Chromosome-specific telomere lengths and the minimal functional telomere revealed by nanopore sequencing.

We developed a method to tag telomeres and measure telomere length by nanopore sequencing in the yeast S. cerevisiae Nanopore allows long-read sequencing through the telomere, through the subtelomere, and into unique chromosomal sequence, enabling assignment of telomere length to a specific chromosome end. We observed chromosome end-specific telomere lengths that were stable over 120 cell divisions. These stable chromosome-specific telomere lengths may be explained by slow clonal variation or may represent a new biological mechanism that maintains equilibrium unique to each chromosome end. We examined the role of RIF1 and TEL1 in telomere length regulation and found that TEL1 is epistatic to RIF1 at most telomeres, consistent with the literature. However, at telomeres that lack subtelomeric Y' sequences, tel1Δ rif1Δ double mutants had a very small, but significant, increase in telomere length compared with the tel1Δ single mutant, suggesting an influence of Y' elements on telomere length regulation. We sequenced telomeres in a telomerase-null mutant (est2Δ) and found the minimal telomere length to be ∼75 bp. In these est2Δ mutants, there were apparent telomere recombination events at individual telomeres before the generation of survivors, and these events were significantly reduced in est2Δ rad52Δ double mutants. The rate of telomere shortening in the absence of telomerase was similar across all chromosome ends at ∼5 bp per generation. This new method gives quantitative, high-resolution telomere length measurement at each individual chromosome end and suggests possible new biological mechanisms regulating telomere length.

Human telomere length is chromosome specific and conserved across individuals.

Short telomeres cause age-related disease and long telomeres predispose to cancer; however, the mechanisms regulating telomere length are unclear. To probe these mechanisms, we developed a nanopore sequencing method, Telomere Profiling, that is easy to implement, precise, and cost effective with broad applications in research and the clinic. We sequenced telomeres from individuals with short telomere syndromes and found similar telomere lengths to the clinical FlowFISH assay. We mapped telomere reads to specific chromosome end and identified both chromosome end-specific and haplotype-specific telomere length distributions. In the T2T HG002 genome, where the average telomere length is 5kb, we found a remarkable 6kb difference in lengths between some telomeres. Further, we found that specific chromosome ends were consistently shorter or longer than the average length across 147 individuals. The presence of conserved chromosome end-specific telomere lengths suggests there are new paradigms in telomere biology that are yet to be explored. Understanding the mechanisms regulating length will allow deeper insights into telomere biology that can lead to new approaches to disease.

Human telomere length is chromosome end-specific and conserved across individuals.

Short telomeres cause age-related disease, and long telomeres contribute to cancer; however, the mechanisms regulating telomere length are unclear. We developed a nanopore-based method, which we call Telomere Profiling, to determine telomere length at nearly single-nucleotide resolution. Mapping telomere reads to chromosome ends showed chromosome end-specific length distributions that could differ by more than six kilobases. Examination of telomere lengths in 147 individuals revealed that certain chromosome ends were consistently longer or shorter. The same rank order was found in newborn cord blood, suggesting that telomere length is determined at birth and that chromosome end-specific telomere length differences are maintained as telomeres shorten with age. Telomere Profiling makes precision investigation of telomere length widely accessible for laboratory, clinical, and drug discovery efforts and will allow deeper insights into telomere biology.