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Research Projects

Loizou Lab

DNA Damage and Repair Mechanisms

The human genome is constantly exposed to endogenous and exogenous sources of DNA damage. DNA repair ensures the integrity of large eukaryotic genomes by minimising mutation rates and thereby suppressing the development of cancer. We are interested in exploring the several highly effective pathways for DNA repair that have distinct specificities and are evolutionary conserved despite partial redundancies. By investigating the genetic interactions of the intricate crosstalk between DNA damage and repair mechanisms our ultimate goal is that our findings may pave the way to rational therapeutic approaches.

Consequences of DNA damage and repair on genomic mutation signatures

Figure 1: Mutations result due to DNA damage and ensuing DNA repair processes. We are developing experimental approaches to identify specific mutational outcomes, known as mutation signatures, that occur within the human genome.

The compilation of somatic mutations is the outcome of one or more mutational processes that have been operative due to DNA damage and repair processes. The resulting mutational signature is determined by the intensity and duration of exposure to each mutational process. We seek to systematically map the contribution of various DNA damage and repair processes to the generation of mutation signatures hence deciphering the contribution of genetic and environmental sources to genome stability (Figure 1).

Synthetic lethal and viable interactions

Figure 2: 1. DNA damage is preferentially repaired by Pathway 1 and 2. mutations in Gene B within this pathway lead to defects in DNA repair. 3. Mutation of Gene A, that is a suppressor of alternative Pathway 2, allows for resumption of DNA repair. 4. Inhibition of Gene A is an alternative approach to allow for the alternative Pathway 2 to be utilized.

Complex interactions between different genes have been a focus of genetic research on model organisms for decades. The concepts of synthetic lethality and synthetic viability have been extensively explored because defects in a specific cellular pathway may (de)sensitize cells for the loss of another. Using genome-scale approaches, including CRISPR screens, we aim to identify new therapeutic targets, at scale, to ameliorate pathologies. Our hypothesis is that through the exploration of genetic, proteomic and chemical space will identify novel regulatory pathways for DNA repair (Figure 2). 

Repair of CRISPR-Cas9 generated DNA breaks

Figure 3: DNA breaks generated by Cas9 are repaired in an error-free manner via homology-directed repair or in an error-prone manner via either classical non-homologous end-joining or microhomology-mediated end-joining.

Repair of DNA double-strand breaks is thought to occur mainly via two major pathways: error-prone non-homologous and joining (NHEJ) and error-free homologous recombination (HR).

An enzyme that generates DNA double strand breaks, and is extensively used to promote gene editing, is Cas9. While large efforts have gone into the utilization of CRISPR-Cas9 for gene editing purposes, comparatively little advancement has been made with regard to the cellular mechanisms by which DNA lesions generated by Cas9 are dealt with.

We are interested in understanding the contribution of DNA repair pathways to the resolution of DNA double-strand breaks generated by different versions of Cas9 (Figure 3).