Yeast as a model of human mitochondrial tRNA base substitutions: Investigation of the molecular basis of respiratory defects
- Arianna Montanari1,6,
- Céline Besagni2,6,
- Cristina De Luca1,
- Veronica Morea3,
- Romina Oliva4,
- Anna Tramontano5,
- Monique Bolotin-Fukuhara2,
- Laura Frontali1, and
- Silvia Francisci1
- 1Department of Cell and Developmental Biology, Pasteur Institute-Fondazione Cenci Bolognetti, University Sapienza, 00185 Rome, Italy
- 2Laboratoire de Génétique Moléculaire, Institut de Genétique et Microbiologie UMR CNRS 8621, Université Paris Sud, 91405 Orsay Cedex, France
- 3CNR-Institute of Molecular Biology and Pathology (IBPM), Pasteur Institute-Fondazione Cenci Bolognetti, University Sapienza, 00185 Rome, Italy
- 4Department of Applied Sciences, University Parthenope, 80133 Naples, Italy
- 5Department of Biochemical Sciences, Pasteur Institute-Fondazione Cenci Bolognetti, University Sapienza, 00185 Rome, Italy
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↵6 These authors contributed equally to this work.
Abstract
We investigate the relationships between acylation defects and structure alterations due to base substitutions in yeast mitochondrial (mt) tRNAUUR Leu. The studied substitutions are equivalent to the A3243G and T3250C human pathogenetic tRNA mutations. Our data show that both mutations can produce tRNAUUR Leu acylation defects, although to a different extent. For mutant A14G (equivalent to MELAS A3243G base substitution), the presence of the tRNA and its defective aminoacylation could be observed only in the nuclear context of W303, a strain where the protein synthesis defects caused by tRNA base substitutions are far less severe than in previously studied strains. For mutant T20C (equivalent to the MM/CPEO human T3250C mutation), the acylation defect was less severe, and a thermosensitive acylation could be detected also in the MCC123 strain. The correlation between the severity of the in vivo phenotypes of yeast tRNA mutants and those obtained in in vitro studies of human tRNA mutants supports the view that yeast is a suitable model to study the cellular and molecular effects of tRNA mutations involved in human pathologies. Furthermore, the yeast model offers the possibility of modulating the severity of yeast respiratory phenotypes by studying the tRNA mutants in different nuclear contexts. The nucleotides at positions 14 and 20 are both highly conserved in yeast and human mt tRNAs; however, the different effect of their mutations can be explained by structure analyses and quantum mechanics calculations that can shed light on the molecular mechanisms responsible for the experimentally determined defects of the mutants.
Keywords
Footnotes
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Reprint requests to: Silvia Francisci, Department of Cell and Developmental Biology, University of Rome I, Piazzale A. Moro 5, 00185 Rome, Italy; e-mail: silvia.francisci{at}uniroma1.it; fax: +39 06 4461980.
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Abbreviations: mt, mitochondrial; cyt, cytoplasmic; MELAS, mitochondrial encephalomyophathy, lactic acidosis, and stroke-like episodes; MM/CPEO, mitochondrial myopathy/chronic progressive external ophthalmoplegia; TUF1, gene encoding for the mitochondrial protein synthesis elongation factor (EF-Tu); RH, reverse Hoogsten; syn−, mitochondrial mutation that leads to mitochondrial protein synthesis deficiency; rho, indicates the state of the mitochondrial DNA: rho+ (wild-type), rho° (absent), rho− (large DNA deletion).
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Article published online ahead of print. Article and publication date are at http://www.rnajournal.org/cgi/doi/10.1261/rna.740108.
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- Received July 24, 2007.
- Accepted October 16, 2007.
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Freely available online through the open access option.
- Copyright © 2008 RNA Society
