ptp4a3  (Santa Cruz Biotechnology)

Journal: Molecular Cancer Therapeutics

Article Title: Tyrosine Kinase Inhibitors Diminish Renal Neoplasms in a Tuberous Sclerosis Model Via Induction of Apoptosis

doi: 10.1158/1535-7163.MCT-22-0224

Figure Lengend Snippet: Differential signaling mechanisms induced by rapamycin compared to imatinib in UMB1949 renal angiomyolipoma and pulmonary LAM tumor cells. Top 20 canonical signaling mechanisms ( A ) and biological functions ( B ) predicted to be activated ( Z -score ≥ 2, orange squares) or inhibited ( Z -score ≤ −2, blue squares) by concentrations of rapamycin or the TKI imatinib determined by comparative RNA-sequencing analysis. GPVI signaling and cell survival and viability biofunctions (indicated with red asterisks) were resolved canonical pathways and biofunctions that exhibit both widely varying significant Z -scores by drug type and significant P values of Fisher exact tests of the likelihood of association of differentially expressed genes in our dataset with curated genes and canonical pathways in IPA knowledge base (see Supplementary Figs. S3A and S3B). RT-qPCR validation of cell survival and viability genes resolved by IPA bioinformatics including ( C ) protein tyrosine phosphatase type A3 ( PTP4A3 ), ( D ) inositol polyphosphate-5-phosphatase D ( INPP5D ), and ( E ) matrix metalloproteinase 13 ( MMP13 ) was performed. Experiments were run in quadruplicates per gene per sample and N = 2 biological samples were used for RT-qPCR analysis. Statistical comparisons were performed using a one-way ANOVA with post hoc Dunnett multiple comparisons test where **, P < 0.005, ***, P = 0.0006, ****, P < 0.0001; and ns, not significant. F, Bioinformatics analysis of differentially expressed genes when pulmonary LAM is treated with rapamycin versus imatinib reveals activation of G 2 –M cell-cycle regulatory mechanisms only in 0.05 μmol/L rapamycin condition. RNA sequencing was performed using ( N = 3) biological replicates of renal AML and ( N = 2) pulmonary LAM cell lysates. Gray dots denote test conditions resulting in nonsignificant values below set threshold (−2 ≥ Z ≥ 2). Principal component analysis cluster renal AML RNA sequence data by drug type ( G ) with variances resolved with first eigenvalue of 0.60, and cluster pulmonary LAM tumor cells ( H ) with first eigenvalue of 0.61 indicating differential drug-induction mechanisms in these cells. Integrin subunit beta 3 ( ITGB3 ) is significantly upregulated in renal AML cells exposed to 1 μmol/L Rapamycin (*, P < 0.01) and 10 μmol/L imatinib (*, P < 0.05; I ) and at all concentrations in pulmonary LAM cells (*, P < 0.001) by RT-qPCR ( J ). Pharmacologic inhibition of ITGB3 in renal AML cells (*, 0.05> P < 0.001; ϕ, 0.05> P < 2.4E−05; K ) and pulmonary LAM tumor cells (*, 0.0005> P < 7.23E−07; ϕ, 1.9E−05> P < 1.0E−0.6; L ) using small molecule CGT at 25 μmol/L concentration significantly reversed cytotoxicity of 1 μmol/L rapamycin and cytocidal effects of 1 and 10 μmol/L imatinib compared with experimental conditions where CGT was not added. RT-qPCR experiments were performed twice using three wells/concentration/experiment. CGT assays were performed twice using three to four wells/concentration/experiment. Error bars denote mean ± SEM of replicate experiments assessed for statistical significance using a one-way ANOVA with post hoc Dunnett multiple comparisons test (RT-qPCR) or multiple T tests (CGT assays). Vehicle = 0.01% DMSO in respective cell culture media.

Article Snippet: Gene expression analysis by RT-qPCR was performed using primers for human PTP4A3 (Hs02341135_m1), INPP5D (Hs00183290_m1), MMP13 (Hs00233992_m1), MTOR (Hs00234508_m1), DEPTOR (Hs00961900_m1), and ITGB3 (Hs01001469_m1; Catalog No. 4331182, Thermo Fisher Scientific) as described previously ( ).

