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 data sets suggested interaction between Tet1 and 
leukemia inhibitory factor (LIF) signaling. LIF signaling is known to promote 
self-renewal and pluripotency 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.