antibodies against ing4  (Proteintech)

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

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.

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, Concentration Assay, Sulforhodamine B Assay

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.

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: Activity Assay, Expressing, Plasmid Preparation, Western Blot, 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.

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: Activity Assay, Plasmid Preparation, Expressing, Isolation, 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 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.

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, Luciferase, Construct, Sequencing

Journal: Journal of Ovarian Research

Article Title: Combinatorial strategies based on CRAd-IL24 and CRAd-ING4 virotherapy with anti-angiogenesis treatment for ovarian cancer

doi: 10.1186/s13048-016-0248-5

Figure Lengend Snippet: Validation of CRAd-mediated expression of IL-24 and ING4 genes. a ) The concentrations of IL-24 protein following infection of the indicated OvCa cell lines and normal IOSE523 cells with CRAd-IL24 at the MOIs of 1 and 10 vp/cell were determined in cell culture supernatants 3 days postinfection using commercial ELISA kit with IL-24 concentration standards. Each bar represents the cumulative mean ± SD (* p ≤ 0.05). b) The relative levels of ING4 gene expression were determined following infection of SKOV3ip.1 cells with CRAd-ING4 at the MOIs of 100, 33, and 11 vp/cell. The ING4 protein band of 29 kDa was detected in cell lysates 3 days postinfection with 100, 33, and 11 vp/cell, but not in mock-infected cells (0 vp/cell) using Western blot with rabbit polyclonal Ab raised against ING4 internal region

Article Snippet: Electrophoretically resolved proteins were transferred to a polyvinylidene fluoride membrane and analyzed for the presence of ING4 polypeptides using polyclonal rabbit antibody raised against ING4 internal region (Assay Biotechnology Company, Inc) diluted 1:1000 for overnight incubation at 4 °C.

Techniques: Biomarker Discovery, Expressing, Infection, Cell Culture, Enzyme-linked Immunosorbent Assay, Concentration Assay, Gene Expression, Western Blot