Stat3 promotes mitochondrial transcription and oxidative respiration during maintenance and induction of naive pluripotency.

Transcription factor Stat3 directs self-renewal of pluripotent mouse embryonic stem (ES) cells downstream of the cytokine leukemia inhibitory factor (LIF). Stat3 upregulates pivotal transcription factors in the ES cell gene regulatory network to sustain naïve identity. Stat3 also contributes to the rapid proliferation of ES cells. Here, we show that Stat3 increases the expression of mitochondrial-encoded transcripts and enhances oxidative metabolism. Chromatin immunoprecipitation reveals that Stat3 binds to the mitochondrial genome, consistent with direct transcriptional regulation. An engineered form of Stat3 that localizes predominantly to mitochondria is sufficient to support enhanced proliferation of ES cells, but not to maintain their undifferentiated phenotype. Furthermore, during reprogramming from primed to naïve states of pluripotency, Stat3 similarly upregulates mitochondrial transcripts and facilitates metabolic resetting. These findings suggest that the potent stimulation of naïve pluripotency by LIF/Stat3 is attributable to parallel and synergistic induction of both mitochondrial respiration and nuclear transcription factors.GM’s laboratory is supported by grants from Armenise-Harvard Foundation and Telethon Foundation (TCP13013). The Cambridge Stem Cell Institute receives core funding from the Wellcome Trust and Medical Research Council. GM was supported by a Human Frontier Science Program Fellowship. AS is a Medical Research Professor.This is the final version of the article. It first appeared from Wiley via http://dx.doi.org/10.15252/embj.20159262

Experimental Determination of Momentum-Resolved Electron-Phonon Coupling

We provide a novel experimental method to quantitatively estimate the electron-phonon coupling and its momentum dependence from resonant inelastic x-ray scattering (RIXS) spectra based on the detuning of the incident photon energy away from an absorption resonance. We apply it to the cuprate parent compound NdBa$_2$Cu$_3$O$_6$ and find that the electronic coupling to the oxygen half-breathing phonon mode is strongest at the Brillouin zone boundary, where it amounts to $\sim 0.17$ eV, in agreement with previous studies. In principle, this method is applicable to any absorption resonance suitable for RIXS measurements and will help to define the contribution of lattice vibrations to the peculiar properties of quantum materials.Comment: 6 pages, 3 figure

Determining the Electron-Phonon Coupling in Superconducting Cuprates by Resonant Inelastic X-ray Scattering: Methods and Results on Nd1+x_{1+x}Ba2−x_{2-x}Cu3_3O7−δ_{7-\delta}

The coupling between lattice vibration quanta and valence electrons can induce charge density modulations and decisively influence the transport properties of materials, e.g. leading to conventional superconductivity. In high critical temperature superconductors, where electronic correlation is the main actor, the actual role of electron-phonon coupling (EPC) is being intensely debated theoretically and investigated experimentally. We present an in-depth study of how the EPC strength can be obtained directly from resonant inelastic x-ray scattering (RIXS) data through the theoretical approach derived by Ament et al. [EPL 95, 27008 (2011)]. The role of the model parameters (e.g. phonon energy $\omega_0$, intermediate state lifetime $1/\Gamma$, EPC matrix element $M$, and detuning energy $\Omega$) is thoroughly analyzed, providing general relations among them that can be used to make quantitative estimates of the dimensionless EPC $g = (M/\omega_0)^2$ without detailed microscopic modeling. We then apply these methods to very high resolution Cu $L_3$ edge RIXS spectra of three Nd$_{1+x}$Ba$_{2-x}$Cu$_3$O$_{7-\delta}$ films. For the insulating antiferromagnetic parent compound the value of $M$ as a function of the in-plane momentum transfer is obtained for Cu-O bond-stretching (breathing) and bond-bending (buckling) phonon branches. For the underdoped and the nearly optimally doped samples, the effects of Coulomb screening and of charge-density-wave correlations on $M$ are assessed. In light of the anticipated further improvements of the RIXS experimental resolution, this work provides a solid framework for an exhaustive investigation of the EPC in cuprates and other quantum materials.Comment: 21 pages, 16 figure

Metabolic control of DNA methylation in naive pluripotent cells.

Naive epiblast and embryonic stem cells (ESCs) give rise to all cells of adults. Such developmental plasticity is associated with genome hypomethylation. Here, we show that LIF-Stat3 signaling induces genomic hypomethylation via metabolic reconfiguration. Stat3-/- ESCs show decreased α-ketoglutarate production from glutamine, leading to increased Dnmt3a and Dnmt3b expression and DNA methylation. Notably, genome methylation is dynamically controlled through modulation of α-ketoglutarate availability or Stat3 activation in mitochondria. Alpha-ketoglutarate links metabolism to the epigenome by reducing the expression of Otx2 and its targets Dnmt3a and Dnmt3b. Genetic inactivation of Otx2 or Dnmt3a and Dnmt3b results in genomic hypomethylation even in the absence of active LIF-Stat3. Stat3-/- ESCs show increased methylation at imprinting control regions and altered expression of cognate transcripts. Single-cell analyses of Stat3-/- embryos confirmed the dysregulated expression of Otx2, Dnmt3a and Dnmt3b as well as imprinted genes. Several cancers display Stat3 overactivation and abnormal DNA methylation; therefore, the molecular module that we describe might be exploited under pathological conditions

