ABSTRACT
The structural maintenance of chromosome (SMC) proteins are key elements in controlling chromosome dynamics. In eukaryotic cells, three essential SMC complexes have been defined: cohesin, condensin, and the Smc5/6 complex. The latter is essential for DNA damage responses; in its absence both repair and checkpoint responses fail. In fission yeast, the UV-C and ionizing radiation (IR) sensitivity of a specific hypomorphic allele encoding the Smc6 subunit, rad18-74 (renamed smc6-74), is suppressed by mild overexpression of a six-BRCT-domain protein, Brc1. Deletion of brc1 does not result in a hypersensitivity to UV-C or IR, and thus the function of Brc1 relative to the Smc5/6 complex has remained unclear. Here we show that brc1Delta cells are hypersensitive to a range of radiomimetic drugs that share the feature of creating lesions that are an impediment to the completion of DNA replication. Through a genetic analysis of brc1Delta epistasis and by defining genes required for Brc1 to suppress smc6-74, we find that Brc1 functions to promote recombination through a novel postreplication repair pathway and the structure-specific nucleases Slx1 and Mus81. Activation of this pathway through overproduction of Brc1 bypasses a repair defect in smc6-74, reestablishing resolution of lesions by recombination.
Subject(s)
Cell Cycle Proteins/genetics , DNA Damage/genetics , DNA Repair/genetics , Recombination, Genetic/genetics , Schizosaccharomyces pombe Proteins/genetics , Schizosaccharomyces pombe Proteins/metabolism , Schizosaccharomyces/genetics , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Endonucleases/genetics , Endonucleases/metabolism , Epistasis, Genetic , Metronidazole/analogs & derivatives , Metronidazole/toxicity , Schizosaccharomyces/drug effects , Schizosaccharomyces/radiation effects , Ultraviolet RaysABSTRACT
The detection of DNA damage activates DNA repair pathways and checkpoints to allow time for repair. Ultimately, these responses must be coordinated to ensure that cell cycle progression is halted until repair is completed. Several multiprotein complexes containing members of the structural maintenance of chromosomes family of proteins have been described, including the condensin and cohesin complexes, that are critical for chromosomal organization. Here we show that the Smc5/Smc6 (Smc5/6) complex is required for a coordinated response to DNA damage and normal chromosome integrity. Fission yeast cells lacking functional Smc6 initiate a normal checkpoint response to DNA damage, culminating in the phosphorylation and activation of the Chk1 protein kinase. Despite this, cells enter a lethal mitosis, presumably without completion of DNA repair. Another subunit of the complex, Nse1, is a conserved member of this complex and is also required for this response. We propose that the failure to maintain a checkpoint response stems from the lack of ongoing DNA repair or from defective chromosomal organization, which is the signal to maintain a checkpoint arrest. The Smc5/6 complex is fundamental to genome integrity and may function with the condensin and cohesin complexes in a coordinated manner.
Subject(s)
Cell Cycle Proteins/metabolism , DNA Damage/physiology , Schizosaccharomyces pombe Proteins/metabolism , Schizosaccharomyces/metabolism , Amino Acid Sequence , Base Sequence , Cell Cycle , Cell Cycle Proteins/genetics , DNA Damage/genetics , DNA Repair/genetics , DNA Repair/physiology , DNA, Fungal/genetics , DNA, Fungal/metabolism , Molecular Sequence Data , Mutation , Schizosaccharomyces/cytology , Schizosaccharomyces/genetics , Schizosaccharomyces pombe Proteins/genetics , Sequence Homology, Amino AcidABSTRACT
SUMMARY: The structural maintenance of chromosomes (SMC) proteins are essential for successful chromosome transmission during replication and segregation of the genome in all organisms. SMCs are generally present as single proteins in bacteria, and as at least six distinct proteins in eukaryotes. The proteins range in size from approximately 110 to 170 kDa, and each has five distinct domains: amino- and carboxy-terminal globular domains, which contain sequences characteristic of ATPases, two coiled-coil regions separating the terminal domains and a central flexible hinge. SMC proteins function together with other proteins in a range of chromosomal transactions, including chromosome condensation, sister-chromatid cohesion, recombination, DNA repair and epigenetic silencing of gene expression. Recent studies are beginning to decipher molecular details of how these processes are carried out.
