Figure 5.

MTO1 interacts with mitoribosomal subunits and affects mitoribosome assembly. (A) Immunoprecipitation of MTO1 from HeLa cells expressing flag-tagged MTO1 from a TET promoter. Immunoprecipitations were performed in the absence and presence of RNase. The eluate was probed by immunoblotting for the presence of the mitoribosomal proteins MRPL37 and MRPS35 as well as for HSP60 and HSP70. (B) Western blot analysis and quantification of mitochondria of WT, heterozygous and homozygous MTO1 mice to assess enzymes connected to the mitochondrial translation. Steady-state levels of mitoribosomal proteins MRPS27/10 and MRPL12 tested. Porin (VDAC) was used as a loading control. Depicted are representative blots. (C–F) Analysis of mitoribosomal assembly by sucrose gradient ultracentrifugation using mitochondrial extracts. Sedimentation of the 28S (small subunit), 39S (large subunit) and 55S (monosome) were determined by western blot analysis. Shown is the quantification of the western blots. The amount relative to the input is depicted. See also Supplementary Material, Figure S4. **P < 0.01, ***P < 0.001.