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Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (RNA-Seq)

ABSTRACT: Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4 and c-Myc (abbreviated as OSKM), in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog, Oct4 and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the vast majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation were attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods.

ORGANISM(S): Mus musculus

PROVIDER: GSE67265 | GEO | 2015/04/30

SECONDARY ACCESSION(S): PRJNA279457

REPOSITORIES: GEO

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Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (RNA-Seq)

Project description:Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4 and c-Myc (abbreviated as OSKM), in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog, Oct4 and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the vast majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation were attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods. poly RNA-Seq was measured before, during and after conversion of mouse embryonic fibroblasts to neural stem cells using OSKM trans-differentiation method.

2015-04-30 | E-GEOD-67265 | biostudies-arrayexpress

Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (ATAC-Seq)

Project description:Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4 and c-Myc (abbreviated as OSKM), in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog, Oct4 and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the vast majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation were attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods. poly RNA-Seq and Chromatin accesibility (ATAC-seq) were measured during conversion of mouse embryonic fibroblasts to neural stem cells using OSKM trans-differentiation method, as well as in mouse emrbyonic fibroblasts, iPSCs and mouse ESCs.

2015-04-30 | E-GEOD-67298 | biostudies-arrayexpress

Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (WGBS)

Project description:Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4 and c-Myc (abbreviated as OSKM), in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog, Oct4 and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the vast majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation were attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods. WGBS (Whole-Genome-Bisulfite-sequencing) were measured during conversion of mouse embryonic fibroblasts to neural stem cells using OSKM trans-differentiation method, as well as in mouse emrbyonic fibroblasts, and mouse ESCs.

2015-05-07 | E-GEOD-68601 | biostudies-arrayexpress

Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (WGBS)

2015-05-07 | GSE68601 | GEO

Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (ATAC-Seq)

2015-04-30 | GSE67298 | GEO

Oct4 binding and Histone modification profiling during OSKM-mediated 2nd reprogramming

Project description:Forced expression of four transcription factors Oct4,Sox2, Klf4 and Myc (OSKM) induces somatic cell reprogramming towards pluripotency. Major efforts have been made to characterize the molecular events involved in this process. Yet, it remains elusive how gene expression change, epigenetic landscape remodelling and cell fate conversion are triggered by expression of these Yamanaka factors.To address this gap,we utilized a secondary inducible reprogramming system and performed genome-wide profilings of Oct4 binding, histone modification(H3K4me3/H3K27me3/H3K4me1/H3K27ac), and gene expression analysis during this process. Through integrative analysis, we revealed stage-specific Oct4 binding and enhancer signatures in consistence with gene expression changes,in which the initial regression of somatic program is followed by the gradual acquisition of pluripotent program. Oct4 preferatially binds to H3K4me1 marked enhancer regions and Oct4 binding is positively correlated with active mark H3K27ac. Moreover,we observed significant enhancer activation of epigenetic related genes, especially acetylation associated genes, prior to pluripotency network activation, suggesting a pivotal role of epigenetic remodelling in the process of pluripotency acquisition and maintenance. We used ChIP-seq to explore the global changes of Oct4 binding profiling during OSKM-mediated somatic cell reprogramming. The exogenous Oct4 was tagged with 3XFlag, thus we used anti-flag antibody to monitor exogenous Oct4 changes; anti-Oct4 antibody which recognizes both endogenous and exogenous Oct4 was used to monitor total Oct4 changes during iPSC induction. In addition, we also examined genome-wide H3K4me1/H3K27ac/H3K4me3/H3K27me3/RNAPII profiling during 3Flag-OSKM 2' reprogramming.

2015-12-31 | E-GEOD-67520 | biostudies-arrayexpress

Expression data from OSKM-mediated 2nd reprogramming cells and the corresponding iPS cell line

Project description:Forced expression of four transcription factors Oct4,Sox2, Klf4 and Myc (OSKM) induces somatic cell reprogramming towards pluripotency. Major efforts have been made to characterize the molecular events involved in this process. Yet, it remains elusive how gene expression change, epigenetic landscape remodelling and cell fate conversion are triggered by expression of these Yamanaka factors. To address this gap, we utilized a secondary inducible reprogramming system and performed genome-wide profilings of Oct4 binding, histone modification (H3K4me3/H3K27me3/H3K4me1/H3K27ac), and gene expression analysis during this process. Through integrative analysis, we revealed stage-specific Oct4 binding and enhancer signatures in consistence with gene expression changes, in which the initial regression of somatic program is followed by the gradual acquisition of pluripotent program. Oct4 preferatially binds to H3K4me1 marked enhancer regions and Oct4 binding is positively correlated with active mark H3K27ac. Moreover, we observed significant enhancer activation of epigenetic related genes, especially acetylation associated genes, prior to pluripotency network activation, suggesting a pivotal role of epigenetic remodelling in the process of pluripotency acquisition and maintenance. We used microarrays to explore the global changes of gene expression during OSKM-mediated somatic cell reprogramming.

