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Cited by 671Neurogenesis and neuroplasticity mechanismsCancer, Hypoxia, and MetabolismMetabolism, Diabetes, and Cancer

Metabolic reprogramming during neuronal differentiation from aerobic glycolysis to neuronal oxidative phosphorylation

Open access
Zheng, Xinde, Boyer, Leah, Jin, Mingji, Mertens, Jerome, Kim, Yongsung, Ma, Li, Ma, Li, Hamm, Michael +2 more·2016·eLife
DOI 10.7554/elife.13374PMC PMC4963198
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Abstract

How metabolism is reprogrammed during neuronal differentiation is unknown. We found that the loss of hexokinase (HK2) and lactate dehydrogenase (LDHA) expression, together with a switch in pyruvate kinase gene splicing from PKM2 to PKM1, marks the transition from aerobic glycolysis in neural progenitor cells (NPC) to neuronal oxidative phosphorylation. The protein levels of c-MYC and N-MYC, transcriptional activators of the HK2 and LDHA genes, decrease dramatically. Constitutive expression of HK2 and LDHA during differentiation leads to neuronal cell death, indicating that the shut-off aerobic glycolysis is essential for neuronal survival. The metabolic regulators PGC-1α and ERRγ increase significantly upon neuronal differentiation to sustain the transcription of metabolic and mitochondrial genes, whose levels are unchanged compared to NPCs, revealing distinct transcriptional regulation of metabolic genes in the proliferation and post-mitotic differentiation states. Mitochondrial mass increases proportionally with neuronal mass growth, indicating an unknown mechanism linking mitochondrial biogenesis to cell size.

AI snapshot

Written from the paper’s full text.

TL;DR

Neuronal differentiation from human NPCs is marked by shutting off aerobic glycolysis and shifting to oxidative phosphorylation, with loss of HK2/LDHA, PKM2-to-PKM1 splicing, and decreased c-MYC/N-MYC. Constitutive HK2/LDHA expression kills differentiating neurons, while increased PGC-1α/ERRγ maintain rather than increase metabolic and mitochondrial gene expression.

Key findings

  • Differentiation from NPCs to neurons involves loss of HK2 and LDHA protein, a PKM splice switch from PKM2 to PKM1, and a neuron-specific OGDH splice form lacking a Ca²⁺-binding motif; c-MYC and N-MYC protein levels fall by ~70% and ~85%, respectively.
  • Neurons show lower glycolytic/TCA metabolite levels and higher oxidative capacity: basal and maximum OCR rise from ~82 and 130 pmoles/min/10 µg in NPCs to ~98 and 183 in 3-week neurons, while lactate/glucose ratio falls from ~1.61 to ~0.35.
  • Constitutive HK2 and LDHA expression during differentiation causes neuronal death: ~40% condensed nuclei versus ~6% in controls, with ~40% GFAP-positive glial cells versus ~4% in controls; extra pyruvate (5 mM) rescues neuronal death but not glial increase.
  • PGC-1α and ERRγ increase ~3–4 fold and ~2.8–3.4 fold, respectively, and are required to maintain expression of glycolysis, TCA, and mitochondrial respiratory complex genes, even though most mitochondrial gene expression is unchanged versus NPCs; mtDNA copy number and protein mass both increase ~4-fold by 3 weeks.

Methods

  • Human BJ iPSC-derived NPCs were differentiated for 1–3 weeks and validated by Sox2/Nestin and MAP2 staining; RNA-seq was performed on two NPC lines with duplicates, and data were mapped with STAR and quantified by FPKM.
  • Metabolic profiling used GC-MS for glycolytic/TCA metabolites, Seahorse for OCR, lactate assays, immunoblotting/immunostaining, RT-PCR, ChIP for c-MYC/N-MYC binding, and lenti-shRNA knockdowns of HK2, LDHA, PGC-1α, or ERRγ.
  • Direct neuronal conversion from fibroblasts was tested with PTB knockdown or Ngn2/Ascl1 overexpression; mouse neuroprogenitor differentiation and published mouse astrocyte/neuron RNA-seq were used for conservation checks.

