Reversible Top1 cleavage complexes are stabilized strand-specifically at the ribosomal replication fork barrier and contribute to ribosomal DNA stability.

Various topological constraints at the ribosomal DNA (rDNA) locus impose an extra challenge for transcription and DNA replication, generating constant torsional DNA stress. The topoisomerase Top1 is known to release such torsion by single-strand nicking and re-ligation in a process involving transient covalent Top1 cleavage complexes (Top1cc) with the nicked DNA. Here we show that Top1ccs, despite their usually transient nature, are specifically targeted to and stabilized at the ribosomal replication fork barrier (rRFB) of budding yeast, establishing a link with previously reported Top1 controlled nicks.

DNA repair and the control of DNA methylation.

The successful establishment and stable maintenance of cell identity are critical for organismal development and tissue homeostasis. Cell identity is provided by epigenetic mechanisms that facilitate a selective readout of the genome. Operating at the level of chromatin, they establish defined gene expression programs during cell differentiation.

DNA ligase 4 stabilizes the ribosomal DNA array upon fork collapse at the replication fork barrier.

DNA double-strand breaks (DSB) were shown to occur at the replication fork barrier in the ribosomal DNA of Saccharomyces cerevisiae using 2D-gel electrophoresis. Their origin, nature and magnitude, however, have remained elusive. We quantified these DSBs and show that a surprising 14% of replicating ribosomal DNA molecules are broken at the replication fork barrier in replicating wild-type cells.

Replication stress leads to genome instabilities in Arabidopsis DNA polymerase delta mutants.

Impeded DNA replication or a deficiency of its control may critically threaten the genetic information of cells, possibly resulting in genome alterations, such as gross chromosomal translocations, microsatellite instabilities, or increased rates of homologous recombination (HR). We examined an Arabidopsis thaliana line derived from a forward genetic screen, which exhibits an elevated frequency of somatic HR. These HR events originate from replication stress in endoreduplicating cells caused by reduced expression of the gene coding for the catalytic subunit of the DNA polymerase delta (POLdelta1).

Distinct roles for SWR1 and INO80 chromatin remodeling complexes at chromosomal double-strand breaks.

INO80 and SWR1 are two closely related ATP-dependent chromatin remodeling complexes that share several subunits. Ino80 was reported to be recruited to the HO endonuclease-induced double-strand break (DSB) at the budding yeast mating-type locus, MAT. We find Swr1 similarly recruited in a manner dependent on the phosphorylation of H2A (gammaH2AX).

The dual nature of homologous recombination in plants.

Homologous recombination creates covalent linkages between DNA in regions of highly similar or identical sequence. Recent results from several laboratories, many of them based on forward and reverse genetics in Arabidopsis, give insights into the mechanisms of the enzymatic machinery and the involvement of chromatin in somatic and meiotic DNA recombination. Also, signaling pathways and interconnections between repair pathways are being discovered.

Recruitment of the INO80 complex by H2A phosphorylation links ATP-dependent chromatin remodeling with DNA double-strand break repair.

The budding yeast INO80 complex is a conserved ATP-dependent nucleosome remodeler containing actin-related proteins Arp5 and Arp8. Strains lacking INO80, ARP5, or ARP8 have defects in transcription. Here we show that these mutants are hypersensitive to DNA damaging agents and to double-strand breaks (DSBs) induced by the HO endonuclease.

The INO80 protein controls homologous recombination in Arabidopsis thaliana.

Homologous recombination (HR) serves a dual role in providing genetic flexibility and in maintaining genome integrity. Little is known about the regulation of HR and other repair pathways in the context of chromatin. We report on a mutant affected in the expression of the Arabidopsis INO80 ortholog of the SWI/SNF ATPase family, which shows a reduction of the HR frequency to 15% of that in wild-type plants.

CENTRIN2 modulates homologous recombination and nucleotide excision repair in Arabidopsis.

A genetic screen of a population of Arabidopsis thaliana lines exhibiting enhanced somatic homologous recombination yielded a mutant affected in expression of a gene encoding a caltractin-like protein (centrin). The hyperrecombinogenic phenotype could be reproduced using RNA interference (RNAi) technology. Both the original mutant and the RNAi plants exhibited a moderate UV-C sensitivity as well as a reduced efficiency of in vitro repair of UV-damaged DNA.