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Using mitoribosomal profiling to investigate human mitochondrial translation

Lookup NU author(s): Dr Fei Gao, Maria Wesolowska, Dr Conor LawlessORCiD, Professor Robert Lightowlers, Professor Zofia Chrzanowska-LightowlersORCiD



This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Background: Gene expression in human mitochondria has various idiosyncratic features. One of these was recently revealed as the unprecedented recruitment of a mitochondrially-encoded tRNA as a structural component of the large mitoribosomal subunit. In porcine particles this is mt-tRNAPhe whilst in humans it is mt-tRNAVal. We have previously shown that when a mutation in mt-tRNAVal causes very low steady state levels, there is preferential recruitment of mt-tRNAPhe. We have investigated whether this altered mitoribosome affects intra-organellar protein synthesis. Methods: By using mitoribosomal profiling we have revealed aspects of mitoribosome behaviour with its template mt-mRNA under both normal conditions as well as those where the mitoribosome has incorporated mt-tRNAPhe. Results: Analysis of the mitoribosome residency on transcripts under control conditions reveals that although mitochondria employ only 22 mt-tRNAs for protein synthesis, the use of non-canonical wobble base pairs at codon position 3 does not cause any measurable difference in mitoribosome occupancy irrespective of the codon. Comparison of the profile of aberrant mt-tRNAPhe containing mitoribosomes with those of controls that integrate mt-tRNAVal revealed that the impaired translation seen in the latter was not due to stalling on triplets encoding either of these amino acids. The alterations in mitoribosome interactions with start codons was not directly attributable to the either the use of non-cognate initiation codons or the presence or absence of 5’ leader sequences, except in the two bicistronic RNA units, RNA7 and RNA14 where the initiation sites are internal. Conclusions: These data report the power of mitoribosomal profiling in helping to understand the subtleties of mammalian mitochondrial protein synthesis. Analysis of profiles from the mutant mt-tRNAVal cell line suggest that despite mt-tRNAPhe being preferred in the porcine mitoribosome, its integration into the human counterpart results in a suboptimal structure that modifies its interaction with mt-mRNAs.

Publication metadata

Author(s): Gao F, Wesolowska M, Agami R, Rooijers K, Loayza-Puch F, Lawless C, Lightowlers RN, Chrzanowska-Lightowlers ZM

Publication type: Article

Publication status: Published

Journal: Wellcome Open Research

Year: 2017

Volume: 2

Print publication date: 11/12/2017

Online publication date: 11/12/2017

Acceptance date: 11/12/2017

Date deposited: 11/12/2017

ISSN (electronic): 2398-502X

Publisher: F1000 Research Limited


DOI: 10.12688/wellcomeopenres.13119.1


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