Unconventional decoding of the AUA codon as methionine by mitochondrial tRNAMet with the anticodon f5CAU as revealed with a mitochondrial in vitro translation system

Chie Takemoto; Linda L. Spremulli; Lisa A. Benkowski; Takuya Ueda; Takashi Yokogawa; Kimitsuna Watanabe

doi:10.1093/nar/gkp001

Unconventional decoding of the AUA codon as methionine by mitochondrial tRNA^Met with the anticodon f⁵CAU as revealed with a mitochondrial in vitro translation system

Chie Takemoto^*, Linda L. Spremulli, Lisa A. Benkowski, Takuya Ueda, Takashi Yokogawa, Kimitsuna Watanabe

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

76 Citations (Scopus)

Abstract

Mitochondrial (mt) tRNA^Met has the unusual modified nucleotide 5-formylcytidine (f⁵C) in the first position of the anticodon. This tRNA must translate both AUG and AUA as methionine. By constructing an in vitro translation system from bovine liver mitochondria, we examined the decoding properties of the native mt tRNA^Met carrying f⁵C in the anticodon compared to a transcript that lacks the modification. The native mt Met-tRNA could recognize both AUA and AUG codons as Met, but the corresponding synthetic tRNA^Met lacking f⁵C (anticodon CAU), recognized only the AUG codon in both the codon-dependent ribosomal binding and in vitro translation assays. Furthermore, the Escherichia coli elongator tRNA^Met_m with the anticodon ac⁴CAU (ac⁴C-4-acetylcytidine) and the bovine cytoplasmic initiator tRNA^Met (anticodon CAU) translated only the AUG codon for Met on mt ribosome. The codon recognition patterns of these tRNAs were the same on E. coli ribosomes. These results demonstrate that the f⁵C modification in mt tRNA^Met plays a crucial role in decoding the nonuniversal AUA codon as Met, and that the genetic code variation is compensated by a change in the tRNA anticodon, not by a change in the ribosome. Base pairing models of f⁵C-G and f⁵C-A based on the chemical properties of f⁵C are presented.

Original language	English
Pages (from-to)	1616-1627
Number of pages	12
Journal	Nucleic acids research
Volume	37
Issue number	5
DOIs	https://doi.org/10.1093/nar/gkp001
Publication status	Published - 2009
Externally published	Yes

ASJC Scopus subject areas

Genetics

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10.1093/nar/gkp001

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@article{472c9fb3dd8e4282a9c313e3c13c551c,

title = "Unconventional decoding of the AUA codon as methionine by mitochondrial tRNAMet with the anticodon f5CAU as revealed with a mitochondrial in vitro translation system",

abstract = "Mitochondrial (mt) tRNAMet has the unusual modified nucleotide 5-formylcytidine (f5C) in the first position of the anticodon. This tRNA must translate both AUG and AUA as methionine. By constructing an in vitro translation system from bovine liver mitochondria, we examined the decoding properties of the native mt tRNAMet carrying f5C in the anticodon compared to a transcript that lacks the modification. The native mt Met-tRNA could recognize both AUA and AUG codons as Met, but the corresponding synthetic tRNAMet lacking f5C (anticodon CAU), recognized only the AUG codon in both the codon-dependent ribosomal binding and in vitro translation assays. Furthermore, the Escherichia coli elongator tRNAMetm with the anticodon ac4CAU (ac4C-4-acetylcytidine) and the bovine cytoplasmic initiator tRNAMet (anticodon CAU) translated only the AUG codon for Met on mt ribosome. The codon recognition patterns of these tRNAs were the same on E. coli ribosomes. These results demonstrate that the f5C modification in mt tRNAMet plays a crucial role in decoding the nonuniversal AUA codon as Met, and that the genetic code variation is compensated by a change in the tRNA anticodon, not by a change in the ribosome. Base pairing models of f5C-G and f5C-A based on the chemical properties of f5C are presented.",

author = "Chie Takemoto and Spremulli, {Linda L.} and Benkowski, {Lisa A.} and Takuya Ueda and Takashi Yokogawa and Kimitsuna Watanabe",

