Project description:Microarray analyses with cells/tissues overexpressing YAP have revealed many transcription targets of YAP (Dong et al, 2007; Zhao et al, 2008). However, as YAP induces transformation of non-cancerous cells, we thought many of known targets of YAP may be indirect consequence of transforming property of YAP. To identify the immediate transcription targets for YAP, we utilized immortalized mammary epithelial MCF-10A cells expressing a tamoxifen inducible, hyperactive (S127/381A) YAP mutant (MCF-10A ERT2-YAP 2SA). MCF-10A ERT2 and MCF-10A ERT2-YAP 2SA are generated. Each cell line was treated with 0.1% of ethanol (solvent) or 1uM of 4-hydroxytamoxifen for 2 or 6 hours. This makes 6 samples per set. The experiments were done in duplicate. The expression data from MCF-10A ERT2 and MCF-10A ERT2-YAP 2SA before tamoxifen treatment can serve as control.
Project description:To study the function of 14-3-3ζ, we established MCF-10A human mammary epithelial cells transduced with 14-3-3ζ (10A.ζ) and vector (10A.Vec) We performed gene expression profiling on 10A.ζ cells and 10A.Vec cells, and normalized to profiling of MCF-10A parental cells
Project description:p63 is critical for epithelial development yet little is known about the transcriptional programmes it regulates. The p63 transactivating (TA) isoforms contain an amino-terminal exon that encodes a p53-like transactivation domain, whereas ÎN-isoforms lack this domain but contain the common DNA binding domain (DBD), suggesting that TAp63 and ÎNp63 isoforms may have opposing functions. By characterising transcriptional changes and cellular effects following modulation of p63 expression, we have defined a vital role for p63 in cellular adhesion. Knockdown of p63 expression caused downregulation of cell adhesion-associated genes, cell detachment and anoikis in mammary epithelial cells and keratinocytes. Conversely, overexpression of the TAp63γ or ÎNp63α isoforms of p63 upregulated cell adhesion molecules, increased cellular adhesion and conferred resistance to anoikis. Total RNA was isolated 48 h after retroviral or adenoviral infection and was subjected to reverse transcription, labelling and hybridization. The knockdown samples were performed in duplicate with shRNA vector control, shRNA targetting all isoforms of p63 (shDBD) and shRNA targetting p63 isoforms containing the transactivating (TA) domains. The overexpression samples were performed in triplicate with vector control, overexpression of the ÎNp63α isoform, and overexpression of the TAp63γ isoforms.
Project description:In this study, we examined temporal changes in gene expression during acinar morphogenesis of normal-like human mammary epithelial cells (MCF-10A) in a three-dimensional (3D) basement membrane cultures. Changes in gene expression in 3D culture of MCF-10A cells were measured at 4, 8, and 12 days.
Project description:Recent studies suggest that PDEF is required for secretory cell differentiation in several epithelial tissues. To investigate PDEF in the mammary gland, we examined the effect of this transcription factor on gene expression using microarray based profiling of MCF-10A cells. These cells are non-transformed mammary epithelial cells that express protein and gene expression programs of basal epithelial cells and undetectable levels of endogenous PDEF. Bioinformatics analysis of the genes induced or repressed by PDEF overexpression in MCF10A cells revealed a striking effect on expression of luminal and myoepithelial cell markers.
Project description:We profiled gene expression and splicing changes in MCF-10A human mammary epithelial cells expressing MYC fused to a portion of estrogen receptor (MYC-ER) (Eilers et al., 1989; Littlewood et al., 1995). We performed RNA-seq, in triplicate, on 3D-grown MCF-10A MYC-ER cells at 0, 8, and 24 hours (h) after 4-OHT-induced MYC activation. As a control for 4-OHT-induced effects, 3D-grown parental MCF-10A cells lacking the MYC-ER fusion protein were treated with 4-OHT at the same timepoints.
