Loss of Cdc13 causes genome instability by a deficiency in replication-dependent telomere capping.

In budding yeast, Cdc13, Stn1, and Ten1 form the telomere-binding heterotrimer CST complex. Here we investigate the role of Cdc13/CST in maintaining genome stability by using a Chr VII disome system that can generate recombinants, chromosome loss, and enigmatic unstable chromosomes. In cells expressing a temperature sensitive CDC13 allele, cdc13F684S, unstable chromosomes frequently arise from problems in or near a telomere.

A Slowed Cell Cycle Stabilizes the Budding Yeast Genome.

During cell division, aberrant DNA structures are detected by regulators called checkpoints that slow division to allow error correction. In addition to checkpoint-induced delay, it is widely assumed, though rarely shown, that merely slowing the cell cycle might allow more time for error detection and correction, thus resulting in a more stable genome. Fidelity by a slowed cell cycle might be independent of checkpoints.

Ontogeny of Unstable Chromosomes Generated by Telomere Error in Budding Yeast.

DNA replication errors at certain sites in the genome initiate chromosome instability that ultimately leads to stable genomic rearrangements. Where instability begins is often unclear. And, early instability may form unstable chromosome intermediates whose transient nature also hinders mechanistic understanding.

Nifty Alleles, a Plethora of Interactions, and Imagination Advance Understanding of Smc5/6's Roles with Chromosomes.

The Smc5/6 complex is involved in DNA replication and repair, but why it is essential is less clear. In this issue of Molecular Cell, Menolfi et al. (2015) use cell cycle-specific alleles to link the essential function of Smc5/6 to resolving replication problems that persist into G2.

Mec1 and Tel1: an arresting dance of resection.

At sites of damage DNA is resected to enable the optimal form of repair, directed by homology. But how does the cell regulate resection while coordinating with the cell cycle? In this issue of , Clerici define missing links between DNA resection and cell cycle arrest during repair, showing that Mec1 can inhibit resection to subsequently activate arrest by Tel1.

DNA replication: failures and inverted fusions.

DNA replication normally follows the rules passed down from Watson and Crick: the chromosome duplicates as dictated by its antiparallel strands, base-pairing and leading and lagging strand differences. Real-life replication is more complicated, fraught with perils posed by chromosome damage for one, and by transcription of genes and by other perils that disrupt progress of the DNA replication machinery. Understanding the replication fork, including DNA structures, associated replisome and its regulators, is key to understanding how cells overcome perils and minimize error.

Checkpoint genes and Exo1 regulate nearby inverted repeat fusions that form dicentric chromosomes in Saccharomyces cerevisiae.

Genomic rearrangements are common, occur by largely unknown mechanisms, and can lead to human diseases. We previously demonstrated that some genome rearrangements occur in budding yeast through the fusion of two DNA sequences that contain limited sequence homology, lie in inverted orientation, and are within 5 kb of one another. This inverted repeat fusion reaction forms dicentric chromosomes, which are well-known intermediates to additional rearrangements.

Choreography of the 9-1-1 checkpoint complex: DDK puts a check on the checkpoints.

Checkpoint proteins respond to DNA damage by halting the cell cycle until the damage is repaired. In this issue of Molecular Cell, Furuya et al. (2010) provide evidence that checkpoint proteins need to be removed from sites of damage in order to properly repair it.

The role of replication bypass pathways in dicentric chromosome formation in budding yeast.

Gross chromosomal rearrangements (GCRs) are large scale changes to chromosome structure and can lead to human disease. We previously showed in Saccharomyces cerevisiae that nearby inverted repeat sequences (∼20-200 bp of homology, separated by ∼1-5 kb) frequently fuse to form unstable dicentric and acentric chromosomes. Here we analyzed inverted repeat fusion in mutants of three sets of genes.

Fusion of nearby inverted repeats by a replication-based mechanism leads to formation of dicentric and acentric chromosomes that cause genome instability in budding yeast.

Large-scale changes (gross chromosomal rearrangements [GCRs]) are common in genomes, and are often associated with pathological disorders. We report here that a specific pair of nearby inverted repeats in budding yeast fuse to form a dicentric chromosome intermediate, which then rearranges to form a translocation and other GCRs. We next show that fusion of nearby inverted repeats is general; we found that many nearby inverted repeats that are present in the yeast genome also fuse, as does a pair of synthetically constructed inverted repeats.

tRNA traffic meets a cell-cycle checkpoint.

The molecular pathways linking DNA-damage checkpoint proteins to cell-cycle progression remain largely unresolved. Findings by Ghavidel et al. (2007) reported in this issue suggest that tRNA trafficking and the transcription factor Gcn4 are key intermediates in the process by which yeast cells detect DNA damage and delay cell-cycle progression at the G1 to S phase transition.

Cycles of chromosome instability are associated with a fragile site and are increased by defects in DNA replication and checkpoint controls in yeast.

We report here that a normal budding yeast chromosome (ChrVII) can undergo remarkable cycles of chromosome instability. The events associated with cycles of instability caused a distinctive "sectoring" of colonies on selective agar plates. We found that instability initiated at any of several sites on ChrVII, and was sharply increased by the disruption of DNA replication or by defects in checkpoint controls.

Do telomeres ask checkpoint proteins: "gimme shelter-in"?

Telomeres are complicated structures designed to allow one thing and avoid another. They allow replication of chromosome ends, an issue mostly about telomerase, which we seem to understand (though details of its regulation are works in progress). Telomeres must also avoid being detected as DNA breaks.

A telomeric repeat sequence adjacent to a DNA double-stranded break produces an anticheckpoint.

Telomeres are complex structures that serve to protect chromosome ends. Here we provide evidence that in Saccharomyces cerevisiae telomeres may contain an anticheckpoint activity that prevents chromosome ends from signaling cell cycle arrest. We found that an internal tract of telomeric repeats inhibited DNA damage checkpoint signaling from adjacent double-strand breaks (DSBs); cell cycle arrest lasted 8-12 h from a normal DSB, whereas it lasted only 1-2 h from a DSB adjacent to a telomeric repeat.

Mec1 and Rad53 inhibit formation of single-stranded DNA at telomeres of Saccharomyces cerevisiae cdc13-1 mutants.

Here we examine the roles of budding-yeast checkpoint proteins in regulating degradation of dsDNA to ssDNA at unprotected telomeres (in Cdc13 telomere-binding protein defective strains). We find that Rad17, Mec3, as well as Rad24, members of the putative checkpoint clamp loader (Rad24) and sliding clamp (Rad17, Mec3) complexes, are important for promoting degradation of dsDNA in and near telomere repeats. We find that Mec1, Rad53, as well as Rad9, have the opposite role: they inhibit degradation.

Toward maintaining the genome: DNA damage and replication checkpoints.

DNA checkpoints play a significant role in cancer pathology, perhaps most notably in maintaining genome stability. This review summarizes the genetic and molecular mechanisms of checkpoint activation in response to DNA damage. The major checkpoint proteins common to all eukaryotes are identified and discussed, together with how the checkpoint proteins interact to induce arrest within each cell cycle phase.