Limitations

  • The paper states that expression levels of metabolic enzymes do not completely reflect biochemical/physiological activities, and post-translational modifications were not explored.
  • The paper notes that mitochondrial structure, dynamics, and matrix calcium were not investigated, and that protein degradation may also downregulate key metabolic enzymes.
  • Evident from the text: the main differentiation model relies on human BJ iPSC-derived NPCs, with some key findings validated in mouse cells; 3-week cultures were used as developing neurons, and rare glial cells emerge later, so longer-term neuronal populations were not the primary focus.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.7554/elife.13374

Cites

77 of 77 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 50 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    PGC-1α and ERRγ increase significantly upon neuronal differentiation.

    “The metabolic regulators PGC-1α and ERRγ increase significantly upon neuronal differentiation”
    Quote in the paperSubstantial claim
  • Findingown data

    Mitochondrial mass increases proportionally with neuronal mass growth.

    “Mitochondrial mass increases proportionally with neuronal mass growth”
    Quote in the paperNot a substantial claim
  • Findingown data

    During neuronal differentiation, pyruvate kinase gene splicing switches from PKM2 to PKM1.

    “a switch in pyruvate kinase gene splicing from PKM2 to PKM1”
    • Also stated in: § Neuron-specific splicing of PKM and OGDH · plain
      “NPCs mainly expressed PKM2, whereas neurons expressed PKM1”
    Quote in the paperSubstantial claim
  • Findingown data

    In human NPC-derived neurons, HK2 and LDHA expression is lost during neuronal differentiation.

    “We found that the loss of hexokinase (HK2) and lactate dehydrogenase (LDHA) expression”
    • Also stated in: § HK2 and LDHA were off during neuronal differentiation · plain
      “HK2 and LDHA mRNAs were about 15% and 25% of those in NPCs, but strikingly, both proteins almost completely disappeared upon differentiation”
    Quote in the paperSubstantial claim
  • Findingreasoning

    Levels of metabolic and mitochondrial genes are unchanged in neurons compared to NPCs.

    “whose levels are unchanged compared to NPCs”
    Quote in the paperSubstantial claim
  • May combine two claims. 43
  • Classified as probably not a substantial claim. 17
  • The quote only partly states this claim. 1

  • May combine two claims.

    Findingcitation

    OGDH has a novel and conserved neuronal splicing shift, resulting in loss of a calcium binding motif.

    “OGDH, a key enzyme in the TCA cycle, has a novel and conserved neuronal splicing shift, resulting in the loss of a calcium binding motif.”
    • Also stated in: § Neuron-specific splicing of PKM and OGDH · plain
      “a stretch of 34 amino acids in OGDH1, containing a Ca2+-binding motif (Armstrong et al., 2014), is replaced by 45 amino acids specific to OGDHneu”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Armstrong et al., 2014

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    In neurons, 3-phosphoglycerate and pyruvate levels dropped to ~52% and 32% of those in NPCs.

    “the levels of 3-phosphoglyerate (3PG) and pyruvate in neurons dropped to ~52% and 32% of those in NPCs”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In neurons, TCA intermediates citrate, α-ketoglutarate, fumarate, and malate were lower than in NPCs (to ~32%, ~17%, ~30%, ~22%), while succinate was ~62%.

    “citrate was ~32% of that in NPCs, α-ketoglutarate was ~17%, fumarate was ~30%, and malate was ~22%; succinate was relatively higher, at ~62%”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    HK2 and LDHA mRNAs were about 15% and 25% of those in NPCs, but both proteins almost completely disappeared upon differentiation.

    “HK2 and LDHA mRNAs were about 15% and 25% of those in NPCs, but strikingly, both proteins almost completely disappeared upon differentiation”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    During mouse neuroprogenitor differentiation, both HK2 and LDHA mRNA levels dropped to ~8.5% and ~11% of NPC levels, and their protein levels were greatly decreased.

    “both HK2 and LDHA mRNA levels dropped, falling to ~8.5% and ~11% of the level in NPCs, respectively, and, consistently, their protein levels were also greatly decreased”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    OGDHneu splicing is conserved in mouse neurons, whereas mouse astrocytes only expressed OGDH1.

    “OGDHneu splicing is conserved in mouse neurons, whereas mouse astrocytes, like human astrocytes, only expressed OGDH1”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    OGDH in NPCs (OGDH1) uses exon 4, whereas the neuronal isoform OGDHneu uses two adjacent exons.

