Nucleic Acids Research, Vol 27, Issue 3 756-763, Copyright © 1999 by Oxford University Press
M Helm, C Florentz, A Chomyn and G Attardi
Post-transcriptional modifications are characteristic features of tRNAs and
have been shown in a number of cases to influence both their structural and
functional properties, including structure stabilization, amino-acylation
and codon recognition. We have developed an approach which allows the
investigation of the post-transcriptional modification patterns of human
mitochondrial wild-type and mutant tRNAs at both the qualitative and the
quantitative levels. Specific tRNA species are long-term labeled in vivo
with [32P]orthophosphate, isolated in a highly selective way, enzymatically
digested to mononucleotides and then subjected to two-dimensional thin
layer chromatographic analysis. The wild-type tRNALysand the corresponding
tRNALyscarrying the A8344G mutation associated with the MERRF (Myoclonic
Epilepsy with Ragged Red Fibers) syndrome exhibit the same modified
nucleotides at the same molar concentrations. By contrast, a quantitatively
different modification pattern was observed between the wild-type
tRNALeu(UUR)and its counterpart carrying the A3243G mutation associated
with the MELAS (Mitochondrial Myopathy, Encephalopathy with Lactic Acidosis
and Stroke-like episodes) syndrome, the latter exhibiting a 50% decrease in
m2G content. Complementary sequencing of tRNALeu(UUR)has allowed the
localization of this modification at position 10 within the D-stem of the
tRNA. The decreased level of this modification may have important
implications for understanding the molecular mechanism underlying the
MELAS-associated mitochondrial dysfunction.
ARTICLES
Search for differences in post-transcriptional modification patterns of mitochondrial DNA-encoded wild-type and mutant human tRNALys and tRNALeu(UUR)
Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.
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