Determining the electron-phonon coupling in superconducting cuprates by resonant inelastic x-ray scattering: Methods and results on Nd1+xBa2-xCu3O7-δ

The coupling between lattice vibration quanta and valence electrons can induce charge-density modulations and decisively influence the transport properties of materials, e.g., leading to conventional superconductivity. In high-critical-temperature superconductors, where electronic correlation is the main actor, the actual role of electron-phonon coupling (EPC) is being intensely debated theoretically and investigated experimentally. We present an in-depth study of how the EPC strength can be obtained directly from resonant inelastic x-ray scattering (RIXS) data through the theoretical approach derived by Ament et\ua0al. [Europhys. Lett. 95, 27008 (2011)]. The role of the model parameters (e.g., phonon energy ω0, intermediate state lifetime 1/Γ, EPC matrix element M, and detuning energy Ω) is thoroughly analyzed, providing general relations among them that can be used to make quantitative estimates of the dimensionless EPC g=(M/ω0)2 without detailed microscopic modeling. We then apply these methods to very high-resolution Cu L3-edge RIXS spectra of three Nd1+xBa2−xCu3O7−δ films. For the insulating antiferromagnetic parent compound, the value of M as a function of the in-plane momentum transfer is obtained for Cu-O bond-stretching (breathing) and bond-bending (buckling) phonon branches. For the underdoped and the nearly optimally doped samples, the effects of Coulomb screening and of charge-density-wave correlations on M are assessed. In light of the anticipated further improvements of the RIXS experimental resolution, this work provides a solid framework for an exhaustive investigation of the EPC in cuprates and other quantum materials

Metabolic control of DNA methylation in naive pluripotent cells.

Naive epiblast and embryonic stem cells (ESCs) give rise to all cells of adults. Such developmental plasticity is associated with genome hypomethylation. Here, we show that LIF-Stat3 signaling induces genomic hypomethylation via metabolic reconfiguration. Stat3-/- ESCs show decreased α-ketoglutarate production from glutamine, leading to increased Dnmt3a and Dnmt3b expression and DNA methylation. Notably, genome methylation is dynamically controlled through modulation of α-ketoglutarate availability or Stat3 activation in mitochondria. Alpha-ketoglutarate links metabolism to the epigenome by reducing the expression of Otx2 and its targets Dnmt3a and Dnmt3b. Genetic inactivation of Otx2 or Dnmt3a and Dnmt3b results in genomic hypomethylation even in the absence of active LIF-Stat3. Stat3-/- ESCs show increased methylation at imprinting control regions and altered expression of cognate transcripts. Single-cell analyses of Stat3-/- embryos confirmed the dysregulated expression of Otx2, Dnmt3a and Dnmt3b as well as imprinted genes. Several cancers display Stat3 overactivation and abnormal DNA methylation; therefore, the molecular module that we describe might be exploited under pathological conditions

Y705 and S727 are required for mitochondrial import and transcriptional activities of STAT3 and regulate proliferation of embryonic and tissue stem cells

The STAT3 transcription factor, acting both in the nucleus and mitochondria, maintains embryonic stem cell pluripotency and promotes their proliferation. In this work, using zebrafish, we determined in vivo that mitochondrial STAT3 regulates mtDNA transcription in embryonic and larval stem cell niches and that this activity affects their proliferation rates. As a result, we demonstrated that STAT3 import inside mitochondria requires Y705 phosphorylation by Jak, while its mitochondrial transcriptional activity, as well as its effect on proliferation, depends on the MAPK target S727. These data were confirmed using mouse embryonic stem cells: while the Y705 mutated STAT3 cannot enter mitochondria, the S727 mutation does not affect the import in the organelle and is responsible for STAT3-dependent mitochondrial transcription. Surprisingly, STAT3-dependent increase of mitochondrial transcription seems independent from STAT3 binding to STAT3 responsive elements. Finally, loss of function experiments, with chemical inhibition of the JAK/STAT3 pathway or genetic ablation of stat3 gene, demonstrated that STAT3 is also required for cell proliferation in the intestine of zebrafish

3D ECM-rich environment sustains the identity of naive human iPSCs

The establishment of in vitro naive human pluripotent stem cell cultures opened new perspectives for the study of early events in human development. The role of several transcription factors and signaling pathways have been characterized during maintenance of human naive pluripotency. However, little is known about the role exerted by the extracellular matrix (ECM) and its three-dimensional (3D) organization. Here, using an unbiased and integrated approach combining microfluidic cultures with transcriptional, proteomic, and secretome analyses, we found that naive, but not primed, hiPSC colonies are characterized by a self-organized ECM-rich microenvironment. Based on this, we developed a 3D culture system that supports robust long-term feeder-free self-renewal of naive hiPSCs and also allows direct and timely developmental morphogenesis simply by modulating the signaling environment. Our study opens new perspectives for future applications of naive hiPSCs to study critical stages of human development in 3D starting from a single cell