    “OGDH in NPCs (OGDH1) uses exon 4, whereas the isoform in neurons, here called OGDHneu, uses two adjacent exons”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Most down-regulated genes during neuronal differentiation were cell cycle related, whereas up-regulated genes were enriched in neuronal function pathways.

    “Most of the down-regulated genes were cell cycle related, whereas up-regulated genes were enriched in neuronal function pathways”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The lactate production/glucose consumption ratio was ~1.61 for NPCs and ~0.35 for neurons.

    “the ratio was ~1.61 lactate/glucose for NPCs, while in neurons, it was ~0.35”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    TCA genes remained at the same level during differentiation, except IDH2 dropped nearly 50%.

    “TCA genes remained at the same level during differentiation; an exception was isocitrate dehydrogenase, IDH2, the mitochondrial isoform of IDH, which dropped nearly 50%”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Except for PDK2, RNA levels of the other three PDKs declined significantly during neuronal differentiation, while PDP1 RNA levels increased ~2.5 fold at 3 weeks.

    “Except for PDK2, RNA levels of the other three PDKs declined significantly during neuronal differentiation; in contrast, PDP1 RNA levels increased ~2.5 fold at 3 weeks of differentiation”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Both PTB knockdown and Ngn2/Ascl1 overexpression generated Tuj1-positive neurons after 3 weeks of transduction.

    “Both approaches generated Tuj1-positive neurons after 3 weeks of transduction.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    No significant difference was found in amounts of mitochondrial respiratory complexes between NPCs and neurons at 1 and 3 weeks, except SDHB of complex II was consistently ~50% lower in neurons.

    “no significant difference was found between NPCs and neurons at 1 and 3 weeks, except for SDHB of complex II, which was consistently ~50% lower in neurons (Figure 5C).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    AMPK pT172 was clearly increased at day 4 of neuronal differentiation compared to control, but returned to control level at day 21.

    “Indeed, AMPK pT172 was clearly increased at day 4 of neuronal differentiation compared to control, but the phosphorylation returned to a level comparable to control at day 21, when extensive cell death had already occurred (Figure 4I).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    After the first week, neuronal mtDNA copy number doubled compared to NPCs, and at 3 weeks was increased to ~four fold.

    “after the first week, the neuronal mtDNA copy number doubled compared to NPCs, and at 3 weeks was increased to ~four fold.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The LDHA signal was clearly detected in GFAP-positive cells with anti-LDHA antibody or anti-FLAG antibody staining, but there was no colocalization with MAP2.

    “The LDHA signal was clearly detected in GFAP-positive cells with anti-LDHA antibody (Figure 4E) or anti-FLAG antibody staining (Figure 4H), but there was no colocalization with MAP2, a neuronal marker.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    PGC-1α was required to maintain the levels of ENO1 and PKM in glycolysis; CS, ACO2, OGDH and SUCLA2 in TCA cycle; TFAM and TFBM1 in mtDNA replication and transcription, and NRF1.

    “We found that PGC-1α was required to maintain the levels of ENO1 and PKM in glycolysis; CS, ACO2, OGDH and SUCLA2 in TCA cycle; TFAM and TFBM1 in mtDNA replication and transcription, and NRF1 (nuclear respiratory factor-1), a key transcription factor that activates nuclear genes encoding respiratory complex subunits (Figure 5F).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    During neuronal differentiation, UCP2 expression levels dropped significantly, and in 3-week neurons the UCP2 level was only 20% of that in NPCs.

    “We found that during neuronal differentiation, UCP2 expression levels dropped significantly, and in 3-week neurons the UCP2 level was only 20% of that in NPCs (Figure 5—figure supplement 1)”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingnone given

    Even though neurons rely on oxidative phosphorylation to generate energy, the expression of TCA and mitochondrial genes is not increased.

    “Surprisingly, even though neurons rely on oxidative phosphorylation to generate energy; the expression of TCA and mitochondrial genes is not increased.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Neurons rely on mitochondrial oxidative phosphorylation to meet energy demand.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Deletion of LDHA impairs maintenance and proliferation of hematopoietic stem and progenitor cells.

