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J. Biol. Chem., Vol. 275, Issue 48, 37907-37914, December 1, 2000
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From the Departments of All mitochondrial tRNAs in Leishmania
tarentolae are encoded in the nuclear genome and imported into
the mitochondrion from the cytosol. One imported tRNA
(tRNATrp) is edited by a C to U modification at the first
position of the anticodon. To determine the in vivo
substrates for mitochondrial tRNA importation as well as tRNA editing,
we examined the subcellular localization and extent of 5'- and 3'-end
maturation of tRNATrp(CCA), tRNAIle(UAU),
tRNAGln(CUG), tRNALys(UUU), and
tRNAVal(CAC). Nuclear, cytosolic, and mitochondrial
fractions were obtained with little cross-contamination, as determined
by Northern analysis of specific marker RNAs. tRNAGln was
mainly cytosolic in localization; tRNAIle and
tRNALys were mainly mitochondrial; and tRNATrp
and tRNAVal were shared between the two compartments. 5'-
and 3'-extended precursors of all five tRNAs were present only in the
nuclear fraction, suggesting that the mature tRNAs represent the
in vivo substrates for importation into the mitochondrion.
Consistent with this model, T7-transcribed mature tRNAIle
underwent importation in vitro into isolated mitochondria
more efficiently than 5'-extended precursor
tRNAIle. 5'-Extended precursor
tRNATrp was found to be unedited, which is consistent with
a mitochondrial localization of this editing reaction. T7-transcribed
unedited tRNATrp was imported in vitro more
efficiently than edited tRNATrp, suggesting the presence of
importation determinants in the anticodon.
End Processing Precedes Mitochondrial Importation and Editing
of tRNAs in Leishmania tarentolae*
§,
, and
Molecular, Cell, and
Developmental Biology and ¶ Microbiology, Immunology, and
Molecular Genetics and the ** Howard Hughes Medical
Institute, University of California,
Los Angeles, California 90095
*
This work was supported in part by a Human Frontier Science
Program grant (to L. S.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Supported in part by a National Science Foundation Graduate
Research Fellowship.
To whom correspondence should be addressed: Howard Hughes
Medical Inst., University of California, 675 Charles E. Young Dr. South, Los Angeles, CA 90095. Tel.: 310-825-4215; Fax: 310-206-8967; E-mail: simpson@hhmi.ucla.edu.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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