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Millipore ing4 antibody
Ing4 Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 90 stars, based on 1 article reviews

ing4 antibody - by Bioz Stars, 2026-05

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Protein expression of ING4 with western blot analysis

Journal: Turkish Journal of Pharmaceutical Sciences

Article Title: Evaluation of a Synthetic Polyethyleneimine Based Polymeric Vector for ING4 Gene Delivery to MCF-7 Breast Cancer Cells

doi: 10.4274/tjps.galenos.2023.72430

Figure Lengend Snippet: Protein expression of ING4 with western blot analysis

Article Snippet: Primary antibody incubation was performed using an ING4 polyclonal antibody from Elabscience (E-AB-33309), followed by Horseradish peroxidase (HRP)-conjugated secondary antibody incubation, both conducted in TBS-T containing 0.5% dry milk either at room temperature for 1 hour or at 4 ° C overnight.

Techniques: Expressing, Western Blot

Figure 1. Loss of ING4 promoted tumor immune escape. A), The correlation of ING4 expression with T cell inﬁltration was assayed using TIME 2. http://timer.cistrome.org/. B), The correlation of ING4 expression with activated CD8+ T cell abundance was assayed using TISIDB (hku.hk). C), Im- munohistochemical analysis of ING4, PD-L1, and CD8 using LUSC tumor tissue specimens. Scale bar: 100 μm. The correlation of ING4 with PD-L1, and CD8 protein expression was assayed (n = 48). D), Western blot analysis of wild type (WT) or ING4−/- H520 cell lysates. The relative PD-L1 level

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Inhibition of CK2/ING4 Pathway Facilitates Non-Small Cell Lung Cancer Immunotherapy.

doi: 10.1002/advs.202304068

Figure Lengend Snippet: Figure 1. Loss of ING4 promoted tumor immune escape. A), The correlation of ING4 expression with T cell inﬁltration was assayed using TIME 2. http://timer.cistrome.org/. B), The correlation of ING4 expression with activated CD8+ T cell abundance was assayed using TISIDB (hku.hk). C), Im- munohistochemical analysis of ING4, PD-L1, and CD8 using LUSC tumor tissue specimens. Scale bar: 100 μm. The correlation of ING4 with PD-L1, and CD8 protein expression was assayed (n = 48). D), Western blot analysis of wild type (WT) or ING4−/- H520 cell lysates. The relative PD-L1 level

Article Snippet: Antibodies, Immunoprecipitation, and Western Blot: ING4 (10617-1- AP), CK2α (10992-1-AP), Tubulin (11224-1-AP), Lamp1(21997-1-AP), PDL1(66248-1), ATG7 (10088-2-AP), LC3B (14600-1-AP), GST (66001-2), and Flag (66008-4) antibodies were purchased from Proteintech.

Techniques: Expressing, Western Blot

Figure 2. ING4 induced PD-L1 autophagic degradation. A), H520 cells were transfected vector (pcDNA3) or Flag-ING4 plasmids for 48 h and Western blot analysis of cell lysates. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. B, H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-ING4/∆NLS plasmids for 48 h and Western blot analysis of cell lysates. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. C, H520 cells were transfected vector (pcDNA3) or Flag-ING4 plasmids for 48 h. Cells were treated with cycloheximide (CHX, 30 μg ml−1) as indicated time course. Cell lysates were subjected to Western blot analysis. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. D, WT or ING4−/- H520 cells were treated with CHX (30 μg ml−1) as indicated time course. Cell lysates were subjected to Western blot analysis. The relative PD-L1 level was quantiﬁed. Results are expressed as means ±

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Inhibition of CK2/ING4 Pathway Facilitates Non-Small Cell Lung Cancer Immunotherapy.

doi: 10.1002/advs.202304068

Figure Lengend Snippet: Figure 2. ING4 induced PD-L1 autophagic degradation. A), H520 cells were transfected vector (pcDNA3) or Flag-ING4 plasmids for 48 h and Western blot analysis of cell lysates. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. B, H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-ING4/∆NLS plasmids for 48 h and Western blot analysis of cell lysates. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. C, H520 cells were transfected vector (pcDNA3) or Flag-ING4 plasmids for 48 h. Cells were treated with cycloheximide (CHX, 30 μg ml−1) as indicated time course. Cell lysates were subjected to Western blot analysis. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. D, WT or ING4−/- H520 cells were treated with CHX (30 μg ml−1) as indicated time course. Cell lysates were subjected to Western blot analysis. The relative PD-L1 level was quantiﬁed. Results are expressed as means ±

