Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Conversion of Fibroblasts into Functional Trophoblast Stem-like Cells

ABSTRACT: Here we demonstrate the generation of stable induced trophoblast stem cells (iTSCs) from fibroblasts by the transient expression of Gata3, Eomes and Tfap2c. Transcriptome and methylome analyses and functional assays such as hemorrhagic lesion formation and placenta contribution suggested a high degree of conversion. Careful examination of the reprogramming process indicated that the cells did not go through a transient pluripotent state. Detect and compare different H2A.X deposition patterns in ES cells,MEFs and iTSC and TS cells, with Illumina HiSeq 2000

ORGANISM(S): Mus musculus

SUBMITTER: Tao Wu

PROVIDER: E-GEOD-70234 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Extensive Nuclear Reprogramming Underlies Lineage Conversion into Functional Trophoblast Stem-like Cells.

Benchetrit Hana H Herman Shay S van Wietmarschen Niek N Wu Tao T Makedonski Kirill K Maoz Noam N Yom Tov Nataly N Stave Danielle D Lasry Rachel R Zayat Valery V Xiao Andrew A Lansdorp Peter M PM Sebban Shulamit S Buganim Yosef Y

Cell stem cell 20150924 5

Induced pluripotent stem cells (iPSCs) undergo extensive nuclear reprogramming and are generally indistinguishable from embryonic stem cells (ESCs) in their functional capacity and transcriptome and DNA methylation profiles. However, direct conversion of cells from one lineage to another often yields incompletely reprogrammed, functionally compromised cells, raising the question of whether pluripotency is required to achieve a high degree of nuclear reprogramming. Here, we show that transient ex ...[more]

PMID: 26412562

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Project description:It is well-known that embryonic stem cells (ESC) are much more sensitive to replication-induced stress than differentiated cells but the underpinning mechanisms are largely unknown. H2A.X, a minor variant of H2A, constitutes only 1-10% of the mammalian genome. H2A.X plays a well-known for role in the DNA damage response and maintaining stability in the genome, including the regions frequently experiencing replication stress, such as the fragile sites. Intriguingly, several recent studies have reported that H2A.X function is elevated in ESC; and others reported that H2A.X function is provoked during cellular reprogramming (in induced pluripotent stem cells, iPSC), indicating that increased proliferation during iPS may trigger replication stress and the H2A.X DNA damage response. However, several studies of genomic instability in iPSC led to different conclusions on this important issue. For example, frequent copy number variants (CNV) were reported at the genomic regions sensitive to replication stress, such as the fragile sites. On the other hand, another study reported the lack of genomic instability in mouse iPS clones that are able to generate “all-iPS” animals in tetraploid complementation assays (4N+ iPSC), indicative of a potential link between pluripotency and genome integrity. However, whether if high level genomic instability occurs in the 4N- iPSC iPSC clones at replication stress sensitive regions is unknown. Moreover, due to the lack of mechanistic insights on genome integrity maintenance, how pluripotency and genome integrity are connected remains elusive. Here we show that H2A.X plays unexpected roles in maintaining pluripotency and genome integrity in ESC and iPSC. In ESC, it is specially enriched at genomic regions sensitive to replication stress so that it protects genome integrity thereat. Faithful H2A.X deposition is critical for genome integrity and pluripotency in iPSC. H2A.X depositions in 4N+ iPSC clones faithfully recapitulate the ESC pattern and therefore, prevent genome instability. On the other hand, insufficient H2A.X depositions in 4N- iPSC clones at such regions lead to genome instability and defects in replication stress response and DNA repair, reminiscent of the H2A.X deficient ESC. Detect and compare different H2A.X deposition patterns in ES cells and iPS cells, with Illumina HiSeq 2000 and Illumina Genome Analyzer IIx

Project description:Purpose: to characterize epigenetic changes following Twist1 mediated Epithelial-Mesenchymal Transition in human Methods: we characterized the epigenetic and transcriptome landscapes using whole genome transcriptome analysis by RNA-seq, DNA methylation by digital restriction enzyme analysis of methylation (DREAM) and histone modifications by CHIP-seq of H3K4me3 and H3K27me3 in immortalized human mammary epithelial cells relative to cells induced to undergo EMT by Twist1. Results: EMT is accompanied by focal hypermethylation and widespread global DNA hypomethylation, predominantly within transcriptionally repressed gene bodies. At the chromatin level, the number of gene promoters marked by H3K4me3 increases by more than one fifth; H3K27me3 undergoes dynamic genomic redistribution characterized by loss at half of gene promoters and overall reduction of peak size by almost one-half. This is paralleled by increased phosphorylation of EZH2 at serine 21. Among genes with highly altered mRNA expression, 23.1% switch between H3K4me3 and H3K27me3 marks, and those point to the master EMT targets and regulators CDH1, PDGFRA and ESRP1. Strikingly, Twist1 increases the number of bivalent genes by more than two fold. Inhibition of the H3K27 methyltransferases EZH2 and EZH1, which form part of the PRC2 complex, results in blocking EMT and stemness properties. Conclusion: Our findings demonstrate that the EMT program requires epigenetic remodeling by the Polycomb/Trithorax complexes leading to increased cellular plasticity which suggests that its inhibition will prevent EMT, and the associated breast cancer metastasis. ChIPseq profiles of human mammary epithelial cells before (HMLE_parental) and after Twist1 transfection (HMLE_Twist) were generated in monolayer (HMLE_Twist2D) and sphere culture by deep sequencing using Illumina GAIIx or Illumina hiseq2000 Please note that the processed data files were generated by pooling both replicate samples after confirming the high correlation of the two replicates.

Dataset Information

Conversion of Fibroblasts into Functional Trophoblast Stem-like Cells

Publications

Extensive Nuclear Reprogramming Underlies Lineage Conversion into Functional Trophoblast Stem-like Cells.

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