Abstract |
Two mutations (G8363A and A8296G) in the mtDNA ( mitochondrial DNA) tRNA(Lys) gene have been associated with severe mitochondrial diseases in a number of reports. Their functional significance, however, remains unknown. We have already shown that homoplasmic cybrids harbouring the A8296G mutation display normal oxidative phosphorylation, although the possibility of a subtle change in mitochondrial respiratory capacity remains an open issue. We have now investigated the pathogenic mechanism of another mutation in the tRNA(Lys) gene (G8363A) by repopulating an mtDNA-less human osteosarcoma cell line with mitochondria harbouring either this genetic variant alone or an unusual combination of the two mutations (A8296G+G8363A). Cybrids homoplasmic for the single G8363A or the A8296G+G8363A mutations have defective respiratory-chain enzyme activities and low oxygen consumption, indicating a severe impairment of the oxidative phosphorylation system. Generation of G8363A cybrids within a wild-type or the A8296G mtDNA genetic backgrounds resulted in an important alteration in the conformation of the tRNA(Lys), not affecting tRNA steady-state levels. Moreover, mutant cybrids have an important decrease in the proportion of amino-acylated tRNA(Lys) and, consequently, mitochondrial protein synthesis is greatly decreased. Our results demonstrate that the pathogenicity of the G8363A mutation is due to a change in the conformation of the tRNA that severely impairs aminoacylation in the absence of changes in tRNA stability. The only effect detected in the A8296G mutation is a moderate decrease in the aminoacylation capacity, which does not affect mitochondrial protein biosynthesis.
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Authors | Belén Bornstein, José Antonio Mas, Clarice Patrono, Miguel Angel Fernández-Moreno, Emiliano González-Vioque, Yolanda Campos, Rosalba Carrozzo, Miguel Angel Martín, Pilar del Hoyo, Filippo M Santorelli, Joaquín Arenas, Rafael Garesse |
Journal | The Biochemical journal
(Biochem J)
Vol. 387
Issue Pt 3
Pg. 773-8
(May 01 2005)
ISSN: 1470-8728 [Electronic] England |
PMID | 15554876
(Publication Type: Comparative Study, Journal Article, Research Support, Non-U.S. Gov't)
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Chemical References |
- DNA, Mitochondrial
- RNA, Transfer, Lys
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Topics |
- Aminoacylation
- Cell Line, Tumor
- DNA, Mitochondrial
(genetics)
- Gene Expression Regulation
(genetics)
- Humans
- MERRF Syndrome
(genetics, physiopathology)
- Mitochondria
(metabolism)
- Mutation
- Phenotype
- Protein Conformation
- RNA, Transfer, Lys
(genetics, physiology)
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