New Mechanisms of the Pseudouridine Synthase Module in Mitoribosome Assembly

PROJECT SUMMARY/ABSTRACT RNA modifiers coordinate essential steps of the mitochondrial ribosome assembly pathway; yet our limited understanding of the signals controlling these RNA editing factors hinders our knowledge of how these important regulators misshape the mitochondrial proteome in complex diseases. In mammals, the RPUSD4 pseudouridine (Y) synthase collaborates with newly identified auxiliary factors to catalyze pseudouridylation of mitochondrial ribosomal RNA (rRNA). Although the factors necessary for mitochondrial rRNA pseudouridylation have been annotated, the molecular mechanisms by which the Y synthase module exerts exquisite specificity towards assembling mitochondrial ribosomes is unknown. Moreover, mounting evidence links dysregulated rRNA pseudouridylation to mitochondrial genetic disorders, underscoring the importance of this underexplored RNA modification regulatory network. To address these shortcomings, we will apply a cross-disciplinary program combining hybrid structural biology, genomics, and biochemistry. This integrative approach will address critical questions in mitochondrial RNA processing, including: 1) How does the RPUSD4 Y synthase exerts strict specificity for its single rRNA modification site? 2) Why do mitochondrial disease-mutations dysregulate the Y synthase module? 3) How does the Y synthase module recognize assembling ribosomes? Defining the intricate molecular signals coordinating the fundamental process of rRNA pseudouridylation will set a framework to launch innovative programs that exploit mitochondrial RNA processing systems for the benefit of mitochondrial disease patients.