note = "Funding Information: JSPS Research Fellowship for Young Scientists (08004075 to C.T.); Grants-in-aid for Scientific Research on Priority Areas from the Ministry of Education, Science, Sports and Culture of Japan (1130802 and 14035206 to K.W.); the Human Frontier Science Program Organization (to K. W.); National Institutes of Health (GM32734 to L.L.S.). Funding for open access charge: Human Frontier Science Program Young Investigator Grant (RGY0067/2007 to C.T.).",

year = "2009",

doi = "10.1093/nar/gkp001",

language = "English",

volume = "37",

pages = "1616--1627",

journal = "Nucleic Acids Research",

issn = "0305-1048",

publisher = "Oxford University Press",

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T1 - Unconventional decoding of the AUA codon as methionine by mitochondrial tRNAMet with the anticodon f5CAU as revealed with a mitochondrial in vitro translation system

AU - Takemoto, Chie

AU - Spremulli, Linda L.

AU - Benkowski, Lisa A.

AU - Ueda, Takuya

AU - Yokogawa, Takashi

AU - Watanabe, Kimitsuna

N1 - Funding Information: JSPS Research Fellowship for Young Scientists (08004075 to C.T.); Grants-in-aid for Scientific Research on Priority Areas from the Ministry of Education, Science, Sports and Culture of Japan (1130802 and 14035206 to K.W.); the Human Frontier Science Program Organization (to K. W.); National Institutes of Health (GM32734 to L.L.S.). Funding for open access charge: Human Frontier Science Program Young Investigator Grant (RGY0067/2007 to C.T.).

PY - 2009

Y1 - 2009

N2 - Mitochondrial (mt) tRNAMet has the unusual modified nucleotide 5-formylcytidine (f5C) in the first position of the anticodon. This tRNA must translate both AUG and AUA as methionine. By constructing an in vitro translation system from bovine liver mitochondria, we examined the decoding properties of the native mt tRNAMet carrying f5C in the anticodon compared to a transcript that lacks the modification. The native mt Met-tRNA could recognize both AUA and AUG codons as Met, but the corresponding synthetic tRNAMet lacking f5C (anticodon CAU), recognized only the AUG codon in both the codon-dependent ribosomal binding and in vitro translation assays. Furthermore, the Escherichia coli elongator tRNAMetm with the anticodon ac4CAU (ac4C-4-acetylcytidine) and the bovine cytoplasmic initiator tRNAMet (anticodon CAU) translated only the AUG codon for Met on mt ribosome. The codon recognition patterns of these tRNAs were the same on E. coli ribosomes. These results demonstrate that the f5C modification in mt tRNAMet plays a crucial role in decoding the nonuniversal AUA codon as Met, and that the genetic code variation is compensated by a change in the tRNA anticodon, not by a change in the ribosome. Base pairing models of f5C-G and f5C-A based on the chemical properties of f5C are presented.

AB - Mitochondrial (mt) tRNAMet has the unusual modified nucleotide 5-formylcytidine (f5C) in the first position of the anticodon. This tRNA must translate both AUG and AUA as methionine. By constructing an in vitro translation system from bovine liver mitochondria, we examined the decoding properties of the native mt tRNAMet carrying f5C in the anticodon compared to a transcript that lacks the modification. The native mt Met-tRNA could recognize both AUA and AUG codons as Met, but the corresponding synthetic tRNAMet lacking f5C (anticodon CAU), recognized only the AUG codon in both the codon-dependent ribosomal binding and in vitro translation assays. Furthermore, the Escherichia coli elongator tRNAMetm with the anticodon ac4CAU (ac4C-4-acetylcytidine) and the bovine cytoplasmic initiator tRNAMet (anticodon CAU) translated only the AUG codon for Met on mt ribosome. The codon recognition patterns of these tRNAs were the same on E. coli ribosomes. These results demonstrate that the f5C modification in mt tRNAMet plays a crucial role in decoding the nonuniversal AUA codon as Met, and that the genetic code variation is compensated by a change in the tRNA anticodon, not by a change in the ribosome. Base pairing models of f5C-G and f5C-A based on the chemical properties of f5C are presented.

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JO - Nucleic Acids Research

JF - Nucleic Acids Research

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ER -

Unconventional decoding of the AUA codon as methionine by mitochondrial tRNA^Met with the anticodon f⁵CAU as revealed with a mitochondrial in vitro translation system

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