ncoa1 antibody  (Cell Signaling Technology Inc)

Journal: Journal of Gastrointestinal Oncology

Article Title: Bromodomain-containing protein 9 activates proliferation and epithelial-mesenchymal transition of colorectal cancer via the estrogen pathway in vivo and in vitro

doi: 10.21037/jgo-23-271

Figure Lengend Snippet: Inactivation of the estrogen pathway contributed to BRD9 knockdown-induced inhibition of proliferation in CRC cells. (A) KEGG pathway analysis based on the BRD9-binding and interacting genes based on TCGA databases. (B) The cnetplot for the molecular function data in GO analysis is based on TCGA databases. (C) GSEA plot showing that BRD9 expression is negatively correlated with ESTROGEN_DEPENDENT_GENE_EXPRESS. (D,E) BRD9 expression positively correlated with NCOA1. (F) Overexpression of BRD9 in SW480 cells and downregulation of BRD9 in HCT116 cells. Western blot was used to determine the protein levels of BRD9, p-MEK, MEK, p-ERK, ERK, and NCOA1. The relative levels of BRD9 (G), the ratio of p-MEK/MEK (H), the ratio of p-ERK/ERK (I), and NCOA1 (J) were measured using ImageJ and normalized to GAPDH. Data are presented as mean ± SD from at least three independent experiments. **P<0.01 and ***P < 0.001 compared with the indicated groups. CRC, colorectal cancer; GO, Gene Ontology; GSEA, gene set enrichment analysis; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; p-MEK, phosphorylated-MEK; p-ERK, phosphorylated-ERK; SD, standard deviation; BP, biological process; CC, cell component; MF, molecular function; KEGG, Kyoto Encyclopedia of Genes and Genomes; NES, normalized enrichment score; FDR, false discovery rate; BRD9, bromodomain-containing protein 9; TPM, transcript per million; NCOA1, nuclear receptor coactivator 1.

Article Snippet: The primary antibodies used included the following: anti-BRD9 antibodies (Abcam, Cambridge, MA, USA; ab137245, 1:1,000), anti-GAPDH antibodies (Abcam, ab9485, 1:1,000), anti-E-cadherin antibodies (Cell Signaling Technology, Danvers, MA, USA; #14472, 1:1,000), anti-N-cadherin antibodies (Cell Signaling Technology, Danvers, RA, USA; #13116, 1:1,000), anti-Vimentin antibodies (Cell Signaling Technology, Danvers, RA, USA; #5741, 1:1,000), anti-MEK2 antibodies (Abcam, ab32517, 1:1,000), anti-ERK antibodies (Abcam, ab32537, 1:1,000), anti-p-MEK1/2 antibodies (Cell Signaling Technology, Danvers, RA, USA; #75262, 1:2,000), anti-p-ERK antibodies (Cell Signaling Technology, Danvers, RA, USA; #4370, 1:2,000), and anti-NCOA1 antibodies (Cell Signaling Technology, Danvers, RA, USA; #2191, 1:1,000).

Techniques: Knockdown, Inhibition, Binding Assay, Expressing, Over Expression, Western Blot, Standard Deviation

Journal: Journal of Gastrointestinal Oncology

Article Title: Bromodomain-containing protein 9 activates proliferation and epithelial-mesenchymal transition of colorectal cancer via the estrogen pathway in vivo and in vitro

doi: 10.21037/jgo-23-271

Figure Lengend Snippet: Knockdown of BRD9 inhibits tumorigenesis in vivo . (A) SW480 cells transfected with control vector or shBRD9 were subcutaneously inoculated into BALB/c nude mice, and three weeks later the tumors were carefully dissected. (B) The average weight and volume of tumors were measured in both groups. (C) A schematic diagram depicting the role of BRD9 in promoting CRC progression via regulating the estrogen pathway. This diagram was reprinted using Figdraw ( https://www.figdraw.com/static/index.html ) with permission number 533420148. Data are presented as mean ± SD from at least three independent experiments. **P<0.01 compared with the control group. BRD9, bromodomain-containing protein 9; CRC, colorectal cancer; sh, short hairpin; GRB2, growth factor receptor-bound protein 2; SOS, son of sevenless; MEK, mitogen-activated protein kinase 1, ERK, mitogen-activated protein kinase 1/3; ESR1, estrogen receptor 1; NCOA1, nuclear receptor coactivator 1; SD, standard deviation.

