Project description:The transcription factor Oct3/4 is essential to maintain pluripotency in mouse embryonic stem (ES) cells. It was reported that the Xpc DNA repair complex is involved in this process. Here we examined the role of Xpc on the transcriptional activation of the target genes by Oct3/4 using the inducible knockout strategy. We found that the removal of the C-terminal region of Xpc, including the interaction sites with Rad23 and Cetn2, showed faint impact on the gene expression profile of ES cells and the functional Xpc-ΔC ES cell lines retained proper gene expression profile as well as pluripotency to contribute chimeric embryos. These data indicated that the C-terminal region of Xpc is dispensable for the transcriptional activity of Oct3/4 in mouse ES cells. An inducible knockout ES cell line of Xpc with the Cre-loxP system was generated and gene expression was measured without Xpc deletion, after 4 days of Xpc deletion or constitutive knocked out cells. Each sample was prepared in triplicate.
Project description:The transcription factor Oct3/4 is essential to maintain pluripotency in mouse embryonic stem (ES) cells. It was reported that the Xpc DNA repair complex is involved in this process. Here we examined the role of Xpc on the transcriptional activation of the target genes by Oct3/4 using the inducible knockout strategy. We found that the removal of the C-terminal region of Xpc, including the interaction sites with Rad23 and Cetn2, showed faint impact on the gene expression profile of ES cells and the functional Xpc-ΔC ES cell lines retained proper gene expression profile as well as pluripotency to contribute chimeric embryos. These data indicated that the C-terminal region of Xpc is dispensable for the transcriptional activity of Oct3/4 in mouse ES cells.
Project description:To characterize the transdifferentiation of embryonic stem (ES) cells into trophoblast stem (TS) cells triggered by forced repression of Oct3/4, we performed whole-genome expression analysis after tetracycline (Tet)-induced knockout of Oct3/4. A Tet-inducible Oct3/4 knockout ES cell line ZHBTc4 was treated with Tet for 4 days in the presence of FGF4 and mouse embryonic fibroblasts (MEFs). A total of 15 samples from Day 0 to Day 4 were analyzed in three biological replicates.
Project description:Differentiated cells can be reprogrammed to an embryonic-like state by transfer of their nuclear contents into oocytes or by fusion with embryonic stem (ES) cells. Little is known about the factors that induce this reprogramming. Here we show that the combination of four factors, Oct3/4, Sox2, c-Myc, and Klf4, can generate pluripotent-like stem cells directly from mouse embryonic or adult fibroblast cultures. Unexpectedly, Nanog was dispensable in this process. These cells, which we designated iPS (induced pluriopotent-like stem) cells, exhibit the morphology and growth properties of ES cells and express ES cell marker genes. Subcutaneous transplantation of iPS cells into nude mice resulted in tumors containing a variety of tissues from all three germ layers. Following injection into blastocysts, iPS cells contributed to mouse embryonic development. These data demonstrate that pluripotent-like cells can be directly generated from fibroblast cultures by the addition of only a few defined factors. Experiment Overall Design: Total RNAs were isolated from indicated cells and labeled with Cy3. Hybridization was performed once for each sample.
Project description:To understand the mechanism underlying the versatility in transcriptional regulation by Sox2, we compared genome-wide binding sites of Sox2 in embryonic stem (ES) cells and trophoblast stem (TS) cells by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq). A tetracycline-inducible Oct3/4 knockout ES cell line ZHBTc4 was treated with Tet for 4 days in the presence of FGF4 and mouse embryonic fibroblasts (MEFs).
Project description:Differentiated cells can be reprogrammed to an embryonic-like state by transfer of their nuclear contents into oocytes or by fusion with embryonic stem (ES) cells. Little is known about the factors that induce this reprogramming. Here we show that the combination of four factors, Oct3/4, Sox2, c-Myc, and Klf4, can generate pluripotent-like stem cells directly from mouse embryonic or adult fibroblast cultures. Unexpectedly, Nanog was dispensable in this process. These cells, which we designated iPS (induced pluriopotent-like stem) cells, exhibit the morphology and growth properties of ES cells and express ES cell marker genes. Subcutaneous transplantation of iPS cells into nude mice resulted in tumors containing a variety of tissues from all three germ layers. Following injection into blastocysts, iPS cells contributed to mouse embryonic development. These data demonstrate that pluripotent-like cells can be directly generated from fibroblast cultures by the addition of only a few defined factors. Keywords: cell type comparison
Project description:We recently identified the DNA repair complex XPC-RAD23B-CETN2 as a stem cell coactivator (SCC) required for OCT4 and SOX2 transcriptional activation. Here we investigate genome-wide the role of SCC in murine ESCs by mapping regions bound by its RAD23B subunit in wild type and Xpc-/- ESCs, and analyzing transcriptional profiles of SCC-depleted ESCs. RAD23B ChIP-seq in wild type murine ESCs was performed in two replicates; normal IgG were used as controls. RAD23B ChIP-seq in Xpc-/- mESCs was compared to normal IgG. Antibodies to immunoprecipitate RAD23B were raised in guinea pigs against the 108-177 peptide of murine RAD23B protein (NP_033037). Anti-sera were affinity-purified. For RNA-seq, we compared wild type JM8.N4 cells with either Rad23b-/- cells, Rad23b-/-Xpc KD1 or Rad23b-/-Xpc KD2 cells. Xpc was knocked down with two independent shRNAs (KD1 and KD2).