Techniques: Transfection, Plasmid Preparation, Western Blot

Figure 3. The interaction of ING4 with PD-L1. A), Immunoprecipitation and Western blot analysis of H520 cell lysates. The relative binding of ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. B), The interaction of ING4 with PD-L1 was assayed by confocal in H520 cells. Scale bar: 25 μm. C), Schematic illustration of PD-L1. D), H520 cells were transfected vector (pcDNA3), his-PD-L1 or his-PD-L1/∆C plasmids as indicated for 48 h. Cell lysates were subjected to Ni-NTA pull-down and Western blot. The relative binding of his-PD-L1 to ING4 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. E), Schematic illustration of ING4. PHD: plant homeodomain. F), H520 cells were transfected PEBG (GST), GST-ING4, or GST-ING4/∆PHD plasmids as indicated. Cell lysates were subjected to GST pull-down and Western blot. The relative binding of GST-ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05.

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Inhibition of CK2/ING4 Pathway Facilitates Non-Small Cell Lung Cancer Immunotherapy.

doi: 10.1002/advs.202304068

Figure Lengend Snippet: Figure 3. The interaction of ING4 with PD-L1. A), Immunoprecipitation and Western blot analysis of H520 cell lysates. The relative binding of ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. B), The interaction of ING4 with PD-L1 was assayed by confocal in H520 cells. Scale bar: 25 μm. C), Schematic illustration of PD-L1. D), H520 cells were transfected vector (pcDNA3), his-PD-L1 or his-PD-L1/∆C plasmids as indicated for 48 h. Cell lysates were subjected to Ni-NTA pull-down and Western blot. The relative binding of his-PD-L1 to ING4 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. E), Schematic illustration of ING4. PHD: plant homeodomain. F), H520 cells were transfected PEBG (GST), GST-ING4, or GST-ING4/∆PHD plasmids as indicated. Cell lysates were subjected to GST pull-down and Western blot. The relative binding of GST-ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05.

Techniques: Immunoprecipitation, Western Blot, Binding Assay, Transfection, Plasmid Preparation

Figure 4. LIR motif of ING4 was required for PD-L1 autophagic degradation and anti-tumor immune escape. A), Immunoprecipitation and Western blot analysis of H520 cell lysates. The relative binding of ING4 to LC3B was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. B), Alignment of ING4 with LIR motif contained proteins, and schematic illustration of ING4 LIR motif. C), in vitro binding analysis of the interaction of ING4 with LC3B as described in methods. Down panel was ponceau S staining. The relative binding of ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. D, H520 cells were transfected PEBG (GST), GST-ING4, GST-F178A plasmids as indicated for 48 h. GST pull-down analysis was performed using cell lysates. The relative binding of ING4 to LC3B was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. E, WT or ATG7−/- H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-F178A plasmids for 48 h and Western blot analysis of cell lysates. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. F, H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-F178A plasmids for 48 cells, and co-localization of PD-L1 with LAMP1 was detected by confocal. Scale bar: 25 μm. Percent colocalization of PD-L1 with LAMP1 was quantiﬁed. Results are expressed as means ± SEM (n = 15 ﬁelds). *P<0.05. G,H, H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-F178A plasmids for 48 h and co-cultured with Jurkat cells for 24 h. IL-2 or IFN-𝛾release from Jurkat cells was assayed. Results are expressed as means ± SEM (n = 3). *P<0.05. I,J), Implanted tumor model analysis of vector (pLenti-CMV), ING4 or F178A stably expressing LLC cells. Tumor volume and weight were measured. Results are expressed as means ± SEM, n = 6. *P<0.05.

