rDNA copy number variation and methylation from birth to sexual maturity

Ribosomal DNA transcription is essential for ribosome biogenesis and the production of proteins. Using a combination of droplet digital PCR and deep bisulfite sequencing, we have quantified both the absolute number as well as the methylation level of individual rDNA transcription units (TU) in blood samples of 139 young healthy individuals and 141 sex- and age-matched individuals with unsolved syndromal developmental delay (DD), ranging from 0.02 to 18.4 years in age. There were no between-group differences in average promoter methylation, absolute copy number (CN), extreme CN, and hypomethylated (0-10%) presumably active CN. This largely excludes rDNA CN as a modulating factor in DD. The absolute CN in all 280 samples was 423.7 ± 108.4 (median 410, range 153 to 1,000) and the active CN was 175.0 ± 36.4 (median 174, range 70 to 376). Similar to adults, the absolute CN did not change from birth to sexual maturity but was strongly (Pearson ρ = 0.64, P < 0.001) correlated with promoter methylation. In contrast to adults, there was no significant correlation between age and promoter methylation and no age-related loss of active copies from birth to < 20 years. The number of completely unmethylated copies even significantly (Pearson ρ = 0.15; P = 0.01) increased during childhood and adolescence. Our results suggest that rDNA promoter methylation and the age-related loss of active rDNA TU, which are a hallmark of the aging process, start only after reaching sexual maturity.