    “deletion of lactate dehydrogenase A (LDHA) impairs maintenance and proliferation of hematopoietic stem and progenitor cells (Wang et al., 2014)”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Wang et al., 2014

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    PGC-1α knockout mice show obvious neurodegenerative pathology.

    “PGC-1α knockout mice show obvious neurodegenerative pathology (Lin et al., 2004)”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Lin et al., 2004 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationown data

    An unknown mechanism links mitochondrial biogenesis to cell size.

    “indicating an unknown mechanism linking mitochondrial biogenesis to cell size”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    In embryonic Xenopus retina, NPCs are less reliant on oxidative phosphorylation for ATP production than differentiated neurons, and the transition from glycolysis to oxidative phosphorylation is tightly coupled to neuronal differentiation.

    “A pioneering study using embryonic Xenopus retina revealed that neural progenitor cells (NPCs) are less reliant on oxidative phosphorylation for ATP production than are non-dividing differentiated neurons, and the transition from glycolysis to oxidative phosphorylation is tightly coupled to neuronal differentiation”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Agathocleous et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    The loss of HK2 and LDHA expression and the PKM2-to-PKM1 splicing switch mark the transition from aerobic glycolysis in NPCs to neuronal oxidative phosphorylation.

    “marks the transition from aerobic glycolysis in neural progenitor cells (NPC) to neuronal oxidative phosphorylation”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Neurons rely on oxidative phosphorylation to meet energy demands.

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Only high-quality NPC lines containing more than 90% Sox2 and Nestin double-positive cells were used for experiments.

    “Only high-quality NPC lines containing more than 90% Sox2 and Nestin double-positive cells were used for experiments.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    The drastic decreases in HK2 and LDHA are consistent with neurons not having high flux aerobic glycolysis and exhibiting low lactate production.

    “The drastic decreases in HK2 and LDHA, the two key enzymes supporting aerobic glycolysis (DeBerardinis and Thompson, 2012; Dang, 2012), are consistent with neurons not having high flux aerobic glycolysis and exhibiting low lactate production.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites DeBerardinis and Thompson, 2012

      Not checked in the free run.

    • cites Dang, 2012

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    MYC directly activates HK2 and LDHA transcription, and the promoter regions of HK2 and LDHA contain MYC binding sites (5'-CACGTG-3'), also known as E box.

    “MYC, a key transcription factor associated with cell cycle progression, directly activates HK2 and LDHA transcription, and the promoter regions of HK2 and LDHA contain MYC binding sites (5'-CACGTG-3'), also known as E box (Kim et al., 2004).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Kim et al., 2004

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Denton et al. (2016) detected this splice form of OGDH in whole brain tissue and pancreatic islets, calling it LS1, and confirmed that LS1 isoform is insensitive to Ca2+.

    “Denton et al. (2016) also detected this splice form of OGDH in the whole brain tissue and pancreatic islets, calling it LS1, and, importantly, they confirmed that LS1 isoform is insensitive to Ca2+.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Denton et al., 2016

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductown data

    The median sequencing read yield for each RNA-seq sample was ~35 million 100-base reads, of which ~85% were mapped.

    “The median sequencing read yield for each sample was ~35 million 100-base reads, of which ~85% were mapped.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Overexpression of PGC-1α in cardiac cells triggers upregulation of hundreds of genes in glycolysis, TCA and mitochondrial respiratory complexes.

    “overexpression of PGC-1α in cardiac cells triggers the upregulation of hundreds of genes in glycolysis, TCA and mitochondrial respiratory complexes (Rowe et al., 2010).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Rowe et al., 2010

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    Neurons cannot tolerate constitutive HK2 and LDHA expression and die during differentiation.

    “The results suggest that neurons cannot tolerate constitutive HK2 and LDHA expression and die during differentiation.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Although mitochondrial mass increases on a per cell basis, mitochondrial density remains largely unchanged during neuronal differentiation.

    “It appears that although mitochondrial mass increases on a per cell basis, mitochondrial density remains largely unchanged during neuronal differentiation.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    A transcriptional circuit including PGC-1 family coactivators, ERRs and NRF is responsible for transcription of metabolic and mitochondrial genes in multiple tissues.

