LANDSCAPING OF THE HUMAN MITOCHONDRIAL RNOME BY COLOC-SEQ

Abstract

Mitochondria possess their own genome but critically depend on the import of many nuclear-encoded macromolecules to ensure its expression. Beside ~1,500 proteins, in apparently all eukaryotes, from protists to humans, some RNAs (tRNAs, 5S rRNA, miRNAs...) are partially redirected into mitochondria where they participate in gene expression processes(1). Therefore, the mitochondrial RNome represents an intricate mixture of transcriptome and importome. While studies of the mitochondrial transcriptome have now been facilitated by such techniques as RNA-seq(2,3), robust identification of nuclear-encoded transcripts imported into the organelles is still challenging since cytosolic contamination remains even after most thorough purification of mitochondria. Our laboratory is currently developing a conceptually novel experimental approach, Controlled Level of Contamination (CoLoC) which allows, by following RNase-mediated depletion dynamics of each transcript, to unequivocally distinguish between RNAs genuinely present inside mitochondria and mere contaminants. Coupled with deep sequencing (CoLoC-seq), this methodology will provide the first global view of the human mitochondrial RNA importome in diverse cell types and conditions. This knowledge will help a better understanding of nuclear-mitochondrial communication and will open ways to exploit RNA targeting pathways for manipulation of the mitochondrial genetic system and development of therapeutic approaches to currently incurable mitochondrial diseases(4).