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Inhibition of CK2/ING4 Pathway Facilitates Non-Small Cell Lung Cancer Immunotherapy.

doi: 10.1002/advs.202304068

Figure Lengend Snippet: Figure 4. LIR motif of ING4 was required for PD-L1 autophagic degradation and anti-tumor immune escape. A), Immunoprecipitation and Western blot analysis of H520 cell lysates. The relative binding of ING4 to LC3B was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. B), Alignment of ING4 with LIR motif contained proteins, and schematic illustration of ING4 LIR motif. C), in vitro binding analysis of the interaction of ING4 with LC3B as described in methods. Down panel was ponceau S staining. The relative binding of ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. D, H520 cells were transfected PEBG (GST), GST-ING4, GST-F178A plasmids as indicated for 48 h. GST pull-down analysis was performed using cell lysates. The relative binding of ING4 to LC3B was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. E, WT or ATG7−/- H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-F178A plasmids for 48 h and Western blot analysis of cell lysates. The relative PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.01. F, H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-F178A plasmids for 48 cells, and co-localization of PD-L1 with LAMP1 was detected by confocal. Scale bar: 25 μm. Percent colocalization of PD-L1 with LAMP1 was quantiﬁed. Results are expressed as means ± SEM (n = 15 ﬁelds). *P<0.05. G,H, H520 cells were transfected vector (pcDNA3), Flag-ING4 or Flag-F178A plasmids for 48 h and co-cultured with Jurkat cells for 24 h. IL-2 or IFN-𝛾release from Jurkat cells was assayed. Results are expressed as means ± SEM (n = 3). *P<0.05. I,J), Implanted tumor model analysis of vector (pLenti-CMV), ING4 or F178A stably expressing LLC cells. Tumor volume and weight were measured. Results are expressed as means ± SEM, n = 6. *P<0.05.

Techniques: Immunoprecipitation, Western Blot, Binding Assay, In Vitro, Staining, Transfection, Plasmid Preparation, Cell Culture, Stable Transfection, Expressing

Figure 8. Inhibitor of CK2 enhanced antitumor immunotherapy. A), H520 cells were treated without or with TBB (10 μM), TBB (10 μM)+ CQ (30 μM), or TBB (10 μM)+ MG132 (20 μM) for 4 h and Western blot analysis of cell lysates. PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. B), WT or ATG7−/- H520 cells were treated without or with TBB (10 μM) for 4 h and Western blot analysis of cell lysates. PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. C), H520 cells were treated with TBB (10 μM) for 0, 0.5, and 2 h. Cell lysates were subjected to immunoprecipitation and Western blot. The relative binding of ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. D), WT or ING4−/- H520 cells were treated without or with TBB (10 μM) for 4 h and Western blot analysis of cell lysates. PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. E,F), co-cultured Jurkat cells with H520 cells were treated with TBB (10 μM) for 24 h. IL-2 or IFN-𝛾production from Jurkat cells was detected. Results are expressed as means ± SEM (n = 3). *P<0.05. G,H), LLC cells were inoculated subcutaneously C57BL/6 mice. Mice were treated control, TBB, anti-PD-1 mice monoclonal antibody, or TBB+anti-PD-1 antibody as described in method. Tumor volume and weight were detected. Results are expressed as means±SEM, n = 5. *P<0.05. I), Relative surface PD-L1 expression in LLC cell implanted tumors was assayed by ﬂow cytometry. MFI: median ﬂuorescence intensity. Results are expressed as means ± SEM (n = 5). *P<0.05. J), The percentage of CD8+ /IFN-𝛾+ T cells in LLC cell implanted tumors was assayed by ﬂow cytometry. Results are expressed as means ± SEM (n = 5). *P<0.05.

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Inhibition of CK2/ING4 Pathway Facilitates Non-Small Cell Lung Cancer Immunotherapy.

