A human mitochondrial poly(A) polymerase mutation reveals the complexities of post-transcriptional mitochondrial gene expression

William C. Wilson, Hue Tran Hornig-Do, Francesco Bruni, Jeong H.o. Chang, Alexis A. Jourdain, Jean Claude Martinou, Maria Falkenberg, Henrik Spåhr, Nils Göran Larsson, Richard J. Lewis, Lorraine Hewitt, Arnaud Baslé, Harold E. Cross, Liang Tong, Robert R. Lebel, Andrew H. Crosby, Zofia M.A. Chrzanowska-Lightowlers, Robert N. Lightowlers

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38 Scopus citations

Abstract

The p.N478D missense mutation in human mitochondrial poly(A) polymerase (mtPAP) has previously been implicated in a form of spastic ataxia with optic atrophy. In this study, we have investigated fibroblast cell lines established from family members. The homozygous mutation resulted in the loss of polyadenylation of all mitochondrial transcripts assessed; however, oligoadenylation was retained. Interestingly, this had differential effects on transcript stability that were dependent on the particular species of transcript. These changes were accompanied by a severe loss of oxidative phosphorylation complexes I and IV, and perturbation of de novo mitochondrial protein synthesis. Decreases in transcript polyadenylation and in respiratory chain complexes were effectively rescued by overexpression of wild-type mtPAP. Both mutated and wild-type mtPAP localized to the mitochondrial RNA-processing granules thereby eliminating mislocalization as a cause of defective polyadenylation. In vitro polyadenylation assays revealed severely compromised activity by the mutated protein, which generated only short oligo(A) extensions on RNA substrates, irrespective of RNA secondary structure. The addition of LRPPRC/SLIRP, a mitochondrial RNA-binding complex, enhanced activity of the wild-type mtPAP resulting in increased overall tail length. The LRPPRC/SLIRP effect although present was less marked with mutated mtPAP, independent of RNA secondary structure. We conclude that (i) the polymerase activity of mtPAP can be modulated by the presence of LRPPRC/SLIRP, (ii) N478D mtPAP mutation decreases polymerase activity and (iii) the alteration in poly(A) length is sufficient to cause dysregulation of post-transcriptional expression and the pathogenic lack of respiratory chain complexes.

Original languageEnglish (US)
Pages (from-to)6345-6355
Number of pages11
JournalHuman molecular genetics
Volume23
Issue number23
DOIs
StatePublished - Dec 1 2014

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Genetics(clinical)

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    Wilson, W. C., Hornig-Do, H. T., Bruni, F., Chang, J. H. O., Jourdain, A. A., Martinou, J. C., Falkenberg, M., Spåhr, H., Larsson, N. G., Lewis, R. J., Hewitt, L., Baslé, A., Cross, H. E., Tong, L., Lebel, R. R., Crosby, A. H., Chrzanowska-Lightowlers, Z. M. A., & Lightowlers, R. N. (2014). A human mitochondrial poly(A) polymerase mutation reveals the complexities of post-transcriptional mitochondrial gene expression. Human molecular genetics, 23(23), 6345-6355. https://doi.org/10.1093/hmg/ddu352