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MCF10A cells in 3-D culture for 15 h after treatment with BRCA1-RNAi vs Luc-RNAi adenovirus

ABSTRACT: BRCA1 exerts transcriptional repression through interaction with CtIP in the C-terminal BRCT domain and ZBRK1 in the central domain. A dozen of genes including angiopoietin-1 (ANG1), a secreted angiogenic factor, are co-repressed by BRCA1 and CtIP based on microarray analysis of mammary epithelial cells in 3-D culture. BRCA1, CtIP and ZBRK1 form a complex that coordinately represses ANG1 expression via a ZBRK1 recognition site in ANG1 promoter. Impairment of this complex upregulates ANG1, which stabilizes endothelial cells forming capillary-like network structure. Consistently, Brca1-deficient mouse mammary tumors exhibit accelerated growth, pronounced vascularization and overexpressed ANG1. These results suggest, besides its role in maintaining genomic stability, BRCA1 directly regulates the expression of angiogenic factors to modulate the tumor microenvironment. Keywords: MCF10A, human mammary epithelial cells, 3-D Matrigel, 15 h, BRCA1-RNAi

ORGANISM(S): Homo sapiens

PROVIDER: GSE4750 | GEO | 2006/05/02

SECONDARY ACCESSION(S): PRJNA104541

REPOSITORIES: GEO

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MCF10A cells in 3-D culture for 15 h after treatment with CtIP-RNAi vs Luc-RNAi adenovirus

2006-05-02 | GSE4751 | GEO

RNAi knock down of human breast cancer cell line MCF10A with BCRA1 or CtIP-RNAi reveals removal of the BRCA1/CtIP/ZBRK1 repressor complex on ANG1 promoter leads to accelerated mammary tumor growth contributed by prominent vasculature

Project description:BRCA1 exerts transcriptional repression through interaction with CtIP in the C-terminal BRCT domain and ZBRK1 in the central domain. A dozen genes, including angiopoietin-1 (ANG1), a secreted angiogenic factor, are corepressed by BRCA1 and CtIP based on microarray analysis of mammary epithelial cells in 3D culture. BRCA1, CtIP, and ZBRK1 form a complex that coordinately represses ANG1 expression via a ZBRK1 recognition site in the ANG1 promoter. Impairment of this complex upregulates ANG1, which stabilizes endothelial cells that form a capillary-like network structure. Consistently, Brca1-deficient mouse mammary tumors exhibit accelerated growth, pronounced vascularization, and overexpressed ANG1. These results suggest that, besides its role in maintaining genomic stability, BRCA1 directly regulates the expression of angiogenic factors to modulate the tumor microenvironment.

2007-10-26 | E-GEOD-4754 | biostudies-arrayexpress

RNAi profiling of human MCF10A cells in 3-D culture for 15 h after treatment with BRCA1-RNAi vs Luc-RNAi adenovirus

Project description:BRCA1 exerts transcriptional repression through interaction with CtIP in the C-terminal BRCT domain and ZBRK1 in the central domain. A dozen of genes including angiopoietin-1 (ANG1), a secreted angiogenic factor, are co-repressed by BRCA1 and CtIP based on microarray analysis of mammary epithelial cells in 3-D culture. BRCA1, CtIP and ZBRK1 form a complex that coordinately represses ANG1 expression via a ZBRK1 recognition site in ANG1 promoter. Impairment of this complex upregulates ANG1, which stabilizes endothelial cells forming capillary-like network structure. Consistently, Brca1-deficient mouse mammary tumors exhibit accelerated growth, pronounced vascularization and overexpressed ANG1. These results suggest, besides its role in maintaining genomic stability, BRCA1 directly regulates the expression of angiogenic factors to modulate the tumor microenvironment. Experiment Overall Design: MCF10A cells seeded at 5x105 cells/60mm plate were infected with adenoviral luciferase- or BRCA1-RNAi in duplicate at 20 MOI for 24h. Infected cells were re-seeded at 5x105 cells in a 60mm plate pre-coated with Growth Factor Reduced Matrigel and covered with the growth medium containing 2% Matrigel at 37Â°C for 15h. RNA was extracted and quality assessed. cDNA synthesized from the harvested RNA was biotin-labeled, hybridized onto Affymetrix HG U133 PLUS 2.0 array (54,676 genes) and stained with streptavidin-phycoerythrin. The hybridized array was analyzed using GeneChipÂ® Scanner 3000 and GCOS 1.2 software (Affymetrix) at the UCI Microarray Core service.

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Transcription profiling of human MCF10A cells in 3-D culture for 15 h after treatment with CtIP-RNAi vs Luc-RNAi adenovirus

Project description:BRCA1 exerts transcriptional repression through interaction with CtIP in the C-terminal BRCT domain and ZBRK1 in the central domain. A dozen of genes including angiopoietin-1 (ANG1), a secreted angiogenic factor, are co-repressed by BRCA1 and CtIP based on microarray analysis of mammary epithelial cells in 3-D culture. BRCA1, CtIP and ZBRK1 form a complex that coordinately represses ANG1 expression via a ZBRK1 recognition site in ANG1 promoter. Impairment of this complex upregulates ANG1, which stabilizes endothelial cells forming capillary-like network structure. Consistently, Brca1-deficient mouse mammary tumors exhibit accelerated growth, pronounced vascularization and overexpressed ANG1. These results suggest, besides its role in maintaining genomic stability, BRCA1 directly regulates the expression of angiogenic factors to modulate the tumor microenvironment. Experiment Overall Design: MCF10A cells seeded at 5x105 cells/60mm plate were infected with adenoviral luciferase- or CtIP-RNAi in duplicate at 20 MOI for 24h. Infected cells were re-seeded at 5x105 cells in a 60mm plate pre-coated with Growth Factor Reduced Matrigel and covered with the growth medium containing 2% Matrigel at 37Â°C for 15h. RNA was extracted and quality assessed. cDNA synthesized from the harvested RNA was biotin-labeled, hybridized onto Affymetrix HG U133 PLUS 2.0 array (54,676 genes) and stained with streptavidin-phycoerythrin. The hybridized array was analyzed using GeneChipÂ® Scanner 3000 and GCOS 1.2 software (Affymetrix) at the UCI Microarray Core service.

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