Resetting Transcription Factor Control Circuitry toward Ground-State Pluripotency in Human

Abstract

SummaryCurrent human pluripotent stem cells lack the transcription factor circuitry that governs the ground state of mouse embryonic stem cells (ESC). Here, we report that short-term expression of two components, NANOG and KLF2, is sufficient to ignite other elements of the network and reset the human pluripotent state. Inhibition of ERK and protein kinase C sustains a transgene-independent rewired state. Reset cells self-renew continuously without ERK signaling, are phenotypically stable, and are karyotypically intact. They differentiate in vitro and form teratomas in vivo. Metabolism is reprogrammed with activation of mitochondrial respiration as in ESC. DNA methylation is dramatically reduced and transcriptome state is globally realigned across multiple cell lines. Depletion of ground-state transcription factors, TFCP2L1 or KLF4, has marginal impact on conventional human pluripotent stem cells but collapses the reset state. These findings demonstrate feasibility of installing and propagating functional control circuitry for ground-state pluripotency in human cells.

A Glycolytic Solution for Pluripotent Stem Cells

“…Previous studies demonstrated that human and mouse naïve PSCs contain mitochondria with more immature morphology than primed PSCs (Zhou et al, 2012). At the same time, human and mouse naïve PSCs exhibit increased OXPHOS (Takashima et al, 2014;Zhou et al, 2012). Now Gu et al (2016) demonstrate that human naïve PSCs, both newly derived and those cultured in previously reported reversion conditions (Takashima et al, 2014;Theunissen et al, 2014), in addition to higher mitochondrial oxygen consumption also display elevated glycolytic rates.…”

“…At the same time, human and mouse naïve PSCs exhibit increased OXPHOS (Takashima et al, 2014;Zhou et al, 2012). Now Gu et al (2016) demonstrate that human naïve PSCs, both newly derived and those cultured in previously reported reversion conditions (Takashima et al, 2014;Theunissen et al, 2014), in addition to higher mitochondrial oxygen consumption also display elevated glycolytic rates. Human naïve PSCs consumed more glucose, produced more lactate, and incorporated more glucose carbons into lactate and into purine and pyrimidine nucleotides, implying increased flux towards glycolysis and the PPP.…”

“…Gu et al (2016) investigated the metabolic changes occurring during the transition between human naïve and primed pluripotent states. These two PSC states respectively represent pre-implantation cells in the inner cell mass and post-implantation cells in the epiblast (Takashima et al, 2014;Theunissen et al, 2014). Previous studies demonstrated that human and mouse naïve PSCs contain mitochondria with more immature morphology than primed PSCs (Zhou et al, 2012).…”

“…In fact, it has been suggested that glycolysis may represent the basal energy source for naïve PSCs and mitochondrial respiration their spare capacity (Carbognin et al, 2016). Accordingly, inhibition of glycolysis is not as detrimental for naïve PSCs as it is for primed PSCs (Takashima et al, 2014). It would be interesting to compare the effects of OXPHOS inhibition.…”

“…LIF is employed in the majority of naïve media (Gu et al, 2016;Takashima et al, 2014) and activates mitochondrial respiration (Carbognin et al, 2016). Activin is included in some naïve media (Gu et al, 2016;Theunissen et al, 2014) and may stimulate glycolysis through the stabilization of Hypoxia-Inducible Factor-1 alpha (HIF1a) (Zhou et al, 2012).…”