Transcriptomics

Dataset Information

64

Acute depletion of Tet1-dependent 5-hydroxymethylcytosine levels impairs LIF/Stat3 signaling and results in loss of embryonic stem cell identity [expression profiling]


ABSTRACT: The TET family of FE(II) and 2-oxoglutarate-dependent enzymes (Tet1/2/3) promote DNA demethylation by converting 5-methylcytosine to 5-hydroxymethylcytosine (5hmC), which they further oxidize into 5-formylcytosine and 5-carboxylcytosine. Tet1 is robustly expressed in mouse embryonic stem cells (mESCs) and has been implicated in mESC maintenance. Here we demonstrate that, unlike genetic deletion, RNAi-mediated depletion of Tet1 in mESCs led to a significant reduction in 5hmC and loss of mESC identity. The differentiation phenotype due to Tet1 depletion positively correlated with the extent of 5hmC loss. Meta-analyses of genomic datasets suggested interaction between Tet1 and leukemia inhibitory factor (LIF) signaling. LIF signaling is known to promote self-renewal and pluripo-tency in mESCs partly by opposing MAPK/ERK mediated differentiation. Withdrawal of LIF leads to differentiation of mESCs. We discovered that Tet1 depletion impaired LIF-dependent Stat3-mediated gene activation by affecting Stat3's ability to bind to its target sites on chromatin. Nanog overexpression or inhibition of MAPK/ERK signaling, both known to maintain mESCs in the absence of LIF, rescued Tet1 depletion, further supporting the dependence of LIF/Stat3 signaling on Tet1. These data support the conclusion that analysis of mESCs in the hours/days immediately following efficient Tet1 depletion reveals Tet1’s normal physiological role in maintaining the pluripotent state that may be subject to homeostatic compensation in genetic models. Genome-wide mapping of 5hmC and microarray gene expression profiling in E14Tg2a mESCs after transfection with indicated siRNAs: Tet1 siRNA #1 (Invitrogen, MSS284895), Tet1 siRNA #2 (Invitrogen, MSS284897), and Control siRNA duplex targeting firefly luciferase.

ORGANISM(S): Mus musculus  

SUBMITTER: Guang Hu   Raja Jothi  Johannes Freudenberg 

PROVIDER: E-GEOD-34886 | ArrayExpress | 2012-01-06

SECONDARY ACCESSION(S): GSE34886PRJNA156265

REPOSITORIES: GEO, ArrayExpress

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Publications

Acute depletion of Tet1-dependent 5-hydroxymethylcytosine levels impairs LIF/Stat3 signaling and results in loss of embryonic stem cell identity.

Freudenberg Johannes M JM   Ghosh Swati S   Lackford Brad L BL   Yellaboina Sailu S   Zheng Xiaofeng X   Li Ruifang R   Cuddapah Suresh S   Wade Paul A PA   Hu Guang G   Jothi Raja R  

Nucleic acids research 20111230 8


The TET family of FE(II) and 2-oxoglutarate-dependent enzymes (Tet1/2/3) promote DNA demethylation by converting 5-methylcytosine to 5-hydroxymethylcytosine (5hmC), which they further oxidize into 5-formylcytosine and 5-carboxylcytosine. Tet1 is robustly expressed in mouse embryonic stem cells (mESCs) and has been implicated in mESC maintenance. Here we demonstrate that, unlike genetic deletion, RNAi-mediated depletion of Tet1 in mESCs led to a significant reduction in 5hmC and loss of mESC iden  ...[more]

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