Garrido, C.; Kornobis, E.; Agier, N.; Ilioaia, O.; Fischer, G.; Xu, Z.

Telomerase counteracts the gradual shortening of telomeres that is occurring at each cell division by elongating the telomeric sequences. This process generates a dynamic balance between elongation and shortening, best represented by a steady-state telomere length (TL) distribution. We developed a computational pipeline for detecting telomeric sequences in Saccharomyces cerevisiae genome directly from raw Oxford Nanopore reads. This method provides a complete view of TL distributions and enables broad exploration of TL diversity, as it is directly applicable to any high-throughput single-molecule sequencing dataset without prior DNA preparation. We analyzed the TL distribution in 100 S. cerevisiae strains representing the genetic and ecological diversity of the species. We obtained the first species-level view of TL distributions at individual chromosome extremities and measured their mean value across all strains at 359 {+/-} 129 bp. Our analysis revealed a large diversity in TL distributions across strains, polyploids exhibiting longer telomeres than diploids and haploids. We confirmed that TL distributions are specific to each chromosome extremity and found that the extent of the inter-extremity differences within a strain varied widely across strains. Each strain displays a distinct TL distribution pattern defined by two components: inter-extremity differences in mean TL and within-extremity TL heterogeneity. Differences in inter-extremity mean TL accounted for most of the observed TL variations between strains. Furthermore, we found that inter-extremity differences partly result from the presence of subtelomeric Y\' elements, which are associated with longer telomeres in cis. Interestingly, despite large TL variations, the length of the shortest telomeres remained relatively constant across strains and fluctuated around 138 {+/-} 45 bp, suggesting a selective constraint at the species level. This study reveals the species-wide TL diversity in S. cerevisiae and highlights key factors shaping TL distributions at individual chromosome extremities.