The involvement of chromatin structure in DNA repair – ChromaRep

The integrity of our genome is constantly challenged by various assaults. DNA damage can occur due to both environmental agents such as UV light or irradiation, and endogenous sources, such as oxidative by-products of cellular metabolism or stalled replication forks. To prevent irreversible mutations occurring through our life span, multiple repair systems emerged during evolution. Depending on the chemical and physical properties of the lesion, different repair processes deal with the reversal of the damage. UV-induced photoproducts and bulky adducts are recognized and eliminated by the process of Nucleotide Excision Repair (NER). DNA double strand breaks (DSBs) are considered the most deleterious damage potentially leading to gross chromosomal rearrangements, whose repair, due to the lack of an undamaged complementary strand, is difficult. These lesions are repaired by the Double Strand Break Repair pathway (DSBR). Our proposed project focuses on these two repair pathways.
All types of DNA repair processes occur in the context of highly structured chromatin. Emerging evidence suggests that the ability of repair factors to detect DNA lesions and to be retained efficiently at the site of damage is determined by histone modifications around the lesion and involves chromatin-remodelling events. In response to DNA damage, chromatin undergoes rapid local and global decondensation, a process that has been proposed to facilitate genome surveillance by enhancing access of DDR proteins to sites of damage. Repair-associated chromatin remodelling is carried out by chromatin-remodelling complexes, which use ATP hydrolysis to remove histones or alter their conformation.
Although there is increasing evidence that chromatin alterations are essential for efficient NER and DSBR, the existing work is based on a candidate approach. Usually, the chromatin modifiers that are known to be involved in gene expression and transcriptional regulation are tested for their function in DNA repair. Moreover, there is emerging evidence that NER and DSBR have overlapping functions and that several common chromatin modifiers and remodelers affect the outcome of both repair processes.

We aim to undertake a systematic and unbiased approach and to perform two siRNA screens to
1. Identify novel chromatin related proteins that are involved in the repair of DSBs
2. Identify novel chromatin related proteins that are involved in NER
3. Compare the chromatin requirements for functional NER and DSBR
For that purpose, we have designed a siRNA library that targets a large number of the annotated chromatin modifiers, remodelers and other chromatin related proteins (1.184 genes including histone acetyltranferases, histone methyltransferases, ubiquitin ligases, ATP dependant chromatin remodelers). Our approach will provide for the first time substantial information about the involvement of chromatin in DNA repair and a thorough comparison of genes that commonly or differentially regulate NER and DSBR.
This application does not involve the purchase of the library, which will be available soon, but the funding of both screenings and the biochemical studies that will follow.