Figure 4. Facilitated OGS and early EMT form a positive feedback loop.

• A, B
  Expression of Hif1α was modulated via a retrovirus system or small‐molecule compounds, oligomycin (1 μM) and 2‐DG (5 mM), during reprogramming. The expression of mesenchymal markers (A) and cell migration (B) was determined with qPCR and live‐cell imaging, respectively, on day 6.
• C
  HIF1α induces EMT by transcriptionally activating key mesenchymal transcriptional factors, including Snai1/2, Twist1/2, and Zeb1/2. Although to a less extent, inducing OGS directly with Pdk1/2 or small molecules also activate these factors.
• D–G
  RepSox (1 μM) was used on days 2–7 to inhibit early EMT during reprogramming with 5C medium. TGFβ (TGFβ1/2/3, 1 ng/ml each) on days 2–7, LSD1i (SP2509, 10 nM), and sequential introduction of Yamanaka factors (OK + M + S) were used to induce early EMT during reprogramming with mES medium (D). Except for OK + M + S group, the four Yamanaka factors were delivered simultaneously. Energy metabolism was determined on day 6 with Seahorse instrument (E and F). The expression of glycolysis markers was determined on day 6 with qPCR (G).
• H
  Key mesenchymal transcriptional factors, including Snai1/2, Twist1/2, and Zeb1/2, induce OGS by transcriptionally activating Hif1α and glycolysis genes.
• I
  Early EMT and facilitated OGS form a positive feedback loop to induce pluripotency.

Data information: Experiments were independently repeated at least five times (n ≥ 5). Error bars represent standard deviations. ***P < 0.001. Additional statistical information is listed in Dataset EV7.