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Modification of the universally unmodified uridine-33 in a mitochondria-imported edited tRNA and the role of the anticodon arm structure on editing efficiency

Published online by Cambridge University Press:  20 August 2002

PAMELA F. CRAIN ,
JUAN D. ALFONZO ,
JEF ROZENSKI ,
STEPHEN T. KAPUSHOC ,
JAMES A. MCCLOSKEY  and
LARRY SIMPSON
PAMELA F. CRAIN
Affiliation:
Department of Medicinal Chemistry, University of Utah, Salt Lake City, Utah 84112, USA
JUAN D. ALFONZO
Affiliation:
Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, California 90095, USA
JEF ROZENSKI
Affiliation:
Department of Medicinal Chemistry, University of Utah, Salt Lake City, Utah 84112, USA Present address: Rega Institute for Medical Research, B-3000 Leuven, Belgium.
STEPHEN T. KAPUSHOC
Affiliation:
Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, California 90095, USA
JAMES A. MCCLOSKEY
Affiliation:
Department of Medicinal Chemistry, University of Utah, Salt Lake City, Utah 84112, USA Department of Biochemistry, University of Utah, Salt Lake City, Utah 84112, USA
LARRY SIMPSON
Affiliation:
Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, California 90095, USA Howard Hughes Medical Institute, University of California, Los Angeles, California 90095, USA
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Abstract

Editing of tRNA has a wide phylogenetic distribution among eukaryotes and in some cases serves to expand the decoding capacity of the target tRNA. We previously described C-to-U editing of the wobble position of the imported tRNATrp in Leishmania mitochondria, which is essential for decoding UGA codons as tryptophan. Here we show the complete set of nucleotide modifications in the anticodon arm of the mitochondrial and cytosolic tRNATrp as determined by electrospray ionization mass spectrometry. This analysis revealed extensive mitochondria-specific posttranscriptional modifications, including the first example of thiolation of U33, the “universally unmodified” uridine. In light of the known rigidity imparted on sugar conformation by thiolation, our discovery of a thiolated U33 suggests that conformational flexibility is not a universal feature of the anticodon structural signature. In addition, the in vivo analysis of tRNATrp variants presented shows a single base-pair reversal in the anticodon stem of tRNATrp is sufficient to abrogate editing in vivo, indicating that subtle changes in anticodon structure can have drastic effects on editing efficiency.

Keywords

editingimportmitochondriamodificationthiolationtRNAU33
Type
Research Article
Information
RNA , Volume 8 , Issue 6 , June 2002 , pp. 752 - 761
Copyright
© 2002 RNA Society

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