    “A transcriptional circuit, including PGC-1 family coactivators, estrogen-related receptors (ERRs) and nuclear respiratory factor (NRF), is responsible for the transcription of metabolic and mitochondrial genes in multiple tissues, such as heart and skeletal muscle (Scarpulla et al., 2012).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Scarpulla et al., 2012 source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    c-MYC and N-MYC redundantly promote neural progenitor cell proliferation in the developing brain, with double knockout mice exhibiting profoundly impaired growth.

    “c-MYC and N-MYC redundantly promote neural progenitor cell proliferation in the developing brain, with the brains of double knockout mice exhibiting profoundly impaired growth (Wey and Knoepfler, 2010).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Wey and Knoepfler, 2010

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Premisecitation

    Functional induced neurons (iN) can be generated directly from fibroblasts by forced expression of neural specific transcription factors, a process called transdifferentiation.

    “Functional induced neurons (iN) can be generated directly from fibroblasts by forced expression of neural specific transcription factors, a process also called transdifferentiation (Vierbuchen et al., 2010).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Vierbuchen et al., 2010 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    PGC-1α is induced in the neonatal mouse heart concurrent with the increase of mitochondrial energy-producing capacity.

    “PGC-1α is induced in the neonatal mouse heart concurrent with the increase of mitochondrial energy-producing capacity (Lehman et al., 2000).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Lehman et al., 2000 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    PGC-1α not only stimulates the induction of NRF1, but also binds to and coactivates the transcriptional function of NRF1 in muscle cells.

    “PGC-1α not only stimulates the induction of NRF1, but also binds to and coactivates the transcriptional function of NRF1 in muscle cells (Wu et al., 1999).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Wu et al., 1999

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    UCP2 is critical for maintaining stem cell glycolytic metabolism, and its level decreases during differentiation.

    “UCP2 is critical for maintaining stem cell glycolytic metabolism, and its level decreases during differentiation (Zhang et al., 2011; Yu et al., 2013).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Zhang et al., 2011

      Not checked in the free run.

    • cites Yu et al., 2013

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  • May combine two claims.

    Premisecitation

    MILS neurons have very low expression of HK2 and LDHA, and do not exhibit a significant increase in mitochondrial density.

    “MILS neurons also have very low expression of HK2 and LDHA, and do not exhibit a significant increase in mitochondrial density (Zheng et al., 2016).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Zheng et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    The critical role of LDHA in aerobic glycolysis has been proven in cancer cells, and LDHA depletion greatly decreases aerobic glycolysis and increases mitochondrial OXPHOS activity.

    “The critical role of LDHA in aerobic glycolysis has been proven in cancer cells, and LDHA depletion greatly decreases aerobic glycolysis and increases mitochondrial OXPHOS activity (Fantin et al., 2006).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Fantin et al., 2006

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    Neurons cannot use aerobic glycolysis or mitochondrial biogenesis to compensate for energy shortage.

    “It appears that neurons cannot use aerobic glycolysis or mitochondrial biogenesis to compensate for energy shortage.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    The A4 isozyme kinetically favors the conversion from pyruvate to lactate, while the B4 isozyme prefers converting lactate to pyruvate.

    “The A4 isozyme kinetically favors the conversion from pyruvate to lactate, while the B4 isozyme prefer converting lactate to pyruvate (Cahn et al., 1962).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Cahn et al., 1962

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In an iPSC-based disease model of MILS, MILS neurons but not their NPCs show severe energy deficiency; MILS NPCs generate more lactate than healthy control NPCs.

    “In an iPSC-based disease model of maternally inherited Leign syndrome (MILS), an early childhood neurodegenerative disease due to ATP synthase defect, we found that MILS neurons but not their NPCs show severe energy deficiency; MILS NPCs generate more lactate than healthy control NPCs.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Zheng et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Interpretationnone given

    Glucose transporter levels dramatically decrease in neurons, which limits glucose uptake and the production of glycolytic pyruvate.

    “Glucose transporter levels dramatically decrease in neurons, which limits glucose uptake and the production of glycolytic pyruvate.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    The decrease in expression of PDKs and the increase in PDPs may result in more active pyruvate dehydrogenase complexes (PDC), as demonstrated by decreased phosphorylation at Ser 300 of PDH E1 alpha protein (PDHA1), promoting pyruvate entry into the TCA cycle.

