log 2 fold changes of gene expression from cells treated with 2.5 µM EB-TCIP versus DMSO at 8 (A) and 24 (B) h with a −log 10 adjusted P-value cut off of 1. EB-TCIP treatment predominantly increases expression of transcripts at both timepoints. Volcano plots portraying log 2 fold changes of cells treated with 2.5 µM EB-TCIP versus 2.5 µM BI3812 (C) or 2.5 µM NEG-1 (D) at 8 hours with a −log 10 P-value cut off of 1.3. EB-TCIP induces higher expression of BCL6 transcripts than BI3812 or NEG-1 at this early timepoint. Dots corresponding to SOCS2, CISH, and CXCL11 are labelled with black borders. (E) Heatmaps of changes in BCL6 target gene expression at 4, 8 and 24 h show that EB-TCIP induces faster and/or higher expression of these select genes. (F) GSEA comparing EB-TCIP treated EWS502 FKBP-E::F cells to BCL6 KO EWS502 parental cells (LFC ≥ 1.65) at 4 (top), 8 (middle) and 24 h (bottom) show significant positive correlation between the two gene sets. RNA-seq data is shown as the average of three independent replicates.

Article Snippet: Probes used in this study include: SOCS2 : Hs00919620_m1, CISH : Hs00367082_G1, BCL6 : Hs00153368_m1, and GAPDH : Hs02786624_G1 (60x primer limiting).

Techniques: Gene Expression, Expressing, Targeted Gene Expression, RNA Sequencing

Journal: Journal of the American Chemical Society

Article Title: Rewiring the fusion oncoprotein EWSR1::FLI1 in Ewing sarcoma with bivalent small molecules

doi: 10.1021/jacs.5c05634

Figure Lengend Snippet: (A) ChIP-seq tornado plots of HA (FKBP-E::F) binding signal of EB-TCIP (red) versus DMSO (black) peaks that are decreasing (DEC; 92), non-significantly changing (NS; 8656), and increasing (INC; 2296) at 24 h. Differential peaks between EB-TCIP and DMSO are shown for all compounds. Compared to BI3812 (brown) and BI3802 ( DEG , purple), EB-TCIP increases FKBP-E::F binding at a subset of genes. Line plots for all compound treatments in each cluster are shown to the right. (B) Scatter plot portraying top enriched motifs of HA binding sites in DMSO treated cells. (C) Scatter plot portraying top motifs of HA increased peaks enriched in EB-TCIP treated cells. The BCL6 motif scores 29 th . (D) Scatter plot portraying top motifs of HA decreased peaks enriched in EB-TCIP treated cells. IGV visualization of input, HA (FKBP-E::F), BCL6, ATAC-seq signal, and RNA-seq signal at the SOCS2 (E) and CISH (F) with treatments DMSO (black), 1 µM BI3812 (brown), 1 µM BI3802 (DEG , purple), and 1 µM EB-TCIP (red). All ChIP-seq and ATAC-seq is portrayed as the average of two independent replicates. RNA-seq is portrayed as the average of three independent replicates.

Article Snippet: Probes used in this study include: SOCS2 : Hs00919620_m1, CISH : Hs00367082_G1, BCL6 : Hs00153368_m1, and GAPDH : Hs02786624_G1 (60x primer limiting).

Techniques: ChIP-sequencing, Binding Assay, RNA Sequencing

Journal: BMC Cancer

Article Title: Favorable prognostic value of SOCS2 and IGF-I in breast cancer

doi: 10.1186/1471-2407-7-136

Figure Lengend Snippet: Kaplan-Meier curve assessment of risk of death in a cohort of 89 patients with invasive breast cancer. Overall survival is shown for patients with high and low SOCS2 and IGF-I expression, respectively. The median was taken as a cut-off. P values were determined using the log-rank test.

Article Snippet: To test for antibody specificity, lysates from COS7 cells transiently transfected with a myc-tagged SOCS2 plasmid were probed with an anti-c-myc antibody (9E10, Santa Cruz Biotechnology, Santa Cruz) and anti-SOCS2 antibodies.

Techniques: Expressing

Journal: BMC Cancer

Article Title: Favorable prognostic value of SOCS2 and IGF-I in breast cancer

doi: 10.1186/1471-2407-7-136

Figure Lengend Snippet: Comparison of SOCS2 and IGF-I expression. (A) Box blots with data from a set of seven tumors comparing the relative mRNA expression levels in tumor (CA) and normal adjacent tissue (TA). (B) SOCS2 protein and corresponding SOCS2 mRNA expression in normal and cancerous tissue of two tumor specimens. SOCS2 protein expression was assessed by immuno-blot with an antibody directed against SOCS2. Actin was used as a loading control.

Article Snippet: To test for antibody specificity, lysates from COS7 cells transiently transfected with a myc-tagged SOCS2 plasmid were probed with an anti-c-myc antibody (9E10, Santa Cruz Biotechnology, Santa Cruz) and anti-SOCS2 antibodies.

Techniques: Expressing

Journal: BMC Cancer

Article Title: Favorable prognostic value of SOCS2 and IGF-I in breast cancer

doi: 10.1186/1471-2407-7-136

Figure Lengend Snippet: Detection of SOCS2 in primary mammary carcinoma samples by immunohistochemistry. Representative immunohistochemical stainings of samples with low SOCS2 (A) and high SOCS2 (B) mRNA expression are shown. Note a strong cytoplasmatic epithelial staining for SOCS2 in tumors with high SOCS2 mRNA levels (B).

Article Snippet: To test for antibody specificity, lysates from COS7 cells transiently transfected with a myc-tagged SOCS2 plasmid were probed with an anti-c-myc antibody (9E10, Santa Cruz Biotechnology, Santa Cruz) and anti-SOCS2 antibodies.

Techniques: Immunohistochemistry, Immunohistochemical staining, Expressing, Staining