Project description:Chickarmane2006 - Stem cell switch reversible
Kinetic modeling approach of the transcriptional dynamics of the embryonic stem cell switch.
This model is described in the article:
Transcriptional dynamics of the embryonic stem cell switch.
Chickarmane V, Troein C, Nuber UA, Sauro HM, Peterson C
PLoS Computational Biology. 2006; 2(9):e123
Abstract:
Recent ChIP experiments of human and mouse embryonic stem cells have elucidated the architecture of the transcriptional regulatory circuitry responsible for cell determination, which involves the transcription factors OCT4, SOX2, and NANOG. In addition to regulating each other through feedback loops, these genes also regulate downstream target genes involved in the maintenance and differentiation of embryonic stem cells. A search for the OCT4-SOX2-NANOG network motif in other species reveals that it is unique to mammals. With a kinetic modeling approach, we ascribe function to the observed OCT4-SOX2-NANOG network by making plausible assumptions about the interactions between the transcription factors at the gene promoter binding sites and RNA polymerase (RNAP), at each of the three genes as well as at the target genes. We identify a bistable switch in the network, which arises due to several positive feedback loops, and is switched on/off by input environmental signals. The switch stabilizes the expression levels of the three genes, and through their regulatory roles on the downstream target genes, leads to a binary decision: when OCT4, SOX2, and NANOG are expressed and the switch is on, the self-renewal genes are on and the differentiation genes are off. The opposite holds when the switch is off. The model is extremely robust to parameter changes. In addition to providing a self-consistent picture of the transcriptional circuit, the model generates several predictions. Increasing the binding strength of NANOG to OCT4 and SOX2, or increasing its basal transcriptional rate, leads to an irreversible bistable switch: the switch remains on even when the activating signal is removed. Hence, the stem cell can be manipulated to be self-renewing without the requirement of input signals. We also suggest tests that could discriminate between a variety of feedforward regulation architectures of the target genes by OCT4, SOX2, and NANOG.
This model is hosted on BioModels Database
and identified by: MODEL7957907314
.
To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models
.
To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication
for more information.
Project description:Chickarmane2006 - Stem cell switch irreversible
Kinetic modeling approach of the transcriptional dynamics of the embryonic stem cell switch.
This model is described in the article:
Transcriptional dynamics of the embryonic stem cell switch.
Chickarmane V, Troein C, Nuber UA, Sauro HM, Peterson C
PLoS Computational Biology. 2006; 2(9):e123
Abstract:
Recent ChIP experiments of human and mouse embryonic stem cells have elucidated the architecture of the transcriptional regulatory circuitry responsible for cell determination, which involves the transcription factors OCT4, SOX2, and NANOG. In addition to regulating each other through feedback loops, these genes also regulate downstream target genes involved in the maintenance and differentiation of embryonic stem cells. A search for the OCT4-SOX2-NANOG network motif in other species reveals that it is unique to mammals. With a kinetic modeling approach, we ascribe function to the observed OCT4-SOX2-NANOG network by making plausible assumptions about the interactions between the transcription factors at the gene promoter binding sites and RNA polymerase (RNAP), at each of the three genes as well as at the target genes. We identify a bistable switch in the network, which arises due to several positive feedback loops, and is switched on/off by input environmental signals. The switch stabilizes the expression levels of the three genes, and through their regulatory roles on the downstream target genes, leads to a binary decision: when OCT4, SOX2, and NANOG are expressed and the switch is on, the self-renewal genes are on and the differentiation genes are off. The opposite holds when the switch is off. The model is extremely robust to parameter changes. In addition to providing a self-consistent picture of the transcriptional circuit, the model generates several predictions. Increasing the binding strength of NANOG to OCT4 and SOX2, or increasing its basal transcriptional rate, leads to an irreversible bistable switch: the switch remains on even when the activating signal is removed. Hence, the stem cell can be manipulated to be self-renewing without the requirement of input signals. We also suggest tests that could discriminate between a variety of feedforward regulation architectures of the target genes by OCT4, SOX2, and NANOG.
This model is hosted on BioModels Database
and identified by: MODEL7957942740
.
To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models
.
To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication
for more information.