    “Fourth, the decrease in expression of PDKs and the increase in PDPs may result in more active pyruvate dehydrogenase complexes (PDC), as demonstrated by decreased phosphorylation at Ser 300 of PDH E1 alpha protein (PDHA1), promoting pyruvate entry into the TCA cycle.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Mitochondrial structure, dynamics and calcium concentration in the mitochondrial matrix are all involved in mitochondrial energy metabolism.

    “mitochondrial structure (cristae organization and size), dynamics (fusion and fission) and calcium concentration in the mitochondrial matrix are all involved in mitochondrial energy metabolism (Mishra and Chan, 2014).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Mishra and Chan, 2014

      Not checked in the free run.

  • May combine two claims.

    Interpretationnone given

    The decreased expression of HK2, and the GLUT1 and GLUT3 glucose transporters reduce glucose entry, terminating high flux glycolysis.

    “Third, the decreased expression of HK2, and the GLUT1 and GLUT3 glucose transporters reduce glucose entry, terminating high flux glycolysis.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    For each time point, two experimental duplicates were used for each independent NPC line and its differentiated neurons.

    “For each time point, two experimental duplicates were used for each independent NPC line and its differentiated neurons.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Hundreds of compounds were found to be capable of changing cellular mitochondrial content; the majority also change cell size accordingly.

    “in a high-throughput screen to identify small molecules interfering with mitochondrial abundance, hundreds of compounds were found to be capable of changing the cellular mitochondrial content; the majority of them also change cell size accordingly (Kitami et al., 2012)”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Kitami et al., 2012

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Picking only type 4 colonies showing sufficient maturity is critical for the purity of NPCs.

    “This step is critical for the purity of NPCs; only colonies (type 4) showing sufficient maturity as described in Figure 1—figure supplement 2 were picked.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Blocking neuronal oxidative utilization of lactate affects neuronal survival and even memory formation.

    “blocking neuronal oxidative utilization of lactate affects neuronal survival and even memory formation (Suzuki et al., 2011; Lee et al., 2012).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Suzuki et al., 2011 source

      Not checked in the free run.

    • cites Lee et al., 2012 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    c-MYC activation in the myocardium of adult mice induces mitochondrial biogenesis and glycolysis but reduces PGC-1α level.

    “c-MYC activation in the myocardium of adult mice induces mitochondrial biogenesis and glycolysis but reduces PGC-1α level (Ahuja et al., 2010).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Ahuja et al., 2010

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    RNA-seq experiments were performed with two NPC lines established from independent iPSC clones.

    “RNA-seq experiments were performed with two NPC lines established from independent iPSC clones.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Only RNA samples with RIN greater than 8.5 were used for library preparation.

    “only samples with RIN greater than 8.5 were used for library preparation.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    E2F upregulates mitochondrial genes, a function conserved from flies to mammals.

    “E2F also upregulates mitochondrial genes, a function conserved from flies to mammals (Ambrus et al., 2013).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Ambrus et al., 2013

      Not checked in the free run.

  • The quote only partly states this claim.

    Interpretationown data

    Glycolytic pyruvate deficiency is a major cause of death.

    “indicating that glycolytic pyruvate deficiency is a major cause of death.”
    Quote in the paperSubstantial claim
56 of 56

  • The paper relies on this without support.

    Little is known about the molecular basis of neurons' preference for oxidative phosphorylation or neuronal metabolic responses to energy crisis and their implications for disease progression.

    “Little is known about the molecular basis of neurons’ preference for oxidative phosphorylation, or neuronal metabolic responses to the energy crisis and their implications for disease progression.”

  • The paper relies on this without support.

    Neurons rely on oxidative phosphorylation to meet energy demands.

    “Neurons rely on oxidative phosphorylation to meet energy demands.”

  • The paper relies on this without support.

    Neurons rely on mitochondrial oxidative phosphorylation to meet energy demand.

    “Neurons rely on mitochondrial oxidative phosphorylation to meet energy demand.”

  • The paper relies on this without support.

    Aerobic glycolysis diverts pyruvate from mitochondrial oxidative phosphorylation for energy production by converting it into secreted lactate.

