ABSTRACT
Telomeres are nucleoprotein complexes that protect the chromosome-ends from eliciting DNA repair while ensuring their complete duplication. Pot1 is a subunit of telomere capping complex that binds to the G-rich overhang and inhibits the activation of DNA damage checkpoints. In this study, we explore new functions of fission yeast Pot1 by using a pot1-1 temperature sensitive mutant. We show that pot1 inactivation impairs telomere DNA replication resulting in the accumulation of ssDNA leading to the complete loss of telomeric DNA. Recruitment of Stn1 to telomeres, an auxiliary factor of DNA lagging strand synthesis, is reduced in pot1-1 mutants and overexpression of Stn1 rescues loss of telomeres and cell viability at restrictive temperature. We propose that Pot1 plays a crucial function in telomere DNA replication by recruiting Stn1-Ten1 and Polα-primase complex to telomeres via Tpz1, thus promoting lagging-strand DNA synthesis at stalled replication forks.
Subject(s)
Chromosomes, Fungal , DNA Replication , Schizosaccharomyces pombe Proteins , Schizosaccharomyces , Telomere , DNA-Binding Proteins/metabolism , Schizosaccharomyces/metabolism , Schizosaccharomyces pombe Proteins/metabolism , Shelterin Complex , Telomere/metabolism , Telomere-Binding Proteins/metabolism , Chromosomes, Fungal/metabolismABSTRACT
Efficient replication of terminal DNA is crucial to maintain telomere stability. In fission yeast, Taz1 and the Stn1-Ten1 (ST) complex play prominent roles in DNA-ends replication. However, their function remains elusive. Here, we have analyzed genome-wide replication and show that ST does not affect genome-wide replication but is crucial for the efficient replication of a subtelomeric region called STE3-2. We further show that, when ST function is compromised, a homologous recombination (HR)-based fork restart mechanism becomes necessary for STE3-2 stability. While both Taz1 and Stn1 bind to STE3-2, we find that the STE3-2 replication function of ST is independent of Taz1 but relies on its association with the shelterin proteins Pot1-Tpz1-Poz1. Finally, we demonstrate that the firing of an origin normally inhibited by Rif1 can circumvent the replication defect of subtelomeres when ST function is compromised. Our results help illuminate why fission yeast telomeres are terminal fragile sites.