Techniques: RNA Sequencing, Quantitative RT-PCR, Biomarker Discovery, Activation Assay, Sequencing, Inhibition, Concentration Assay, Cell Culture

Journal: SAGE Open Medicine

Article Title: SNHG19 promotes the proliferation and metastasis of hepatocellular carcinoma through regulating the miR-137/protein tyrosine phosphatase 4A3 axis

doi: 10.1177/20503121251334272

Figure Lengend Snippet: SNHG19 promote PTP4A3 expression by competitive binding miR-137 in HCC cells. (a) The binding sites of miR-137 within the 3ʹ-UTR of PTP4A3. (b) Luciferase activity of HCC cells transfected with PTP4A3-WT or PTP4A3-MUT reporter. (c) The expression of PTP4A3 mRNA in HCC cells transfected with miR-137 mimic, SNHG19 siRNA, or SNHG19 siRNA plus miR-137 inhibitor. (d) Western blots identified PTP4A3 protein expression changes, GAPDH was used as a control. (e) Luciferase assay of HCC cells transfected with PTP4A3-WT or PTP4A3-MUT reporter together with SNHG19 siRNA or SNHG19 siRNA plus miR-137 inhibitor. (f) TCGA dataset revealed a significant positive correlation between PTP4A3 mRNA and SNHG19 in HCC tissues. ** p < 0.01.

Article Snippet: We detected the signal on the membrane after incubating the secondary antibody for 1 h. Antibodies against PTP4A3 (Affinity Biosciences, 1:1000, USA) was used.

Techniques: Expressing, Binding Assay, Luciferase, Activity Assay, Transfection, Western Blot, Control

Journal: SAGE Open Medicine

Article Title: SNHG19 promotes the proliferation and metastasis of hepatocellular carcinoma through regulating the miR-137/protein tyrosine phosphatase 4A3 axis

doi: 10.1177/20503121251334272

Figure Lengend Snippet: SNHG19 promotes the growth and metastasis of HCC cell via regulating miR-137/PTP4A3 axis. (a) Western blots identified PTP4A3 protein expression changes in transfected HCC cells, GAPDH was used as a control. (b) The proliferative ability of HCC cells was determined by CCK8 assay. (c) The migratory ability of HepG2 and Huh7 cells was assessed by the scratch wound assay. Scale bar, 200 μm. (d) The invasive capacity of HepG2 and Huh7 cells was assessed by the Transwell assay. Scale bar, 50 μm. (e) The proliferative capacity of HepG2 and Huh7 cells was assessed by Clone formation assay. * p < 0.05, ** p < 0.01.

Article Snippet: We detected the signal on the membrane after incubating the secondary antibody for 1 h. Antibodies against PTP4A3 (Affinity Biosciences, 1:1000, USA) was used.

Techniques: Western Blot, Expressing, Transfection, Control, CCK-8 Assay, Scratch Wound Assay Assay, Transwell Assay, Tube Formation Assay

Journal: SAGE Open Medicine

Article Title: SNHG19 promotes the proliferation and metastasis of hepatocellular carcinoma through regulating the miR-137/protein tyrosine phosphatase 4A3 axis

doi: 10.1177/20503121251334272

Figure Lengend Snippet: SNHG19 exerts its oncogenic activity in vivo. (a) The excision tumor in nude mice of HCC xenograft model. (b) Differences in tumor volume among groups. (c) PCR identified SNHG19 and miR-497-5p expression changes. (d) The expression of PTP4A3 was examined by immunohistochemical staining of sections from the xenograft model. Scale bar, 25 μm. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: We detected the signal on the membrane after incubating the secondary antibody for 1 h. Antibodies against PTP4A3 (Affinity Biosciences, 1:1000, USA) was used.