    “As aerobic glycolysis diverts pyruvate from mitochondrial oxidative phosphorylation for energy production by converting it into secreted lactate”

  • The paper relies on this without support.

    HK1 is still expressed in neurons.

    “HK1 is still expressed in neurons”

  • The paper relies on this without support.

    In proliferating NPCs, mitochondria are used for generation of biosynthetic precursors and mitochondria themselves have to be duplicated for daughter cells.

    “mitochondria are used for generation of biosynthetic precursors in these proliferating cells, and mitochondria themselves have to be duplicated for daughter cells.”

  • The paper relies on this without support.

    The expression levels of metabolic enzymes alone do not completely reflect their biochemical/physiological activities.

    “the expression levels of metabolic enzymes alone do not completely reflect their biochemical/physiological activities”
7 of 7

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • In multiple tissue contexts, adult stem cells primarily engage in aerobic glycolysis.

    “In multiple tissue contexts, adult stem cells have been found to primarily engage in aerobic glycolysis.”

    From Regulation and function of the mammalian tricarboxylic acid cycle (2023) · cites this paper as [205]

  • Upon lineage commitment and differentiation, cells increase their oxidation of pyruvate in the TCA cycle and display enhanced OXPHOS.

    “Upon lineage commitment and differentiation, however, cells increase their oxidation of pyruvate in the TCA cycle and display enhanced OXPHOS (203, 204, 205).”

    From Regulation and function of the mammalian tricarboxylic acid cycle (2023) · cites this paper as [205]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • NPC passage differs between the results and Figure 3 caption

    Minor

    The results say the HK2 and LDHA knockdown experiment used passage-3 NPCs, while the Figure 3C caption says passage 2.

  • Figure 1 caption reverses the TCA and PDH panel assignments

    Minor

    In the caption's ordered list, panel H corresponds to TCA and panel I to PDH. The results instead cite panel I for TCA genes and panel H for PDH-complex genes.

2 of 2

Cites

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Cited by

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Related

Authors
Zheng, X., Boyer, L., Jin, M., Mertens, J., Kim, Y., Ma, L., Ma, L., Hamm, M., Gage, F. H., & Hunter, T.
Affiliations
  • Salk Institute for Biological Studies (US)
  • Laboratory of Genetics, Salk Institute, La Jolla, United States
Corresponding author
  • Gage, Fred H
  • Hunter, Tony
ORCID
Venue
eLife
Publisher
eLife Sciences Publications, Ltd
Published
2016-06-10
Volume
5
ISSN
2050-084X (ISSN-L)
DOI
10.7554/elife.13374
PMC
PMC4963198
OpenAlex
W2416068580
Semantic Scholar
fbed50febe18cd07bb8e0c7a00e748286b5070dc
Cited by
671
References
77
Citations by year
2026: 82 · 2025: 76 · 2024: 84 · 2023: 81 · 2022: 74 · 2021: 85 · 2020: 65 · 2019: 49 · 2018: 39 · 2017: 33 · 2016: 1 · 2013: 1
FWCI
20.80
Type
journal article
Language
en
Open access
Licence
Locations
  • eLife (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • California Institute for Regenerative Medicine (TR2-01778)
  • Salk Pioneer postdoctoral fellowship
  • The G. Harold and Leila Y. Mathers Foundation
  • JPB Foundation
  • Leona M. and Harry B. Helmsley Charitable Trust (#2012-PG-MED002)
  • National Institutes of Health (CA14195, CA80100, CA82683)
Topics
  • Life Sciences › Neuroscience › Developmental Neuroscience › Neurogenesis and neuroplasticity mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Cancer, Hypoxia, and Metabolism
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolism, Diabetes, and Cancer
Keywords
metabolic reprogramming, neuronal differentiation, aerobic glycolysis, oxidative phosphorylation, hexokinase 2, LDHA, c-Myc, MYCN, PGC-1α, estrogen-related receptor gamma, mitochondrial biogenesis, neural progenitor cells, neuronal cell death, mitochondrial mass
MeSH terms
Aerobiosis, Cell Differentiation, Cells, Cultured, Glycolysis, Humans, Metabolism, Oxidative Phosphorylation, Gene Expression Regulation, Developmental, Neural Stem Cells

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-09.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-09.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

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