Article Snippet: The primary antibodies used included the following: anti-BRD9 antibodies (Abcam, Cambridge, MA, USA; ab137245, 1:1,000), anti-GAPDH antibodies (Abcam, ab9485, 1:1,000), anti-E-cadherin antibodies (Cell Signaling Technology, Danvers, MA, USA; #14472, 1:1,000), anti-N-cadherin antibodies (Cell Signaling Technology, Danvers, RA, USA; #13116, 1:1,000), anti-Vimentin antibodies (Cell Signaling Technology, Danvers, RA, USA; #5741, 1:1,000), anti-MEK2 antibodies (Abcam, ab32517, 1:1,000), anti-ERK antibodies (Abcam, ab32537, 1:1,000), anti-p-MEK1/2 antibodies (Cell Signaling Technology, Danvers, RA, USA; #75262, 1:2,000), anti-p-ERK antibodies (Cell Signaling Technology, Danvers, RA, USA; #4370, 1:2,000), and anti-NCOA1 antibodies (Cell Signaling Technology, Danvers, RA, USA; #2191, 1:1,000).

Techniques: Knockdown, In Vivo, Transfection, Control, Plasmid Preparation, Standard Deviation

Journal: Reproductive biomedicine online

Article Title: METTL3 and METTL14-mediated N 6 -methyladenosine modification promotes cell proliferation and invasion in a model of endometriosis.

doi: 10.1016/j.rbmo.2022.10.010

Figure Lengend Snippet: FIGURE 3 Validation of the N6-methyladenosine (m6A) methylation level and mRNA level of the genes enriched in estrogen signalling. (A) The m6A methylation level of MMP2, NCOA1, TLR4, PPP2R2D and YWHAZ in the ectopic endometrium (black bar) and eutopic endometrium (EU) (hatched gray bar) was validated using MeRIP quantitative polymerase chain reaction; (B) the expression of MMP2, NCOA1, TLR4, PPP2R2D and YWHAZ in the ectopic endometrium (black bar) and eutopic endometrium (hatched gray bar) using quantitative reverse transcription polymerase chain reaction; and (C) immunohistochemistry was carried out to assess the protein levels of MMP2, NCOA1, PPP2R2D and YWHAZ in the ectopic endometrium and eutopic endometrium; magnification, × 200. Two-tailed Student's t-tests were used for comparisons between ectopic endometrium and eutopic endometrium. All samples were assayed in triplicate.

Article Snippet: The polyclonal rabbit immunoglobulin G (IgG) antibody against human NCOA1 (51114-1-AP) (ProteinTech, Rosemont, IL, USA), polyclonal rabbit IgG antibody against human MMP2 (10373-2-AP) (ProteinTech, Rosemont, IL, USA), polyclonal rabbit antibody against human PPP2R2D (bs-19968P) (BiosSS Woburn, MA, USA), polyclonal rabbit IgG antibody against human YWHAZ (14881-1-AP) (Proteintech, Sankt LeonRot, Germany), were diluted at 1:1000.

Techniques: Biomarker Discovery, Methylation, Real-time Polymerase Chain Reaction, Expressing, Reverse Transcription, Polymerase Chain Reaction, Immunohistochemistry, Two Tailed Test

Journal: Reproductive biomedicine online

Article Title: METTL3 and METTL14-mediated N 6 -methyladenosine modification promotes cell proliferation and invasion in a model of endometriosis.

doi: 10.1016/j.rbmo.2022.10.010

Figure Lengend Snippet: FIGURE 5 Knockdown of METTL3, METTL14, or both, contributes to the cellular phenotype of endometriosis. (A–B) Quantitative polymerase chain reaction and Western blotting analysis of the expression of NCOA1, MMP2 For photomicrographs, please provide scale bars. PPP2R2D and YWHAZ after the knockdown of METTL3 and(or) METTL14. Cell viability assay of the cells with METTL3 and METTL14 knockdown; (C) the CCK-8 assay of the cell viability after the knockdown of METTL3 and METTL14; (D) the scratch assay of the cell migration after the knockdown of METTL3 and METTL14; (E) transwell assay of the effect of the METTL3 and METTL14 knockdown on the invasion and migration of endometrial stromal cells. NC: negative control; si-METTL14 was transfected with siMETTL14; si-METTL3 was transfected with siMETTL3; si-METTL3+si-METTL14 were transfected with si-METTL3 and si-METTL14. All samples were assayed in triplicate.

Article Snippet: The polyclonal rabbit immunoglobulin G (IgG) antibody against human NCOA1 (51114-1-AP) (ProteinTech, Rosemont, IL, USA), polyclonal rabbit IgG antibody against human MMP2 (10373-2-AP) (ProteinTech, Rosemont, IL, USA), polyclonal rabbit antibody against human PPP2R2D (bs-19968P) (BiosSS Woburn, MA, USA), polyclonal rabbit IgG antibody against human YWHAZ (14881-1-AP) (Proteintech, Sankt LeonRot, Germany), were diluted at 1:1000.

Techniques: Knockdown, Real-time Polymerase Chain Reaction, Western Blot, Expressing, Viability Assay, CCK-8 Assay, Wound Healing Assay, Migration, Transwell Assay, Negative Control, Transfection