Mitochondria contain dedicated ribosomes (mitoribosomes), which synthesize the mitochondrial-encoded core components of the oxidative phosphorylation complexes. The RNA and protein components of mitoribosomes are encoded on two different genomes (mitochondrial and nuclear) and are assembled into functional complexes with the help of dedicated factors inside the organelle. Defects in mitoribosome biogenesis are associated with severe human diseases, yet the molecular pathway of mitoribosome assembly remains poorly understood. Here, we applied a multidisciplinary approach combining biochemical isolation and analysis of native mitoribosomal assembly complexes with quantitative mass spectrometry and mathematical modeling to reconstitute the entire assembly pathway of the human mitoribosome. We show that, in contrast to its bacterial and cytosolic counterparts, human mitoribosome biogenesis involves the formation of ribosomal protein-only modules, which then assemble on the appropriate ribosomal RNA moiety in a coordinated fashion. The presence of excess protein-only modules primed for assembly rationalizes how mitochondria cope with the challenge of forming a protein-rich ribonucleoprotein complex of dual genetic origin. This study provides a comprehensive roadmap of mitoribosome biogenesis, from very early to late maturation steps, and highlights the evolutionary divergence from its bacterial ancestor.

线粒体包含专用的核糖体（线粒体核糖体），它合成氧化磷酸化复合物的线粒体编码的核心成分。线粒体核糖体的 RNA 和蛋白质成分编码在两个不同的基因组（线粒体和核）上，并在细胞器内部专用因子的帮助下组装成功能复合物。线粒体核糖体生物发生的缺陷与严重的人类疾病有关，但对线粒体组装的分子途径仍然知之甚少。在这里，我们应用了一种多学科方法，将天然线粒体组装复合物的生化分离和分析与定量质谱和数学建模相结合，以重建人类线粒体的整个组装途径。我们表明，与其细菌和胞质对应物相比，人类线粒体核糖体生物发生涉及仅核糖体蛋白模块的形成，然后这些模块以协调的方式组装在适当的核糖体 RNA 部分上。为组装准备的过量纯蛋白质模块的存在使线粒体如何应对形成双遗传来源的富含蛋白质的核糖核蛋白复合物的挑战合理化。本研究提供了线粒体核糖体生物发生的全面路线图，从非常早期到晚期成熟步骤，并强调了与其细菌祖先的进化差异。