Techniques: Activity Assay, In Vivo, Expressing, Immunohistochemical staining, Staining

Journal: SAGE Open Medicine

Article Title: SNHG19 promotes the proliferation and metastasis of hepatocellular carcinoma through regulating the miR-137/protein tyrosine phosphatase 4A3 axis

doi: 10.1177/20503121251334272

Figure Lengend Snippet: SNHG19 promote PTP4A3 expression by competitive binding miR-137 in HCC cells. (a) The binding sites of miR-137 within the 3ʹ-UTR of PTP4A3. (b) Luciferase activity of HCC cells transfected with PTP4A3-WT or PTP4A3-MUT reporter. (c) The expression of PTP4A3 mRNA in HCC cells transfected with miR-137 mimic, SNHG19 siRNA, or SNHG19 siRNA plus miR-137 inhibitor. (d) Western blots identified PTP4A3 protein expression changes, GAPDH was used as a control. (e) Luciferase assay of HCC cells transfected with PTP4A3-WT or PTP4A3-MUT reporter together with SNHG19 siRNA or SNHG19 siRNA plus miR-137 inhibitor. (f) TCGA dataset revealed a significant positive correlation between PTP4A3 mRNA and SNHG19 in HCC tissues. ** p < 0.01.

Article Snippet: Immunohistochemistry was performed using antibody PTP4A3 (Affinity Biosciences, USA).

Techniques: Expressing, Binding Assay, Luciferase, Activity Assay, Transfection, Western Blot, Control

Journal: SAGE Open Medicine

Article Title: SNHG19 promotes the proliferation and metastasis of hepatocellular carcinoma through regulating the miR-137/protein tyrosine phosphatase 4A3 axis

doi: 10.1177/20503121251334272

Figure Lengend Snippet: SNHG19 promotes the growth and metastasis of HCC cell via regulating miR-137/PTP4A3 axis. (a) Western blots identified PTP4A3 protein expression changes in transfected HCC cells, GAPDH was used as a control. (b) The proliferative ability of HCC cells was determined by CCK8 assay. (c) The migratory ability of HepG2 and Huh7 cells was assessed by the scratch wound assay. Scale bar, 200 μm. (d) The invasive capacity of HepG2 and Huh7 cells was assessed by the Transwell assay. Scale bar, 50 μm. (e) The proliferative capacity of HepG2 and Huh7 cells was assessed by Clone formation assay. * p < 0.05, ** p < 0.01.

Article Snippet: Immunohistochemistry was performed using antibody PTP4A3 (Affinity Biosciences, USA).

Techniques: Western Blot, Expressing, Transfection, Control, CCK-8 Assay, Scratch Wound Assay Assay, Transwell Assay, Tube Formation Assay

Journal: SAGE Open Medicine

Article Title: SNHG19 promotes the proliferation and metastasis of hepatocellular carcinoma through regulating the miR-137/protein tyrosine phosphatase 4A3 axis

doi: 10.1177/20503121251334272

Figure Lengend Snippet: SNHG19 exerts its oncogenic activity in vivo. (a) The excision tumor in nude mice of HCC xenograft model. (b) Differences in tumor volume among groups. (c) PCR identified SNHG19 and miR-497-5p expression changes. (d) The expression of PTP4A3 was examined by immunohistochemical staining of sections from the xenograft model. Scale bar, 25 μm. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Immunohistochemistry was performed using antibody PTP4A3 (Affinity Biosciences, USA).

Techniques: Activity Assay, In Vivo, Expressing, Immunohistochemical staining, Staining

