[Show abstract][Hide abstract] ABSTRACT: Recent findings show that chromatin dynamics and nuclear organization are not only important for gene regulation and DNA replication, but also for the maintenance of genome stability. In yeast, nuclear pores play a role in the maintenance of genome stability by means of the evolutionarily conserved family of SUMO-targeted Ubiquitin ligases (STUbLs). The yeast Slx5/Slx8 STUbL associates with a class of DNA breaks that are shifted to nuclear pores. Functionally Slx5/Slx8 are needed for telomere maintenance by an unusual recombination-mediated pathway. The mammalian STUbL RNF4 associates with Promyelocytic leukaemia (PML) nuclear bodies and regulates PML/PML-fusion protein stability in response to arsenic-induced stress. A subclass of PML bodies support telomere maintenance by the ALT pathway in telomerase-deficient tumors. Perturbation of nuclear organization through either loss of pore subunits in yeast, or PML body perturbation in man, can lead to gene amplifications, deletions, translocations or end-to-end telomere fusion events, thus implicating SUMO and STUbLs in the subnuclear organization of select repair events.
Cell Research 02/2011; 21(3):474-85. · 11.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Asymmetric cell division drives the generation of differentiated cells and maintenance of stem cells. In budding yeast, autonomously replicating sequence (ARS) plasmids lacking centromere elements are asymmetrically segregated into the mother cell, where they are thought to contribute to cellular senescence. This phenomenon has been proposed to result from the active retention of plasmids through an interaction with nuclear pores.
To investigate the mother-daughter segregation bias of plasmids, we used live-cell imaging to follow the behavior of extrachromosomal DNA. We show that both an excised DNA ring and a centromere-deficient ARS plasmid move freely in the nucleoplasm yet show a strong segregation bias for the mother cell. Computational modeling shows that the geometrical shape of the dividing yeast nucleus and length of mitosis severely restrict the passive diffusion of episomes into daughter nuclei. Predictions based on simulated nuclear division were tested with mutants that extend the length of mitosis. Finally, explaining how various anchors can improve mitotic segregation, we show that plasmid partitioning is improved by tethering the plasmid to segregating structures, such as the nuclear envelope and telomeres.
The morphology and brevity of mitotic division in budding yeast impose physical constraints on the diffusion of material into the daughter, obviating the need for a retention mechanism to generate rejuvenated offspring.
Current biology: CB 01/2011; 21(1):25-33. · 10.99 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recent findings demonstrate that chromatin dynamics and nuclear organization are not only important for gene regulation but also for the maintenance of genome stability. Thanks to novel techniques that allow the visualization of specific chromatin domains in living cells, recent studies have demonstrated that the spatial dynamics of double-strand breaks and modifying enzymes can influence repair. The importance of the spatial organization in the repair of DNA damage has been confirmed by demonstrating that perturbation of nuclear organization can lead to gene amplifications, deletions, translocations and end-to-end telomere fusion events.
[Show abstract][Hide abstract] ABSTRACT: DNA damage during replication requires an integration of checkpoint response with replication itself and distinct repair pathways, such as replication pausing, recombination and translesion synthesis. Here we focus on recent advances in our understanding of how protein posttranslational modifications contribute to the maintenance of fork integrity. In particular, we examine the role of histone modifications and chromatin remodeling complexes in this process.
DNA repair 06/2009; 8(9):1089-100. · 3.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Budding yeast telomeres and cryptic mating-type loci are enriched at the nuclear envelope, forming foci that sequester silent information regulators (SIR factors), much as heterochromatic chromocenters in higher eukaryotes sequester HP1. Here we examine the impact of such subcompartments for regulating transcription genome-wide. We show that the efficiency of subtelomeric reporter gene repression depends not only on the strength of SIR factor recruitment by cis-acting elements, but also on the accumulation of SIRs in such perinuclear foci. To monitor the effects of disrupting this subnuclear compartment, we performed microarray analyses under conditions that eliminate telomere anchoring, while preserving SIR complex integrity. We found 60 genes reproducibly misregulated. Among those with increased expression, 22% were within 20 kb of a telomere, confirming that the nuclear envelope (NE) association of telomeres helps repress natural subtelomeric genes. In contrast, loci that were down-regulated were distributed over all chromosomes. Half of this ectopic repression was SIR complex dependent. We conclude that released SIR factors can promiscuously repress transcription at nontelomeric genes despite the presence of "anti-silencing" mechanisms. Bioinformatic analysis revealed that promoters bearing the PAC (RNA Polymerase A and C promoters) or Abf1 binding consenses are consistently down-regulated by mislocalization of SIR factors. Thus, the normal telomeric sequestration of SIRs both favors subtelomeric repression and prevents promiscuous effects at a distinct subset of promoters. This demonstrates that patterns of gene expression can be regulated by changing the spatial distribution of repetitive DNA sequences that bind repressive factors.
Genome Research 02/2009; 19(4):611-25. · 13.85 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recent findings suggest important roles for nuclear organization in gene expression. In contrast, little is known about how nuclear organization contributes to genome stability. Epistasis analysis (E-MAP) using DNA repair factors in yeast indicated a functional relationship between a nuclear pore subcomplex and Slx5/Slx8, a small ubiquitin-like modifier (SUMO)-dependent ubiquitin ligase, which we show physically interact. Real-time imaging and chromatin immunoprecipitation confirmed stable recruitment of damaged DNA to nuclear pores. Relocation required the Nup84 complex and Mec1/Tel1 kinases. Spontaneous gene conversion can be enhanced in a Slx8- and Nup84-dependent manner by tethering donor sites at the nuclear periphery. This suggests that strand breaks are shunted to nuclear pores for a repair pathway controlled by a conserved SUMO-dependent E3 ligase.