Somatic mutations found in the healthy blood compartment of a 115-yr-old woman demonstrate oligoclonal hematopoiesis

Abstract

The somatic mutation burden in healthy white blood cells (WBCs) is not well known. Based on deep whole-genome sequencing, we estimate that approximately 450 somatic mutations accumulated in the nonrepetitive genome within the healthy blood compartment of a 115-yr-old woman. The detected mutations appear to have been harmless passenger mutations: They were enriched in noncoding, AT-rich regions that are not evolutionarily conserved, and they were depleted for genomic elements where mutations might have favorable or adverse effects on cellular fitness, such as regions with actively transcribed genes. The distribution of variant allele frequencies of these mutations suggests that the majority of the peripheral white blood cells were offspring of two related hematopoietic stem cell (HSC) clones. Moreover, telomere lengths of the WBCs were significantly shorter than telomere lengths from other tissues. Together, this suggests that the finite lifespan of HSCs, rather than somatic mutation effects, may lead to hematopoietic clonal evolution at extreme ages.

Figure 1.

Mean telomere length of W115 tissues. DNA from W115 tissues was isolated using the Qiagen DNA isolation kit and the Promega Wizard kit. Both DNA isolates were measured twice for telomere length (T/S ratio). (*) Blood DNA was isolated only with the Promega Wizard kit.

Figure 2.

Somatic variant detection pipeline SNV and indel detection. Whole-genome sequences of W115 blood and brain tissues were mapped to hg19. Variants were called using both GATK and SAMtools. The SNVs that overlapped between the two genotyping algorithms were considered most trustworthy and were used for further analysis with high stringency (HS-SNV) and low stringency (LS-SNV) filters. Indels were passed through a high stringency (HS-indel) and low stringency (LS-indel) read-depth filter. Candidate somatic SNVs and indels were confirmed by validation with Ion Torrent PGM sequencing and/or Sanger sequencing. The number of confirmed SNVs and indels was extrapolated to account for those not validated. The number of SNVs was also extrapolated to the whole genome, whereas for indel detection the whole genome was assessed.

Figure 3.

Presence of single nucleotide mutations detected in blood in other W115 tissues. Box plot of the VAF values for the 214 confirmed somatic mutations detected in blood for a variety of other tissues. On each box, the central mark is the median VAF; the edges of the box are the 25th and 75th percentiles, the whiskers extend to the most extreme data points not considered outliers, and outliers are plotted individually as red crosses.

Figure 4.

Enrichment and depletion of somatic mutations in functional genomic elements tracked by ENCODE. Enrichment depletion analysis: (y-axis) −log P-values indicate the enrichment or depletion of somatic mutations (blue bar), dbSNP variants (green bar), and ClinVar (red bar) variants for each functional element tracked by ENCODE. The lowest possible P-value = 1 × 10⁻⁶, or –log P-value = 6. Comparative analysis: (Black stars) Variant set is significantly enriched (star above bars) or depleted (star below bars) relative to ClinVar variants, P < 1 × 10⁻⁶. Pink star (BU ORChID track only) Variant set is significantly enriched relative to dbSNP variants. Comparisons of other variant sets did not yield significant differences. (‡) Tracks are not specific for cell lines. (*) Track not available for H1 hESC, track for H7 human embryonic stem cell line used instead.

Figure 5.

Gaussian fit for VAF values of 201 confirmed somatic mutations. (Gray) Histogram of VAF values of 201 mutations confirmed by Ion Torrent PGM; (red) Gaussian fit of clone A; (green) Gaussian fit of clone B; (blue) Gaussian fit of background mutations; (black) resultant mixture of Gaussians.