Reanalysis of mtDNA mutations of human primordial germ cells (PGCs) reveals NUMT contamination and suggests that selection in PGCs may be positive

Abstract

The resilience of the mitochondrial genome (mtDNA) to a high mutational pressure depends, in part, on negative purifying selection against detrimental mutations in the germline. Floros et al. reported a sharp increase in the synonymity of mtDNA mutations between early and late- stage primordial germ cells (PGCs), concomitant with a shift from glycolytic to oxidative metabolism, as evidenced by comparing data from pooled samples of early vs late PGCs. They thus asserted that this metabolic shift exposed deleterious mtDNA mutations to negative selection. We re- analyzed Floros data to resolve a perceived inconsistency of the pattern of synonymity change and discovered a significant contamination of pooled PGC mutations with nuclear sequence derived from mtDNA (NUMT). We determined that contamination was caused by co- amplification of the NUMT sequence by cross-specific PCR primers. Importantly, when we removed NUMT-derived sequence variants from pooled PGC data, the evidence of purifying selection in late PGCs was abolished. We then turned to mutations of single PGCs, also from the same group, for which no synonymity analysis was reported. We found no evidence of NUMT contamination of single PGCs mutations. This is consistent with the use of a different set of PCR primers which are unable to amplify NUMT since they were positioned outside the NUMT sequence. Importantly, we further demonstrated that single PGC mutations show a significant decrease of synonymity with increased mutant fraction. This observation is incompatible with predominantly purifying selection of mtDNA mutations in PGCs at these mutant fractions and suggests that selection may be predominantly positive.