2015-12-31 | GSE67462 | GEO

Expression data from OSKM-mediated 2nd reprogramming cells and the corresponding iPS cell line

Project description:Forced expression of four transcription factors Oct4,Sox2, Klf4 and Myc (OSKM) induces somatic cell reprogramming towards pluripotency. Major efforts have been made to characterize the molecular events involved in this process. Yet, it remains elusive how gene expression change, epigenetic landscape remodelling and cell fate conversion are triggered by expression of these Yamanaka factors.To address this gap,we utilized a secondary inducible reprogramming system and performed genome-wide profilings of Oct4 binding, histone modification(H3K4me3/H3K27me3/H3K4me1/H3K27ac), and gene expression analysis during this process. Through integrative analysis, we revealed stage-specific Oct4 binding and enhancer signatures in consistence with gene expression changes,in which the initial regression of somatic program is followed by the gradual acquisition of pluripotent program. Oct4 preferatially binds to H3K4me1 marked enhancer regions and Oct4 binding is positively correlated with active mark H3K27ac. Moreover,we observed significant enhancer activation of epigenetic related genes, especially acetylation associated genes, prior to pluripotency network activation, suggesting a pivotal role of epigenetic remodelling in the process of pluripotency acquisition and maintenance. We used microarrays to explore the global changes of gene expression during OSKM-mediated somatic cell reprogramming. time series design with bulk population samples collected at different time point of 2nd reprogramming as well as a corresponding iPS cell line. Each sample include two replicates. 2nd mouse embryonic fibroblasts(sample day0) were treated with ES medium supplemented with 1Âµg/ml Doxcycline and 50Âµg/ml Vitamin C for 15 days (sample day1-day15); 72 hours after dox and Vc withdrawl, dox-independent iPS cells are collected(sample day18).

2015-12-31 | E-GEOD-67462 | biostudies-arrayexpress

Oct4 binding and Histone modification profiling during OSKM-mediated 2nd reprogramming

Project description:Forced expression of four transcription factors Oct4,Sox2, Klf4 and Myc (OSKM) induces somatic cell reprogramming towards pluripotency. Major efforts have been made to characterize the molecular events involved in this process. Yet, it remains elusive how gene expression change, epigenetic landscape remodelling and cell fate conversion are triggered by expression of these Yamanaka factors. To address this gap,we utilized a secondary inducible reprogramming system and performed genome-wide profilings of Oct4 binding, histone modification (H3K4me3/H3K27me3/H3K4me1/H3K27ac), and gene expression analysis during this process. Through integrative analysis, we revealed stage-specific Oct4 binding and enhancer signatures in consistence with gene expression changes, in which the initial regression of somatic program is followed by the gradual acquisition of pluripotent program. Oct4 preferatially binds to H3K4me1 marked enhancer regions and Oct4 binding is positively correlated with active mark H3K27ac. Moreover, we observed significant enhancer activation of epigenetic related genes, especially acetylation associated genes, prior to pluripotency network activation, suggesting a pivotal role of epigenetic remodelling in the process of pluripotency acquisition and maintenance.

2015-12-31 | GSE67520 | GEO

Cooperative binding of Oct4, Sox2, and Klf4 with stage-specific transcription factors orchestrates reprogramming [RNA-seq]

Project description:Oct4, Sox2, Klf4, and cMyc (OSKM) reprogram somatic cells to pluripotency. To gain a mechanistic understanding of their function, we mapped OSKM-binding, stage-specific transcription-factors (TFs), and chromatin-states in discrete reprogramming stages and performed loss- and gain-of-function experiments. We found that early in reprogramming OSK extensively bind somatic-enhancers and initiate their decommissioning by recruiting Hdac1. Concurrently, OSK engage other sites, including specific pluripotency-enhancers, and induce the relocation of somatic TFs to these sites and away from somatic-enhancers, extending somatic-enhancer decommissioning genome-wide. Pluripotency-enhancer selection early in reprogramming occurs predominantly at sites with high OSK-motif densities and requires collaborative binding by OSK. Most pluripotency-enhancers are selected later and occupied by OS and stage-specific-TFs like Esrrb. Overexpression of stage-specific-TFs influences reprogramming efficiency by changing OSK-occupancy, somatic-enhancer decommissioning, and pluripotency-enhancer selection. We propose that collaborative interactions among OSK and with stage-specific-TFs direct both somatic-enhancer decommissioning and pluripotency-enhancer selection, which drives the enhancer reorganization underlying reprogramming

2017-01-19 | GSE90894 | GEO

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