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• Analysis in yeast suggests that POLG p.G268A is neither a pathological mutation nor a neutral SNP
  Enrico Baruffini
  Published on: 27 April 2016
• POLG p.G268A and p.G517V are not pathogenic mutations
  Conrad Smith
  Published on: 27 April 2016

• Published on: 27 April 2016

  Analysis in yeast suggests that POLG p.G268A is neither a pathological mutation nor a neutral SNP

  • Enrico Baruffini, Post doc researcher

  It was with great interest that I read the study by Tang and co- authors [1], in which they discovered twenty-five novel mutations in DNA polymerase gamma. Based on the presence of p.G268A substitution in heterozygosis in 19 subjects from a cohort of 2697 unrelated patients, they proposed to reclassify this mutation as a neutral polymorphism or a polymorphic modifier rather than a pathological mutation. Smith and co- auth...

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  It was with great interest that I read the study by Tang and co- authors [1], in which they discovered twenty-five novel mutations in DNA polymerase gamma. Based on the presence of p.G268A substitution in heterozygosis in 19 subjects from a cohort of 2697 unrelated patients, they proposed to reclassify this mutation as a neutral polymorphism or a polymorphic modifier rather than a pathological mutation. Smith and co- authors [2] confirmed that POLG G268A is not pathogenic but represent neutral polymorphism on the basis of the presence of this mutations in three healthy subjects and on the basis of the frequencies of the allele p.G268A in their patient cohort, which were similar to those found in a control population of European origin by the NHLBI exome sequencing project [3].

  I and co-authors previously demonstrated that in yeast the mip1G224A allele, corresponding to the human p.G268A substitution, was associated with a 2.2-fold increase in petite frequency, which is the frequency of mutants with large deletions and/or loss of mtDNA [4]. A further analysis made in our laboratory on yeast mip1 mutations corresponding to human substitutions classified as neutral SNPs [5] conserved from yeasts to mammals (E. Baruffini and T. Lodi, unpublished results), suggested the existence of a class of substitutions, that also includes G224A, which are not neutral. These mutations caused a 1.5 to 2.5-fold increase in petite frequency, which was lower compared to the petite frequency caused by mutations recognized as pathological, for which at least a 10-fold increase in petite frequency or the total loss of mtDNA were observed in our laboratory [4, 6-9]. This class includes also the mip1E900G and mip1Q766C alleles, corresponding to the human p.E1143G and p.R964C substitutions, respectively. The former (2-fold increase in petite frequency) is a substitution classified in humans as a phenotypic modifier that can modulate disease mutations [10, 11]. The latter (2-fold increase) seems to be pathological in humans when in heterozygosis with p.A862T [8,12] or with p.A962T [1] but not pathological in homozygosis [13]; however it is associated with mitochondrial toxicity susceptibility to stavudine both in humans [13] and in yeast [14] and the mutant DNA polymerase gamma showed an impaired polymerase activity [15].

  Conclusion: analysis in yeast suggests that p.G268A is neither a pathological mutation nor a neutral SNP. It can be classified, together with R964C and E1143G, as a "phenotypic modifier", as suggested by Copeland [10], as an "unclassified variant", as suggested by Tang and co- authors [1] or as an "Ecogenetic Single Nucleotide Variant (ESNV)", as suggested by Saneto and Naviaux for genetic variations whose phenotypic expression is determined by interaction with genetic, epigenetic and environmental factors [16]. In general, these observations suggest that the validation of the pathological significance of novel mutations, whose discovery is exponentially increasing thanks to the next generation sequencing, can take advantage, when possible, of experimental analysis in model systems to confirm or disavow their role in the disease.