Journal: bioRxiv

Article Title: Targeting of PTP4A3 overexpression sensitises HGSOC cells towards chemotherapeutic drugs

doi: 10.1101/2024.10.28.620719

Figure Lengend Snippet: ( A ) Expression of PTP4A3 was assessed in OVCAR 3, OVCAR 4 and Kuramochi cells by Western immunoblot and ( B ) this was quantified by densitometry, with PTP4A3 normalised to β-actin. ( C ) NCBI GEO repository RNA-seq data was analysed for gene expression in the three HGSOC cell lines, normalised to Kuramochi levels. ( D ) Cell lysates were analysed for phosphorylation of AKT/PKB (pSer473) and ERK1-2 (pThr202- Tyr204). Antibodies that recognise total AKT/PKB and ERK1-2 were used as controls. Expression of PTP4A3 is shown for comparison. ( E ) Turnover of PTP4A3 was analysed by incubating with or without either MG132 (2.5 µM, 2 h) or BafA1 (100 nM, 1 h) before lysates were collected with β-actin as loading control. Results are representative of at least three independent experiments. Error bars = ± S.E.M. **** p< 0.0001 by unpaired t -test.

Article Snippet: Membranes were incubated overnight at 4 °C on a rocker with the following antibodies in Blocking buffer: β-actin (Sigma-Aldrich, Gillingham, UK A5441), LC3B (Cell Signaling Technology, Leiden, Netherlands 2775), S6K1 (9202), p-S6K1 (9205), ERK1/2 (9102), p-ERK1/2 (9106), ULK1 (4773), p-ULK1 (6888), p-AKT (9271), PTP4A3 (Santa Cruz Biotechnology, Heidelberg, Germany sc-130355), AKT , pan-PTP4A3 (Bio-Techne MAB32191), and GFP (Thermo Fisher Scientific, Horsham, UK A-11122), then washed thrice with TBS-T and incubated with species-appropriate HRP- conjugated secondary antibodies (Thermo Fisher Scientific) diluted in Blocking buffer.

Techniques: Expressing, Western Blot, RNA Sequencing, Gene Expression, Phospho-proteomics, Comparison, Control

Journal: bioRxiv

Article Title: Targeting of PTP4A3 overexpression sensitises HGSOC cells towards chemotherapeutic drugs

doi: 10.1101/2024.10.28.620719

Figure Lengend Snippet: OVCAR 3 cells were transiently transfected with either pEGFP (empty vector), pEGFP-PTP4A1 or pEGFP-PTP4A3. ( A ) Cells were incubated with or without 100 nM BafA1 for 1 h. The phosphorylation of S6K1 (pThr389) was used to confirm the inhibition of mTORC1 signalling and β- actin was used as a loading control, while GFP was probed as a transfection efficiency control. ( B ) Basal autophagy activity was quantified by densitometry of LC3B-II and normalised to β-actin. ( C ) Alternatively, cells were incubated with or without EBSS for 2 h in the presence or absence of BafA1 (100 nM) for the last hour. ( D ) Activatable autophagy was quantified by densitometry of LC3B-II normalised to β-actin expression. Data was pooled from three independent experiments and error bars = ±S.E.M. * p< 0.05, *** p< 0.001, **** p< 0.0001 by two-way ANOVA test.

Article Snippet: Membranes were incubated overnight at 4 °C on a rocker with the following antibodies in Blocking buffer: β-actin (Sigma-Aldrich, Gillingham, UK A5441), LC3B (Cell Signaling Technology, Leiden, Netherlands 2775), S6K1 (9202), p-S6K1 (9205), ERK1/2 (9102), p-ERK1/2 (9106), ULK1 (4773), p-ULK1 (6888), p-AKT (9271), PTP4A3 (Santa Cruz Biotechnology, Heidelberg, Germany sc-130355), AKT , pan-PTP4A3 (Bio-Techne MAB32191), and GFP (Thermo Fisher Scientific, Horsham, UK A-11122), then washed thrice with TBS-T and incubated with species-appropriate HRP- conjugated secondary antibodies (Thermo Fisher Scientific) diluted in Blocking buffer.