doi: 10.1002/advs.202304068

Figure Lengend Snippet: Figure 8. Inhibitor of CK2 enhanced antitumor immunotherapy. A), H520 cells were treated without or with TBB (10 μM), TBB (10 μM)+ CQ (30 μM), or TBB (10 μM)+ MG132 (20 μM) for 4 h and Western blot analysis of cell lysates. PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. B), WT or ATG7−/- H520 cells were treated without or with TBB (10 μM) for 4 h and Western blot analysis of cell lysates. PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. C), H520 cells were treated with TBB (10 μM) for 0, 0.5, and 2 h. Cell lysates were subjected to immunoprecipitation and Western blot. The relative binding of ING4 to PD-L1 was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. D), WT or ING4−/- H520 cells were treated without or with TBB (10 μM) for 4 h and Western blot analysis of cell lysates. PD-L1 level was quantiﬁed. Results are expressed as means ± SEM (n = 3). *P<0.05. E,F), co-cultured Jurkat cells with H520 cells were treated with TBB (10 μM) for 24 h. IL-2 or IFN-𝛾production from Jurkat cells was detected. Results are expressed as means ± SEM (n = 3). *P<0.05. G,H), LLC cells were inoculated subcutaneously C57BL/6 mice. Mice were treated control, TBB, anti-PD-1 mice monoclonal antibody, or TBB+anti-PD-1 antibody as described in method. Tumor volume and weight were detected. Results are expressed as means±SEM, n = 5. *P<0.05. I), Relative surface PD-L1 expression in LLC cell implanted tumors was assayed by ﬂow cytometry. MFI: median ﬂuorescence intensity. Results are expressed as means ± SEM (n = 5). *P<0.05. J), The percentage of CD8+ /IFN-𝛾+ T cells in LLC cell implanted tumors was assayed by ﬂow cytometry. Results are expressed as means ± SEM (n = 5). *P<0.05.

Techniques: Western Blot, Immunoprecipitation, Binding Assay, Cell Culture, Control, Expressing, Cytometry

ER+ breast cancer cells overexpressing ING4 are more sensitive to hormone deprivation and to tamoxifen treatment. Notes: T47D (A) or MCF7 (B) cells ectopically expressing the pMIG vector or ING4 were hormone-deprived in media containing CSS for 3 days (day 0, d0), followed by 7 days in media containing FS, CSS, CSS+10 nM E2, or CSS+E2 in the presence of OHT in 10, 100 nM, or 1 μM. Cell growth was assessed by SRB colorimetric assay. * p -value <0.001, ** p -value <0.002. Error bars represent minimal and maximal values. Abbreviations: ER+, estrogen receptor-positive; ING4, inhibitor of growth 4; CSS, charcoal-stripped serum; FS, full serum; E2, 17β-estradiol; OHT, 4-hydroxy tamoxifen; SRB, sulforhodamine B.

Journal: Breast Cancer : Targets and Therapy

Article Title: Tumor suppressor ING4 inhibits estrogen receptor activity in breast cancer cells

doi: 10.2147/BCTT.S119691

Figure Lengend Snippet: ER+ breast cancer cells overexpressing ING4 are more sensitive to hormone deprivation and to tamoxifen treatment. Notes: T47D (A) or MCF7 (B) cells ectopically expressing the pMIG vector or ING4 were hormone-deprived in media containing CSS for 3 days (day 0, d0), followed by 7 days in media containing FS, CSS, CSS+10 nM E2, or CSS+E2 in the presence of OHT in 10, 100 nM, or 1 μM. Cell growth was assessed by SRB colorimetric assay. * p -value <0.001, ** p -value <0.002. Error bars represent minimal and maximal values. Abbreviations: ER+, estrogen receptor-positive; ING4, inhibitor of growth 4; CSS, charcoal-stripped serum; FS, full serum; E2, 17β-estradiol; OHT, 4-hydroxy tamoxifen; SRB, sulforhodamine B.

Article Snippet: Nuclear and cytoplasmic fractions were analyzed by Western blot using antibodies against ERα (Cell Signaling, Danvers, MA, USA), ING4 (EMD Millipore, Temecula, CA, USA), histone H3 (Cell Signaling), and tubulin (Cell Signaling), and phospho-extracellular signal-regulated kinase (ERK) (Cell Signaling) at 1:1,000 dilution.

Techniques: Expressing, Plasmid Preparation, Colorimetric Assay

ING4 retards estrogen-dependent growth of T47D cells. Notes: (A) T47D cells ectopically expressing the pMIG vector or ING4 were grown in media containing FS, CSS, or CSS with 10 nM E2 for 14 days. Cells were fixed at each time point and relative cell numbers were determined by SRB colorimetric assay. pMIG, closed circle; ING4, open circle; FS, solid lines; CSS, long serrated lines; CSS+10 nM E2, short serrated lines. ( B ) Ectopic expression of ING4 did not affect EC50 but reduced minimal and maximal effective concentration of E2 for growth. Cells were grown in media containing CSS and various concentrations of E2 for 10 days. Cell growth was assessed by SRB assay and used to generate an EC50 curve. All values are normalized to the values of pMIG treated with 1 μM E2 (maximum E2 concentration). Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal-stripped serum; FS, full serum; E2, 17β-estradiol; SRB, sulforhodamine B; EC50, half-maximal effective concentration.

