Extended Data Fig. 7: The rate of mtDNA synthesis depends on the rNTP/dNTP ratio.
From: Ribonucleotide incorporation into mitochondrial DNA drives inflammation
a, rNTP/dNTP ratios from cell extracts of immortalized wild-type MEFs determined by LC-MS. Cells were either left untreated or treated with 5-FU at 5 μM for 16 h (n = 5 independent cultures). b, De novo mtDNA synthesis in Yme1l+/+ and Yme1l−/− mitochondria in the presence or absence of exogenous dNTPs. Isolated mitochondria were pulse labelled for 1 h. Steady state levels of mtDNA and the level of newly synthesized mtDNA are shown in the left-hand side upper panels. The input fraction was analysed by immunoblotting (left-hand side lower panel). Quantification of de novo synthesized mtDNA in Yme1l+/+ and Yme1l−/− mitochondria is shown on the right panel. n = 3 biologically independent experiments. c, De novo mtDNA synthesis in mitochondria isolated from Yme1l+/+ and Yme1l−/− MEFs. Isolated mitochondria were pulsed labelled for 45 min and further incubated for 4 and 8 h. De novo and steady state mtDNA levels are shown (left panel). Newly synthesized mtDNA was quantified relative to mtDNA steady state levels in Yme1l+/+ and Yme1l−/− mitochondria (right panel). n = 2 biologically independent experiments. P values were calculated using unpaired two-tailed Student t-test (a) or using one-way ANOVA with Tukey’s multiple comparison test (b). Data are presented as mean ± SD.
