Telomerase, telomeric DNA and associated proteins together represent a complex, finely tuned and functionally conserved mechanism that ensures genome integrity by protecting and maintaining chromosome ends. Changes in its components can threaten an organism's viability. Nevertheless, molecular innovation in telomere maintenance has occurred multiple times during eukaryote evolution, giving rise to species/taxa with unusual telomeric DNA sequences, telomerase components or telomerase-independent telomere maintenance. The central component of telomere maintenance machinery is telomerase RNA (TR) as it templates telomere DNA synthesis, its mutation can change telomere DNA and disrupt its recognition by telomere proteins, thereby leading to collapse of their end-protective and telomerase recruitment functions.
Using a combination of bioinformatic and experimental approaches, we examine a plausible scenario of evolutionary changes in TR underlying telomere transitions.
We identified plants harbouring multiple TR paralogs whose template regions could support the synthesis of diverse telomeres. In our hypothesis, formation of unusual telomeres is associated with the occurrence of TR paralogs that can accumulate mutations, and through their functional redundancy, allow for the adaptive evolution of the other telomere components.
Experimental analyses of telomeres in the examined plants demonstrate evolutionary telomere transitions corresponding to TR paralogs with diverse template regions.This research was supported by the Czech Science Foundation project 20-01331X. VP, and Bal31-NGS data generation was supported by the Czech Science Foundation project 23-06643S. Collaboration with Potato Research Institute, v.v.i., was supported by the European Regional Development Fund Project ‘SINGING PLANT’ (CZ.02.01/0.0/0.0/16_026/0008446).Introduction
Materials and Methods
Plant material and preparation of DNA and RNA
Fluorescence in situ hybridization and microscopy
Analysis of terminal restriction fragments (TRF) and their sensitivity to BAL31 (BAL31-TRF)
Telomere repeat amplification protocol (TRAP) and sequencing of products
Transcriptional contribution of TR paralogs in Fagopyrum
Identification of abundant tandem repeats in genomic data
Identification of TR sequences
Analysis of syntenic relationships among TR loci using the CoGe platform
Results
Computational identification of TR genes and telomere motifs across land plants
Selection of plant species indicating presence of an unusual telomere sequence and validation
Experimental validation in a subset of species
FISH signals with candidate telomere probes
Sensitivity of TRF signals and telomere read counts to BAL31 nuclease digestion
Synthesis of variant telomere repeats and differential contribution of TR paralogs
Origin and fate of TR paralogs
Discussion
Acknowledgements
Competing interests
Author contribution