Journal: Breast Cancer : Targets and Therapy

Article Title: Tumor suppressor ING4 inhibits estrogen receptor activity in breast cancer cells

doi: 10.2147/BCTT.S119691

Figure Lengend Snippet: ING4 retards estrogen-dependent growth of T47D cells. Notes: (A) T47D cells ectopically expressing the pMIG vector or ING4 were grown in media containing FS, CSS, or CSS with 10 nM E2 for 14 days. Cells were fixed at each time point and relative cell numbers were determined by SRB colorimetric assay. pMIG, closed circle; ING4, open circle; FS, solid lines; CSS, long serrated lines; CSS+10 nM E2, short serrated lines. ( B ) Ectopic expression of ING4 did not affect EC50 but reduced minimal and maximal effective concentration of E2 for growth. Cells were grown in media containing CSS and various concentrations of E2 for 10 days. Cell growth was assessed by SRB assay and used to generate an EC50 curve. All values are normalized to the values of pMIG treated with 1 μM E2 (maximum E2 concentration). Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal-stripped serum; FS, full serum; E2, 17β-estradiol; SRB, sulforhodamine B; EC50, half-maximal effective concentration.

Techniques: Expressing, Plasmid Preparation, Colorimetric Assay, Concentration Assay, Sulforhodamine B Assay

$ING4 inhibits ERα transcription activity without affecting ERα protein expression. Notes: (A) Amounts of the nuclear ERα protein were comparable between the pMIG vector and ING4 expressing T47D or MCF7 cells. Cells were treated for 24 h with FS, CSS, CSS+10 nM E2, CSS+10 nM E2+1 μM OHT, or CSS+10 nM E2+100 nM ICI182,780. Total lysate, cytosolic, and nuclear fractions were analyzed by Western blot with antibodies against ING4, ERα, or phospho-ERK. Antibodies against histone H3 (nuclear) and α-tubulin (cytosolic) were used for loading control. ( B ) ING4 inhibits the expression of an ER-target gene, PDZK1 . PDZK1 mRNA levels in the cells treated with the same assay conditions in (A) were quantified by qRTPCR normalized to GAPDH as sample control. Relative expression was normalized to pMIG (FS) samples. “-” represents no OHT or ICI added. Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal stripped serum; FS, full serum; E2, 17β-estradiol; OHT, 4-hydroxy tamoxifen; ICI, ICI182,780; ER, estrogen receptor; qRTPCR, quantitative reverse transcription polymerase chain reaction; ERK, extracellular signal-regulated kinase.$

Journal: Breast Cancer : Targets and Therapy

Article Title: Tumor suppressor ING4 inhibits estrogen receptor activity in breast cancer cells

doi: 10.2147/BCTT.S119691

Figure Lengend Snippet: ING4 inhibits ERα transcription activity without affecting ERα protein expression. Notes: (A) Amounts of the nuclear ERα protein were comparable between the pMIG vector and ING4 expressing T47D or MCF7 cells. Cells were treated for 24 h with FS, CSS, CSS+10 nM E2, CSS+10 nM E2+1 μM OHT, or CSS+10 nM E2+100 nM ICI182,780. Total lysate, cytosolic, and nuclear fractions were analyzed by Western blot with antibodies against ING4, ERα, or phospho-ERK. Antibodies against histone H3 (nuclear) and α-tubulin (cytosolic) were used for loading control. ( B ) ING4 inhibits the expression of an ER-target gene, PDZK1 . PDZK1 mRNA levels in the cells treated with the same assay conditions in (A) were quantified by qRTPCR normalized to GAPDH as sample control. Relative expression was normalized to pMIG (FS) samples. “-” represents no OHT or ICI added. Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal stripped serum; FS, full serum; E2, 17β-estradiol; OHT, 4-hydroxy tamoxifen; ICI, ICI182,780; ER, estrogen receptor; qRTPCR, quantitative reverse transcription polymerase chain reaction; ERK, extracellular signal-regulated kinase.