Techniques: Transfection, Plasmid Preparation, Incubation, Phospho-proteomics, Inhibition, Control, Activity Assay, Expressing

Journal: bioRxiv

Article Title: Targeting of PTP4A3 overexpression sensitises HGSOC cells towards chemotherapeutic drugs

doi: 10.1101/2024.10.28.620719

Figure Lengend Snippet: Kuramochi ( A ) and OVCAR 4 ( B ) cells were infected with lentiviral particles harbouring either scrambled control shRNA (Scr) or PTP4A3-targeting shRNA (shPTP4A3). The silencing of PTP4A3 expression was quantified by Western immunoblotting. ( C, D ) Kuramochi-Scr (K- Scr), ( E, F ) Kuramochi-shPTP4A3 (K-shPTP4A3), ( G, H ) OVCAR 4-Scr (4-Scr) and ( I, J ) OVCAR 4-shPTP4A3 (4-shPTP4A3) cells were incubated with EBSS for 0, 1, 2 or 4 h and BafA1 (100 nM) was added for the final hour. ( C, E, G, I ) The phosphorylation of S6K1 (pThr389) was assessed by western blot to confirm inhibition of mTORC1 signalling. β-actin was used as a loading control. ( D, F, H, J ) Activation of autophagy was quantified by densitometry of LC3B-II normalised to β-actin. Data was pooled from at least three independent experiments and error bars = ±S.E.M. * p< 0.05, ** p< 0.01 by two-way ANOVA test.

Article Snippet: Membranes were incubated overnight at 4 °C on a rocker with the following antibodies in Blocking buffer: β-actin (Sigma-Aldrich, Gillingham, UK A5441), LC3B (Cell Signaling Technology, Leiden, Netherlands 2775), S6K1 (9202), p-S6K1 (9205), ERK1/2 (9102), p-ERK1/2 (9106), ULK1 (4773), p-ULK1 (6888), p-AKT (9271), PTP4A3 (Santa Cruz Biotechnology, Heidelberg, Germany sc-130355), AKT , pan-PTP4A3 (Bio-Techne MAB32191), and GFP (Thermo Fisher Scientific, Horsham, UK A-11122), then washed thrice with TBS-T and incubated with species-appropriate HRP- conjugated secondary antibodies (Thermo Fisher Scientific) diluted in Blocking buffer.

Techniques: Infection, Control, shRNA, Expressing, Western Blot, Incubation, Phospho-proteomics, Inhibition, Activation Assay

Journal: bioRxiv

Article Title: Targeting of PTP4A3 overexpression sensitises HGSOC cells towards chemotherapeutic drugs

doi: 10.1101/2024.10.28.620719

Figure Lengend Snippet: ( A ) Kuramochi-WT, Scr and shPTP4A3 and ( B ) OVCAR 4-WT, Scr and shPTP4A3 cells were treated with increasing concentrations of JMS-053 (iPRL; 0 - 25 µM) for 2 h. Cell extracts were analysed for phosphorylation of ULK1 (pSer757), S6K1 (pThr389), AKT/PKB (pSer473) and ERK1-2 (pThr202-Tyr204). LC3B was used as a marker of autophagy activity, β-actin was used as a loading control, and PTP4A3 knockdown was also confirmed. ( C-F ) The basal phosphorylation of AKT and ERK in WT, Scr and shPTP4A3 cells (no JMS-053 treatment) in ( C, D ) Kuramochi and ( E, F ) OVCAR 4 cells was determined. Data was pooled from three independent experiments and error bars = ±S.E.M.

Article Snippet: Membranes were incubated overnight at 4 °C on a rocker with the following antibodies in Blocking buffer: β-actin (Sigma-Aldrich, Gillingham, UK A5441), LC3B (Cell Signaling Technology, Leiden, Netherlands 2775), S6K1 (9202), p-S6K1 (9205), ERK1/2 (9102), p-ERK1/2 (9106), ULK1 (4773), p-ULK1 (6888), p-AKT (9271), PTP4A3 (Santa Cruz Biotechnology, Heidelberg, Germany sc-130355), AKT , pan-PTP4A3 (Bio-Techne MAB32191), and GFP (Thermo Fisher Scientific, Horsham, UK A-11122), then washed thrice with TBS-T and incubated with species-appropriate HRP- conjugated secondary antibodies (Thermo Fisher Scientific) diluted in Blocking buffer.

Techniques: Phospho-proteomics, Marker, Activity Assay, Control, Knockdown