Figure 6.
Perturbation of rRNA synthesis leads to mitochondrial impairment followed by increased oxidative stress. A, The bars represent abundance of YY1 transcripts by qPCR normalized to HPRT in control and TIF-IADATCreERT2 mice 2 weeks after tamoxifen (n = 3). B, The graph shows UCP-2 gene expression by qPCR. The bars represent abundance of UCP-2 transcripts normalized to the levels HPRT 2 and 3 weeks after tamoxifen treatment (n = 3). C, Two weeks after tamoxifen injection, reduced COX activity in TIF-IADATCreERT2 mutants is measured as optical density compared with control mice (n = 3). D–I, Brain sections through the ventral midbrain were analyzed for neuroketals (D, E) as marker for ROS-induced lipid damage (brown), for nitrosylated proteins (NITT) (F, G), for 8-hydroxydeoxyguanosine (8-OHdG) (H, I), as marker for ROS-induced DNA damage (brown), in combination with TH staining (light blue) to identify DA. The insets show higher magnification of immunostained cells. Scale bars, 50 μm. J, Quantification of DA neurons positive for oxidative stress markers NITT, neuroketals, and 8-OHdG in control and mutant mice 2 and 4 weeks after injection with tamoxifen. Differences are expressed as fold change compared with the mean of the controls at different stages. Higher levels of oxidative stress markers were observed in the mutants (n = 4). *p < 0.05; **p < 0.01. Error bars represent SEM. K, Diagram showing the sequence of events triggered by nucleolar damage. The decrease and increase of the parameters indicated (left) are depicted in green or red, respectively. The bars indicate the time points (weeks) analyzed after injection of tamoxifen to induce TIF-IA loss in adult mice. L, Schematic representation showing the molecular mechanisms shared between TIF-IA mutation and MPTP pharmacological models. Nucleolar damage as a consequence of TIF-IA mutation induces p53 and inhibits mTOR, causing mitochondrial dysfunction and increased oxidative damage. In the pharmacological model, mitochondrial dysfunction caused by MPTP leads to increased oxidative stress. This increase inhibits mTOR and rRNA synthesis, probably by downregulation of TIF-IA activity, causing nucleolar damage and additional consequences on cell survival.