EMRES

Establishing the meiotic recombination-initiation epigenetic code in the yeast Saccharomyces cerevisiae

 Partecipanti

Mappa

 Word cloud

Esplora la "nuvola delle parole (Word Cloud) per avere un'idea di massima del progetto.

 Obiettivo del progetto (Objective)

'Meiotic recombination is initiated by programmed DNA double strand breaks (DSBs) catalysed by the evolutionary conserved Spo11 protein and associated factors. In the yeast S. cerevisiae, the DSB sites are non-randomly distributed along the chromosomes: certain genomic regions have higher propensity to form DSBs than others, explaining the expansion and contraction of the genetic versus physical maps. The cis- and trans-acting factors that select the DSB sites in the chromatin architecture and make a particular region 'hot' or ‘cold’ are not elucidated. The aim of this project is to decipher the role of histone modifications and other types of chromatin structural elements that orchestrate the recruitment and/or the cleavage activity of the DSB-forming machinery along the chromosomes. In wild type and a panel of mutant strains defective in histone modifying and remodeling activities, we will assess the spatial and temporal dynamics of the histone modification landscape upon DSB formation on the genome-wide scale using the ChIP-Chip, ChIP-Seq, Re-ChIP and FAIRE techniques. The chromatin-state of hot and cold regions will be mapped both at natural and ectopic hot/cold spots using the Gal4BD-Spo11 protein that is suitable for the targeted stimulation of recombination at naturally cold but permissive (warmable) regions but not at other cold (refractory) regions located near centromeres and telomeres. Successful completion of these studies will (1) include the construction of complete, genome-wide maps of meiotic chromatin-state in wild type and mutant yeast strains, and (2) improve our knowledge on how Spo11-dependent DSB formation is differentially controlled in relation to the chromatin context and coupled to homologous recombination, and differ from other sources of DNA lesions (in particular replication fork collapse) also leading to genomic rearrangements by homologous recombination.'

Introduzione (Teaser)

Budding yeast is being used to discover more about meiotic recombination - a common method of DNA repair in large complex organisms as well as microorganisms such as bacteria.

Descrizione progetto (Article)

Meiosis is a special kind of cell division that helps sexually reproducing populations adapt and evolve. Meiotic recombination begins when programmed DNA double-strand breaks (DSBs) are set in action by the Spo11 protein. This is an enzyme that cuts the DNA molecule due to receive new genetic information.

In the yeast Saccharomyces cerevisiae, it has been found that the DSB sites don't just randomly place themselves along chromosomes; certain genomic areas are more likely to form DSBs than others. Histones also play a part in this process. These are the main protein components of chromatin found in cell nuclei of eukaryotes and are responsible for arranging DNA into its most basic structural form - the nucleosome.

The EU-funded project 'Establishing the meiotic recombination-initiation epigenetic code in the yeast Saccharomyces cerevisiae' (EMRES) aimed to discover how histone modifications are involved in choosing and activating sites where meiotic recombination is set off. All work was done using the model organism S. cerevisiae (budding yeast).

Researchers constructed a number of deletion mutants in order to block genes coding for histone-modifying enzymes. Meiosis was followed and extensively described from the meiotic S phase, DSB and crossover formation, sporulation efficiency through to spore viability. The 'chip-on-chip' technique, used to measure binding sites for proteins, was used for genome-wide distribution of H3K56ac. This marker is vital for the proper assembly of nucleosomes and maintaining genome stability during DNA replication. This mapping showed a random distribution process of the H3K56ac histone mark, and no correlation was found between the sites of meiotic DSB formation and H3K56ac.

Having changed the original experimental strategy due to early study results, project partners developed a new, more adaptable means of examining histone modification during meiosis - a plasmid shuffling-based screen. After testing the screen, EMRES concluded that mutations of histone H3K4R and H3K4Q markers reiterate the effect of set1 deletion, which increases the capabilities of mutants following DNA damage. Following this, the research team was able to proceed with testing all the established histone mutants on the process of meiotic recombination.