  References 1. Tang S, Wang J, Lee NC, Milone M, Halberg MC, Schmitt ES, Craigen WJ, Zhang W, Wong LJ. Mitochondrial DNA polymerase gamma mutations: an ever expanding molecular and clinical spectrum. J Med Genet 2011;48:669-681. 2. Smith C. POLG p.G268A and p.G517V are not pathogenic mutations. J MedGenet 2011; eletter 3. NHLBI exome sequencing project, https://esp.gs.washington.edu/drupal/ 4. Baruffini E, Lodi T, Dallabona C, Puglisi A, Zeviani M, Ferrero I. Genetic and chemical rescue of the Saccharomyces cerevisiae phenotype induced by mitochondrial DNA polymerase mutations associated with progressive external ophthalmoplegia in humans. Hum Mol Genet 2006;15:2846 -2855. 5. dbSNP, http://www.ncbi.nlm.nih.gov/snp/ 6. Baruffini E, Ferrero I, Foury F. Mitochondrial DNA defects in Saccharomyces cerevisiae caused by functional interactions between DNA polymerase gamma mutations associated with disease in human. Biochim Biophys Acta 2007;1772:1225-1235. 7. Spinazzola A, Invernizzi F, Carrara F, Lamantea E, Donati A, Dirocco M, Giordano I, Meznaric-Petrusa M, Baruffini E, Ferrero I, Zeviani M. Clinical and molecular features of mitochondrial DNA depletion syndromes. J Inherit Metab Dis 2009;32:143-158. 8. Stricker S, Pr?ss H, Horvath R, Baruffini E, Lodi T, Siebert E, Endres M, Zschenderlein R, Meisel A.J. A variable neurodegenerative phenotype with polymerase gamma mutation. J Neurol Neurosurg Psychiatry 2009;80:1181 -1182. 9. Baruffini E, Horvath R, Dallabona C, Czermin B, Lamantea E, Bindoff L, Invernizzi F, Ferrero I, Zeviani M, Lodi T. Predicting the contribution of novel POLG mutations to human disease through analysis in yeast model. Mitochondrion 2011;11:182-190. 10. Human DNA Polymerase Gamma Mutation Database, http://tools.niehs.nih.gov/polg/ 11. Chan SS, Longley MJ, Copeland WC. Modulation of the W748S mutation in DNA polymerase gamma by the E1143G polymorphismin mitochondrial disorders. Hum Mol Genet 2006;15:3473-3483. 12. Wong LJ, Naviaux RK, Brunetti-Pierri N, Zhang Q, Schmitt ES, Truong C, Milone M, Cohen BH, Wical B, Ganesh J, Basinger AA, Burton BK, Swoboda K, Gilbert DL, Vanderver A, Saneto RP, Maranda B, Arnold G, Abdenur JE, Waters PJ, Copeland WC. Molecular and clinical genetics of mitochondrial diseases due to POLG mutations. Hum Mutat 2008;29:E150-E172. 13. Yamanaka H, Gatanaga H, Kosalaraksa P, Matsuoka-Aizawa S, Takahashi T, Kimura S, Oka S. Novel mutation of human DNA polymerase gamma associated with mitochondrial toxicity induced by anti-HIV treatment. J Infect Dis 2007;195:1419-1425. 14. Baruffini E, Lodi T. Construction and validation of a yeast model system for studying in vivo the susceptibility to nucleoside analogues of DNA polymerase gamma allelic variants. Mitochondrion 2010;10:183-187. 15. Bailey CM, Kasiviswanathan R, Copeland WC, Anderson KS. R964C mutation of DNA polymerase gamma imparts increased stavudine toxicity by decreasing nucleoside analog discrimination and impairing polymerase activity. Antimicrob Agents Chemother 2009;53:2610-2612. 16. Saneto RP, Naviaux RK. Polymerase gamma disease through the ages. Dev Disabil Res Rev 2010;16:163-174.

  Conflict of Interest:

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  Conflict of Interest:
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• Published on: 27 April 2016

  POLG p.G268A and p.G517V are not pathogenic mutations

  • Conrad Smith, Clinical Scientist
  • Other Contributors:
    • Carl Fratter, Julie Evans, Anneke Seller, Michael Parker, Sharon Pagdin, Mary Sweeney, Garry Brown and Joanna Poulton

  In their recently published paper describing mutations in mitochondrial DNA polymerase gamma, Tang et al.(1) propose that the POLG p.G268A (c.803G>C) and p.G517V (c.1550G>T) variants which have previously been reported as pathogenic mutations should be considered as unclassified variants that may represent rare neutral polymorphisms or polymorphic modifiers.

  We have also identified these variants in our...

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  In their recently published paper describing mutations in mitochondrial DNA polymerase gamma, Tang et al.(1) propose that the POLG p.G268A (c.803G>C) and p.G517V (c.1550G>T) variants which have previously been reported as pathogenic mutations should be considered as unclassified variants that may represent rare neutral polymorphisms or polymorphic modifiers.

  We have also identified these variants in our own cohort of 627 patients that were referred with a suspected disorder of mitochondrial DNA maintenance. We detected p.G268A in 9 individuals (allele frequency 0.72%) of ages from 3 months to 63 years. Symptoms were variable ranging from severe early onset mitochondrial DNA depletion syndrome to milder late onset neuropathy/ataxia/ophthalmoplegia. In all 9 cases the variant occurred as a heterozygous change with no other pathogenic POLG mutation, suggesting that p.G268A is not a recessive mutation. Parental samples were available in 3 cases, and for each of these we found the variant in an unaffected parent, suggesting that p.G268A is very unlikely to be a dominant mutation. Furthermore, one of the 9 index cases was later found to be compound heterozygous for pathogenic mutations in another gene associated with mtDNA maintenance disorders, DGUOK. In another case the proband had a similarly affected sibling who did not have p.G268A.

  Similarly, we have identified p.G517V 8 times (allele frequency 0.64%), always as a heterozygous change with no other pathogenic POLG mutation. The age of the individuals ranged from 1-81 yrs and the phenotype varied from infantile epilepsy/failure to thrive to adult onset ophthalmoplegia/ataxia/myopathy. The unaffected parent of 1 of these individuals was also heterozygous for p.G517V. A second affected individual was later found to have pathogenic mutations in RRM2B.

  Furthermore, the allele frequencies of p.G268A and p.G517V in our patient cohort are remarkably similar to those found in a large control population of European origin (2700 alleles) by the NHLBI exome sequencing project (2); 0.59% and 0.86% respectively.

  Therefore, our data, taken together with that for Tang et al. and the NHLBI exome sequencing project, confirm that POLG p.G268A and p.G517V are not pathogenic and are highly likely to represent neutral polymorphisms.

  References. (1) Tang S, Wang J, Lee NC, Milone M, Halberg MC, Schmitt ES, Craigen WJ, Zhang W, Wong LJ. Mitochondrial DNA polymerase gamma mutations: an ever expanding molecular and clinical spectrum. J Med Genet 2011;48:669-681. (2) https://esp.gs.washington.edu/drupal/

  Conflict of Interest:

  None declared

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  Conflict of Interest:
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