期刊
RNA
卷 16, 期 5, 页码 926-941出版社
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1261/rna.1914110
关键词
mitochondrial tRNA import; RNA aptamers; small synthetic RNAs; SELEX; in-gel FRET
资金
- Centre National de Recherche Scientifique ( CNRS)
- Universite de Strasbourg ( UdS)
- Association Francaise contre les Myopathies ( AFM)
- Agence Nationale de la Recherche ( ANR)
- Fondation pour la Recherche Medicale ( FRM)
- Russian Foundation for Basic Research ( RFBR)
- ARCUS cooperation program Alsace-Russia-Ukraine
In the yeast Saccharomyces cerevisiae, nuclear DNA-encoded tRNA(CUU)(Lys) is partially imported into mitochondria. We previously found that the synthetic transcripts of yeast tRNA(Lys) and a number of their mutant versions could be specifically internalized by isolated yeast and human mitochondria. The mitochondrial targeting of tRNA(Lys) in yeast was shown to depend on the cytosolic precursor of mitochondrial lysyl-tRNA synthetase and the glycolytic enzyme enolase. Here we applied the approach of in vitro selection (SELEX) to broaden the spectrum of importable tRNA-derived molecules. We found that RNAs selected for their import into isolated yeast mitochondria have lost the potential to acquire a classical tRNA-shape. Analysis of conformational rearrangements in the importable RNAs by in-gel fluorescence resonance energy transfer ( FRET) approach permitted us to suggest that protein factor binding and subsequent import require formation of an alternative structure, different from a classic L-form tRNA model. We show that in the complex with targeting protein factor, enolase 2, tRK1 adopts a particular conformation characterized by bringing together the 3'-end and the T Psi C loop. This is a first evidence for implication of RNA secondary structure rearrangement in the mechanism of mitochondrial import selectivity. Based on these data, a set of small RNA molecules with significantly improved efficiency of import into yeast and human mitochondria was constructed, opening the possibility of creating a new mitochondrial vector system able to target therapeutic oligoribonucleotides into deficient human mitochondria.
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