Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling by array of human MCF10A cells expressing YAP to identify the targets of the YAP oncogene

ABSTRACT: YAP is an oncogene and an inducer of Epithelial-to-Mesenchymal Transition (EMT). We used microarrays to detail the global program of gene expression to identify YAP target genes. PUBLICATION ABSTRACT:; The Hippo pathway defines a novel signaling cascade regulating cell proliferation and survival in Drosophila, which involves the negative regulation of the transcriptional coactivator Yorkie by the kinases Hippo and Warts. We have recently shown that the human ortholog of Yorkie, YAP, maps to a minimal amplification locus in mouse and human cancers, and that it mediates dramatic transforming activity in MCF10A primary mammary epithelial cells. Here we show that LATS proteins (mammalian orthologs of Warts) interact directly with YAP in mammalian cells and that ectopic expression of LATS1, but not LATS2, effectively suppresses the YAP phenotypes. Furthermore, shRNA-mediated knockdown of LATS1 phenocopies YAP overexpression. Since this effect can be suppressed by simultaneous YAP knockdown, it suggests that YAP is the primary target of LATS1 in mammalian cells. Expression profiling of genes induced by ectopic expression of YAP or by knockdown of LATS1 reveals a subset of potential Hippo pathway targets implicated in epithelial-to-mesenchymal transition (EMT), suggesting that this is a key feature of YAP signaling in mammalian cells. Experiment Overall Design: MCF10A cells were infected with retrovirus constructs (vector or YAP) and puromycin was used to select for transduced cells. Cells were split and grown to ~60-75%% confluency at which point they were harvested for RNA. Vector vs. YAP comparison was done in duplicate.

ORGANISM(S): Homo sapiens

SUBMITTER: Daniel Haber

PROVIDER: E-GEOD-10196 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Negative regulation of YAP by LATS1 underscores evolutionary conservation of the Drosophila Hippo pathway.

Zhang Jianmin J Smolen Gromoslaw A GA Haber Daniel A DA

Cancer research 20080401 8

The Hippo pathway defines a novel signaling cascade regulating cell proliferation and survival in Drosophila, which involves the negative regulation of the transcriptional coactivator Yorkie by the kinases Hippo and Warts. We have recently shown that the human ortholog of Yorkie, YAP, maps to a minimal amplification locus in mouse and human cancers, and that it mediates dramatic transforming activity in MCF10A primary mammary epithelial cells. Here, we show that LATS proteins (mammalian ortholog ...[more]

PMID: 18413746

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Project description:Regulation of organ size is important for development and tissue homeostasis. In Drosophila, Hippo signaling controls organ size by regulating the activity of a TEAD transcription factor, Scalloped, through modulation of its coactivator protein Yki. The role of mammalian Tead proteins in growth regulation, however, remains unknown. Here we examined the role of mouse Tead proteins in growth regulation. In NIH3T3 cells, cell density and Hippo signaling regulated the activity of Tead proteins by modulating nuclear localization of a Yki homologue, Yap, and the resulting change in Tead activity altered cell proliferation. Tead2-VP16 mimicked Yap overexpression, including increased cell proliferation, reduced cell death, promotion of EMT, lack of cell contact inhibition, and promotion of tumor formation. Growth promoting activities of various Yap mutants correlated with their Tead-coactivator activities. Tead2-VP16 and Yap regulated largely overlapping sets of genes. However, only a few of the Tead/Yapregulated genes in NIH3T3 cells were affected in Tead1-/-;Tead2-/- or Yap-/- embryos. Most of the previously identified Yap-regulated genes were not affected in NIH3T3 cells or mutant mice. In embryos, levels of nuclear Yap and Tead1 varied depending on cell types. Strong nuclear accumulation of Yap and Tead1 were seen in myocardium, correlating with requirements of Tead1 for proliferation. However, their distribution did not always correlate with proliferation. Taken together, mammalian Tead proteins regulate cell proliferation and contact inhibition as a transcriptional mediator of Hippo signaling, but the mechanisms by which Tead/Yap regulate cell proliferation differ depending on cell types, and Tead, Yap and Hippo signaling may play multiple roles in mouse embryos. We used microarrays to know the gene expression profiles regurated by Tead2-VP16, Tead2-EnR, Yap, and cell density in NIH3T3 cells. Keywords: Cell density, genetic modification Tead2-VP16-, Tead2-EnR-, Yap- and control vector-expressing cells were cultured at low or high density for RNA extraction and hybridization on Affymetrix microarrays.

Dataset Information

Transcription profiling by array of human MCF10A cells expressing YAP to identify the targets of the YAP oncogene

Publications

Negative regulation of YAP by LATS1 underscores evolutionary conservation of the Drosophila Hippo pathway.

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