Human COUP-TF II/NR2F2 Antibody

Detection of Human COUP-TF II/NR2F2 by Knockdown Validated Reduction of COUP-TFII or nucleolin decreases RAR beta 2 transcription in MCF-7 cells.MCF-7 (A) and T47D (B) cells were transfected with control siRNA or an siRNA targeting nucleolin for 48 h. T47D cells were treated with EtOH or 1 µM atRA for 24 h. Q-PCR for nucleolin (NCL) and RARB2. Values are the average of triplicates. C, Western blot showing COUP-TFII and RAR beta 2 expression after transfection with siCOUP-TFII. Values are relative to beta -actin. MCF-7 were transfected with siControl or siCOUP-TFII for 48 h and treated with 1 µM atRA for 6 h. Q-PCR was also performed for RRIG1. P<0.001 * versus control or ** versus atRA. Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0038278), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human COUP-TF II/NR2F2 by Knockdown Validated Loss of miR-101 promotes cancer metastasis through de-repression of COUP-TFII expression.(a) LNCaP and PC3 cells were individually treated with 50 nM of miR-101 and miR-27a mimic for 48 h and then invasion assays were performed for an additional 16 h (n=3). *P<0.05 (two-sided Student's t-test) compared with the control group. (b) Representative western blot showed the levels of COUP-TFII, E-cadherin, vimentin and beta -actin in PC3 cells stably knocked down by COUP-TFII. (c) LNCaP and PC3 cells stably knocked-down of COUP-TFII were used to perform cell invasion assay. Invaded cells were stained and counted (lower panel) (n=3). Representative western blot showed the knockdown efficiency of COUP-TFII (upper panel). *: P<0.05 (two-sided Student's t-test) compared with control group. (d) PC3 cells carrying an inducible miR-101 in the absence and presence of doxycycline, and re-expression of COUP-TFII were used to perform invasion assays (n=3). Representative western blot showed the levels of COUP-TFII and beta -actin (left). Invaded cells were counted and results are shown in the right panel. *: P<0.05 (two-sided Student's t-test) (e) 22RV-1 cells were treated with miR-101 inhibitor (antisense RNA) in conjunction with knockdown of COUP-TFII and used for invasion assays (n=3). Representative western blot show the levels of COUP-TFII and beta -actin (left). Invaded cells were counted and result is shown in the right panel. *: P<0.05 (two-sided Student's t-test). (f) LNCaP cells containing a construct with inducible expression of COUP-TFII shRNA and constructs expressing with anti-miR-101 and anti-mir-27a were orthotopically injected into NOD-SCID mouse prostate. In addition these cells also contain a luciferase reporter to detect cancer cells. After the tumour size was bigger than 50 mm3, drinking water with or without doxycycline (1 mg ml−1) was given to the mice for 6 weeks to induce the COUP-TFII shRNA to repress COUP-TFII expression. Representative bioluminescence results show the status of tumour growth in different groups. (g) Quantification result of luminance from IVIS (control: n=5; anti-miR-101/27a(−): n=6; anti-miR-101/27a(+): n=7. *: P<0.05 (two-sided Student's t-test). (h) Immunohistochemical stain showed the metastatic tumour cells located in the mouse lymph node are positive for AR. Scale bar: 100 μM (low power); and 200 μM (high power). SCID, severe combined immunodeficient. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/ncomms11418), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human COUP-TF II/NR2F2 by Knockdown Validated Loss of miR-101 promotes cancer metastasis through de-repression of COUP-TFII expression.