Telomere Protection and Maintenance in Arabidopsis thaliana

Telomeres are the physical ends of linear chromosomes in eukaryotes. Telomeres not only protect chromosome ends from being recognized as double-strand breaks but also maintain the chromosome terminal sequences. These processes involve a number of telomere-related proteins. A major challenge in the field is to elucidate the full constitution of telomere-associated proteins and to understand how different protein complexes are regulated at chromosome termini. Here, I report the identification and characterization of STN1 (Suppressor of cdc thirteen, 1), CTC1 (Conserved Telomere maintenance Component 1) and TEN1 (Telomeric pathways in association with Stn1, 1) in Arabidopsis. CTC1/STN1/TEN1 (CST) forms a trimeric complex that specifically associates with telomeres. Loss of any component of the CST induces catastrophic telomere loss, disrupted telomere end architecture, and massive chromosome end-to-end fusions. Thus, CST plays an essential role in chromosome end protection. I also show that CST function at telomeres is independent of a previously characterized capping complex KU70/KU80, and that ATR is responsible for a checkpoint response in plants lacking CTC1/STN1. Additionally, I present data showing that Arabidopsis POT1a (Protection Of Telomere 1, a) has evolved as a telomerase recruitment factor. Unlike POT1 in other eukaryotes which binds and protects ss telomeric DNA, AtPOT1a interacts with telomerase RNA (TER). Based on an evolutionary analysis, we found that the POT1a lineage is under positive selection in the Brassicaceae family in which Arabidopsis belongs. Mutations of two positive selection sites significantly reduce POT1a?s activity in vivo. These data suggest POT1a is under pressure to evolve from a telomeric DNA binding protein to a TER binding protein. I also discovered that POT1a interacts with the novel telomere capping protein CTC1 in vitro and in vivo. Thus, I hypothesize that POT1a acts as a telomerase recruitment factor linking this enzyme to the chromosome termini via interacting with TER and CTC1. Finally, I dissected the functional domains of POT1a and demonstrated that both the N-terminus and the C-terminus of POT1a are required for its function in vivo. In summary, my work has uncovered several new and essential telomereassociated proteins that provide new insight into mechanisms of chromosome end protection and maintenance.