Techniques: Activity Assay, Expressing, Plasmid Preparation, Western Blot, Reverse Transcription Polymerase Chain Reaction

ING4 inhibits ligand-independent ER activity. Notes: (A) ICI treatment, not OHT, of the vector pMIG expressing T47D cells results in diminished PDZK1 expression comparable to ING4 expressing cells. Cells were treated with 1 μM OHT or 1 μM ICI in FS, CSS, or CSS+E2, for 24 h and harvested for total RNA isolation. PDZK1 mRNA was quantified by qRTPCR, normalized to GAPDH . ( B ) ICI, not OHT, suppresses hormone-independent growth of pMIG expressing T47D cells in a dose-dependent manner. Cells were grown in CSS media for 10 days with OHT in incremental concentrations of 100 nM, 1, or 10 μM, or with ICI in incremental concentrations of 100 nM, 1, or 10 μM. “-” represents no OHT or ICI added. Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal stripped serum; FS, full serum; E2, 17β-estradiol; OHT, 4-hydroxy tamoxifen; ER, estrogen receptor; ICI, ICI182,780; qRTPCR, quantitative reverse transcription polymerase chain reaction.

Journal: Breast Cancer : Targets and Therapy

Article Title: Tumor suppressor ING4 inhibits estrogen receptor activity in breast cancer cells

doi: 10.2147/BCTT.S119691

Figure Lengend Snippet: ING4 inhibits ligand-independent ER activity. Notes: (A) ICI treatment, not OHT, of the vector pMIG expressing T47D cells results in diminished PDZK1 expression comparable to ING4 expressing cells. Cells were treated with 1 μM OHT or 1 μM ICI in FS, CSS, or CSS+E2, for 24 h and harvested for total RNA isolation. PDZK1 mRNA was quantified by qRTPCR, normalized to GAPDH . ( B ) ICI, not OHT, suppresses hormone-independent growth of pMIG expressing T47D cells in a dose-dependent manner. Cells were grown in CSS media for 10 days with OHT in incremental concentrations of 100 nM, 1, or 10 μM, or with ICI in incremental concentrations of 100 nM, 1, or 10 μM. “-” represents no OHT or ICI added. Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal stripped serum; FS, full serum; E2, 17β-estradiol; OHT, 4-hydroxy tamoxifen; ER, estrogen receptor; ICI, ICI182,780; qRTPCR, quantitative reverse transcription polymerase chain reaction.

Techniques: Activity Assay, Plasmid Preparation, Expressing, Isolation, Reverse Transcription Polymerase Chain Reaction

ING4 represses transcription of selective ER-target genes containing ERE. Notes: (A) Expression of a luciferase reporter construct containing an ERE promoter sequence in T47D cells. Cells were treated with CSS (−) for 2 days and additional 24 h with (E2) or without (−) 10 nM E2. ( B ) Relative mRNA expression of the ER-target genes, TFF1 , PDZK1 , MYC , and PGR in cells treated with CSS (−E2) or 10 nM E2 (+E2) for 4, 24, or 72 h. Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal stripped serum; FS, full serum; E2, 17β-estradiol; ER, estrogen receptor; ERE, estrogen response element.

Journal: Breast Cancer : Targets and Therapy

Article Title: Tumor suppressor ING4 inhibits estrogen receptor activity in breast cancer cells

doi: 10.2147/BCTT.S119691

Figure Lengend Snippet: ING4 represses transcription of selective ER-target genes containing ERE. Notes: (A) Expression of a luciferase reporter construct containing an ERE promoter sequence in T47D cells. Cells were treated with CSS (−) for 2 days and additional 24 h with (E2) or without (−) 10 nM E2. ( B ) Relative mRNA expression of the ER-target genes, TFF1 , PDZK1 , MYC , and PGR in cells treated with CSS (−E2) or 10 nM E2 (+E2) for 4, 24, or 72 h. Abbreviations: ING4, inhibitor of growth 4; CSS, charcoal stripped serum; FS, full serum; E2, 17β-estradiol; ER, estrogen receptor; ERE, estrogen response element.

Techniques: Expressing, Luciferase, Construct, Sequencing

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