(a) LNCaP and PC3 cells were individually treated with 50 nM of miR-101 and miR-27a mimic for 48 h and then invasion assays were performed for an additional 16 h (n=3). *P<0.05 (two-sided Student's t-test) compared with the control group. (b) Representative western blot showed the levels of COUP-TFII, E-cadherin, vimentin and beta -actin in PC3 cells stably knocked down by COUP-TFII. (c) LNCaP and PC3 cells stably knocked-down of COUP-TFII were used to perform cell invasion assay. Invaded cells were stained and counted (lower panel) (n=3). Representative western blot showed the knockdown efficiency of COUP-TFII (upper panel). *: P<0.05 (two-sided Student's t-test) compared with control group. (d) PC3 cells carrying an inducible miR-101 in the absence and presence of doxycycline, and re-expression of COUP-TFII were used to perform invasion assays (n=3). Representative western blot showed the levels of COUP-TFII and beta -actin (left). Invaded cells were counted and results are shown in the right panel. *: P<0.05 (two-sided Student's t-test) (e) 22RV-1 cells were treated with miR-101 inhibitor (antisense RNA) in conjunction with knockdown of COUP-TFII and used for invasion assays (n=3). Representative western blot show the levels of COUP-TFII and beta -actin (left). Invaded cells were counted and result is shown in the right panel. *: P<0.05 (two-sided Student's t-test). (f) LNCaP cells containing a construct with inducible expression of COUP-TFII shRNA and constructs expressing with anti-miR-101 and anti-mir-27a were orthotopically injected into NOD-SCID mouse prostate. In addition these cells also contain a luciferase reporter to detect cancer cells. After the tumour size was bigger than 50 mm3, drinking water with or without doxycycline (1 mg ml−1) was given to the mice for 6 weeks to induce the COUP-TFII shRNA to repress COUP-TFII expression. Representative bioluminescence results show the status of tumour growth in different groups. (g) Quantification result of luminance from IVIS (control: n=5; anti-miR-101/27a(−): n=6; anti-miR-101/27a(+): n=7. *: P<0.05 (two-sided Student's t-test). (h) Immunohistochemical stain showed the metastatic tumour cells located in the mouse lymph node are positive for AR. Scale bar: 100 μM (low power); and 200 μM (high power). SCID, severe combined immunodeficient. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/ncomms11418), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human COUP-TF II/NR2F2 by Knockdown Validated FOXM1 and CENPF are downstream targets of COUP-TFII in PCa.(a) Gene expression profile of COUP-TFII knockdown PC3 cells (GSE33182) was downloaded from GEO database and analysed using GenePattern. Genes with P<0.05, FOLD CHANGE >1.5-fold are represented as a heatmap. The gene symbol of FOXM1 and CENPF genes after knockdown of COUP-TFII are annotated in red colour. The others were FOXM1 and CENPF co-regulated genes annotated in black colour. siCtrl: knockdown scramble control; siCII: knockdown of COUP-TFII. (b) A representative western blot shows the level of COUP-TFII in PC3 cells transfected with two different siRNAs against COUP-TFII for 72 h (upper panel). Levels of FOXM1 and CENPF are quantified by RT-qPCR (lower panel) (n=3). *P<0.05 (two-sided Student's t-test) compared with the control group (siCON). (c) A cartoon shows the FOXM1 and CENPF loci. COUP-TFII-binding sites are annotated by a black rectangle (upper panel). ChIP results are shown in the PC3 cells treated with control and two different siRNAs against COUP-TFII for 72 h (lower panel) (n=3). *P<0.05 (two-sided Student's t-test) compared with control. (d) FOXM1 and CENPF promoter activities were measured in PC3 cells carrying an inducible COUP-TFII gene in the absence or the presence of doxycycline for 48 h (n=3). *P<0.05 (two-sided Student's t-test). (e) FOXM1 and CENPF promoter activities were measured in control or stably knocked-down COUP-TFII in PC3 cells for 48 h (n=3). *P<0.05 (two-sided Student's t-test). (f) PC3 cells carrying with inducible COUP-TFII-Flag gene were treated with or without doxycycline for 48 h and then transfected with individual siRNA against FOXM1 and CENPF or double knockdown of FOXM1 and CENPF for an additional 48 h. western blot (left panel) shows the level of COUP-TFII, CENPF and FOXM1 expression. Invasion assay (right panel) was performed by using those cells (n=3). *P<0.05 (two-sided Student's t-test) compared with control. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/ncomms11418), licensed under a CC-BY license. Not internally tested by R&D Systems.