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Characterization and localization of mitochondrial DNA-encoded tRNAs and nuclear DNA-encoded tRNAs in the sea anemone Metridium senile

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Abstract

The mitochondrial (mt) genome of the sea anemone Metridium senile contains genes for only two transfer RNAs (tRNAs), tRNAf-Met and tRNATrp. Experiments were conducted to seek evidence for the occurrence of functional tRNAs corresponding to these genes and for the participation of nuclear DNA-encoded tRNAs in mt-protein synthesis. RNA sequences corresponding to the two mt-tRNA genes were located in mitochondria and it was shown that 3′-CC (and possibly A, but no other nucleotide) is added post-transcriptionally to the 3′ end of at least 50 % of mt-tRNAf-Met molecules and to a small fraction of the mt-tRNATrp molecules. Using specific oligonucleotide primers based on expected nuclear DNA-encoded tRNAs in a series of RACE experiments, we located the nuclear genes for tRNAGln, tRNAIle, tRNAi-Met, tRNAVal and tRNAThr. Data from Northern blot analyses indicated that mtDNA-encoded tRNAf-Met is limited to mitochondria but that nuclear DNA-encoded tRNAVal and tRNAi-Met are present in the cytoplasm and in mitochondria. These data provide direct evidence that in M. senile, mature, functional tRNAs are transcribed from the mtDNA-encoded tRNAf-Met and tRNATrp genes, and are consistent with the interpretation that both nuclear DNA-encoded tRNAVal and tRNAi-Met are utilized in mitochondrial and cytosolic protein synthesis.

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Acknowledgments

We thank Michael Bastiani, Brenda L. Bass, Raymond F. Gesteland, David P. Goldenberg for discussions during the course of this work, and Robert Schackmann for oligonucleotides (partially subsidized by National Institutes of Health Grant CA-42014) This work was supported by National Institutes of Health Grant GM-18375 and funds from the University of Utah, and was submitted by C. T. Beagley in partial fulfillment of the requirements for the degree of Doctor of Philosophy (Biology), College of Science, University of Utah.

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  1. Department of Biology, University of Utah, Salt Lake City, UT, 84121, USA

    C. Timothy Beagley & David R. Wolstenholme

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  1. C. Timothy Beagley
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Correspondence to C. Timothy Beagley.

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Communicated by S. Hohmann.

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Beagley, C.T., Wolstenholme, D.R. Characterization and localization of mitochondrial DNA-encoded tRNAs and nuclear DNA-encoded tRNAs in the sea anemone Metridium senile . Curr Genet 59, 139–152 (2013). https://doi.org/10.1007/s00294-013-0395-9

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