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. 2016 Feb 4;35(7):724–742. doi: 10.15252/embj.201592862

Figure EV4. PGC‐1β‐dependent mitochondrial biogenesis modulates cellular senescence.

Figure EV4

  1. (left) Representative images and (right) histograms showing the distribution of MitoTracker green (MTG) immunofluorescence in proliferating and senescent (3 days after 10‐Gy X‐ray) wild‐type and PGC‐1β −/− MEFs. Scale bar = 10 μm. Dashed red lines indicate median MitoTracker green intensity (50–100 cells were quantified per condition).
  2. Kinetics of mtDNA copy number after 10‐Gy X‐ray in wild‐type and PGC‐1β −/− MEFs. Data are mean ± SEM of n = 3 independent experiments.
  3. Expression of the mitochondrial outer membrane protein VDAC in wild‐type and PGC‐1β −/− MEFs 1 day after 10‐Gy X‐ray. Data are mean ± SEM of n = 3 independent experiments.
  4. Knockdown efficiency of PGC‐1β in human MRC5 fibroblasts infected with a control vector (shScr) and with a shPGC‐1β vector by Western blot and qPCR.
  5. Quantification of mitochondrial mass (NAO intensity), mitochondrial ROS (MitoSOX intensity), mean number (N) of 53PB1 foci and Edu‐positive cells in proliferating and senescent MRC5 fibroblasts infected with a control (shScr) and shPGC‐1β vector. Mitochondrial mass and ROS measurements were performed 3 days after 20‐Gy X‐ray (data are mean ± SD of n = 3 technical replicates). 53BP1 foci and EdU incorporation were analysed 10 days after 20‐Gy X‐ray (data are mean ± SD of 80–100 cells/condition for 53BP1 analysis and 10 random planes for the EdU incorporation analysis).
  6. Quantification of Sen‐β‐Gal‐positive cells in proliferating and senescent (10 days after 20‐Gy X‐ray) MRC5 fibroblasts infected with control (shScr) and shPGC‐1β vectors. Data are mean ± SD 10 random planes.
  7. Representative image of FLAG‐PGC‐1β immunostaining 2 days following the transfection of a pcDNA empty vector and pcDNAf:PGC‐1β in wild‐type MEFs. Scale bar = 20 μm.
  8. mRNA expression of PGC‐1β in proliferating and senescent (10 days after 10‐Gy X‐ray) wild‐type MEFs following the transfection with pcDNA empty vector and pcDNAf:PGC‐1β. Data are mean ± SEM of n = 3 independent experiments.
  9. (top) Representative image of FLAG‐PGC‐1β (red) and the mitochondrial protein SDHA (green) double immunostaining and (bottom) histograms showing the distribution of SDHA fluorescence, 2 days after transfection with a pcDNA empty vector and pcDNAf:PGC‐1β in wild‐type MEFs. Scale bar = 10 μm. Dashed red lines indicate median SDHA fluorescence (100 cells were quantified per condition).