The assembly pathway of the mammalian mitochondrial ribosome.

• Applicant: Professorin Dr. Ricarda Richter-Dennerlein
• Subject Area: Biochemistry
• Term: since 2016
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 280677030

The human mitochondrial ribosome (mitoribosome) is of central importance as it synthesizes the mitochondrial core components of the oxidative phosphorylation (OXPHOS) system. Defects in mitoribosome biogenesis and function leads to OXPHOS deficiency and to severe early-onset mitochondrial diseases mostly affecting high-energy demanding tissues. Thus, it is crucial to dissect the molecular mechanism of mitoribosome assembly and the consequences of defects during this complex pathway to understand the associated pathophysiological clinical presentation. Although the mitoribosome derived from a bacterial ancestor, its structure and composition is distinct from its bacterial and cytosolic counterparts. Thus, we cannot simply transfer our knowledge from other systems to understand the biogenesis of the human mitoribosome. Our recent work and studies by other research groups presented the final steps of large and small mitoribosomal subunits (mtLSU, mtSSU) maturation explaining the function of some essential auxiliary factors. However, we are just at the beginning to understand this process. In the on-going project and proposed work package for the 4th funding period within the Emmy Noether program we aim to investigate early steps during mtSSU and mtLSU biogenesis and to reveal the function of involved assembly factors. We apply complementary approaches to investigate the assembly pathway in vivo and in vitro by dissecting ribosome assembly intermediates in genetic-engineered knockout and mutant cell lines followed by biochemical and structural analyses. We especially focus on mitochondrial-specific features of the human mitoribosome such as the central protuberance of the mtLSU and the foot of the mtSSU. The central protuberance contains mainly mitochondria-specific proteins, a tRNA and a functional peptidyl tRNA hydrolase (mL62) as structural constituents. First, we ask how the dual function of mL62 as a structural component of the mtLSU and as a potential ribosome rescue factor is regulated. Knockout and catalytic inactive mutant variants, respectively, will be used to distinguish the cellular consequences of loss of mL62 being a ribosomal protein from mL62 acting as a ribosome rescue factor. Second, we will study the assembly and the role of the foot of the mtSSU, which represents a hotspot for mitochondrial disease-associated mutations. Third, we will focus on the role of GTPases as potential quality control and anti-association factors during mtSSU biogenesis. Our final studies within the Emmy Noether project will provide crucial mechanistic insights into the assembly of the mitoribosome in human health and disease and will advance our understanding of pathophysiological defects during this essential process.

• DFG Programme: Independent Junior Research Groups