Antonio Barrientos, Ph.D., is a professor of Neurology, Biochemistry and Molecular Biology with the Department of Neurology at the University of Miami Miller School of Medicine. He is also a member of the Biochemistry, Neuroscience and Pharmacology graduate programs, with secondary appointment to Biochemistry and Molecular Biology at the University of Miami.
The Human Mitochondrial mRNA Folding Landscape
Antoni Barrientos1 J. Conor Moran1, Amir Brivanlou2, Flavia Fontanesi1, Silvi Rouskin2
1Department of Neurology. Department of Biochemistry and Molecular Biology. University of Miami Miller School of Medicine; 1600 NW 10th Ave. Miami, FL-33136 (USA).
2Department of Microbiology, Harvard Medical School, 77 Ave. Louis Pasteur. Boston, MA-02115 (USA).
Messenger RNAs (mRNAs) fold to adopt secondary and tertiary structures that often have essential regulatory functions in their synthesis, processing, translation, and turnover. In mammalian cells, mitochondria contain a genome that encodes for 13 protein components of the oxidative phosphorylation system that drives aerobic energy transduction. These 13 proteins are translated from 9 monocistronic transcripts and 2 bicistronic mRNAs with overlapping reading frames. The native structures of mitochondrial mRNAs (mt-mRNAs) have not been investigated, and therefore fundamental molecular determinants of mitochondrial gene expression remain unknown.
Here, we have optimized and applied a mitochondrial dimethyl sulfate (DMS) probing technique coupled with mutational profiling through next-generation sequencing (Mito-DMS-MaPseq) to assess the native architecture of all mitochondrial mRNAs. The data provide insights into mitochondrial mRNA biology and, in combination with biochemical data, support translation regulatory mechanisms. These include programmed translational frameshifting, termination-reinitiation events, and mRNA-programmed translational pausing. The approach described here will allow elucidation of mt-mRNA structuromes and interactions in any cell type or tissue and help illuminate novel roles of RNA in mitochondrial gene expression regulation in health and disease.