Fig 1: Nuclear localization of TCA cycle enzymes in somatic cell reprogramming.a Schematic diagram of TCA cycle enzymes in regulating epigenetics in the nucleus. b–i Nuclear location of TCA cycle enzymes (green) during the early stages of somatic cell reprogramming with SKOM. Isolated nuclei were stained with antibodies targeting Pdha1 (b), Pcb (c), Aco2 (d), Cs (e), Idh3a (f), Ogdh (g), Sdha (h), or Mdh2 (i) together with Lamin B1 (red) and DAPI (blue). Scale bars, 5 µm. The nuclei/mitochondria (N/M) fluorescent ratio of each TCA cycle enzyme during somatic cell reprogramming with SKOM present in the source files was shown at the lower panel. Data are presented as the mean ± S.D from three independent experiments containing at least 30 cells each. A two-tailed unpaired Student’s t-test was used. j, k, The western blot analysis of PDHA1, PCB, CS, ACO2, and IDH3A in the mitochondria (j) or nucleus (k) during somatic cell reprogramming induced by SKOM. The TOMM20 was used as an indicator of mitochondrial content, while H3 were used as loading controls. l The quantification of PDHA1, PCB, CS, ACO2, and IDH3A in the mitochondria (upper panel) or nucleus (lower panel) are presented on the right. Data are presented as the mean ± S.D and n = 3 independent experiments. A two-tailed unpaired Student’s t test was used.
Fig 2: Translocation of TCA cycle enzymes in primed-to-naive transition and totipotency stem cells.a, b Immunostaining of Pdha1, Pcb, Aco2, Cs, Idh3a, Ogdh, Sdha, and Mdh2 (purple) in mouse epiblast stem cells (EpiSCs) (a) or OG2 embryonic stem cells (naive stem cells) (b). The mito-DsRed (Mito) was used as a marker of mitochondria. Oct4::GFP is an indicator of embryonic stem cells and DAPI (blue) for indicating the nucleus. Scale bars, 5 µm. c The nuclei/mitochondria fluorescent (N/M) ratio of TCA cycle enzymes in EpiSCs and OG2 ESCs (Naive). Data are presented as the mean ± S.D of at least 30 cells from three independent experiments. A two-tailed unpaired Student’s t test was used. d, Immunostaining of Pdha1, Pcb, Aco2, Cs, Idh3a, Ogdh, Sdha, and Mdh2 (purple) in 2-cell stage-like cells (2CLCs). Oct4::GFP was used to track embryonic stem cells, 2C::tdTomato (red) was used to track 2CLCs, and DAPI (blue) for indicating the nucleus. Scale bars, 5 µm. Three independent experiments were repeated with similar results and one representative picture was presented (a, b, d).
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