Embryonic stem (ES) cell pluripotency is sustained by a network of transcription
factors centred on Oct4, Sox2 and Nanog. Whilst Oct4 and Sox2 expression is
relatively uniform, ES cells fluctuate between states of high Nanog expression
possessing high self-renewal efficiency, and low Nanog expression exhibiting
increased differentiation propensity. Moreover, modulation in the level of Nanog
expression determines the efficiency of ES cell self-renewal.
To identify genes regulated by Nanog, genome-wide transcriptional profiling was
performed on ES cells expressing different Nanog levels and Nanog-null ES cells
expressing a Nanog-ERT2 fusion protein in which nuclear Nanog activity can be
regulated by tamoxifen. Surprisingly, only a minor fraction of the genes to which
Nanog binds showed significant changes in response to Nanog induction. Prominent
amongst Nanog-responsive genes is Estrogen-related receptor b (Esrrb). Nanog binds
directly to Esrrb, enhances binding and pause-release of RNAPolII from the Esrrb
promoter and stimulates Esrrb transcription. Consistent with these findings, elevation
of Nanog produces a cell population that expresses uniformly high Esrrb levels.
Moreover, double fluorescent reporter lines show that Esrrb and Nanog levels are
strongly correlated in individual cells. Loss of Nanog is required for downregulation
of Esrrb, which coincides with commitment to differentiate.
Esrrb overexpression results in LIF independent self-renewal, and blocks neural
differentiation, even in the absence of Nanog. Cell fusion experiments between ES
and neural stem (NS) cells show that elevated Esrrb levels allow the reprogramming
of the NS cell genome in the absence of Nanog. Esrrb can rescue stalled
reprogramming during the derivation of Nanog-/- induced pluripotent stem (iPS)
cells. Moreover, targeted knock-in of Esrrb at the Nanog locus rescues the ability of
Nanog null ES cells to maintain germ cell development beyond E12. Finally, Esrrb
deletion abolishes the defining ability of Nanog to confer LIF-independent selfrenewal
to ES cells.
Together these data identify Esrrb as a critical downstream mediator of Nanog
function