SSA - Genetic and Drug Screening in DNA Repair Deficient Zebrafish for Novel Targets in the Treatment of Neurological Disease and Cancer

Lead Research Organisation: University of Sheffield
Department Name: Biomedical Science

Abstract

A cell can experience ~1 million DNA lesions per day from endogenous and exogenous genotoxins. A variety of lesions also result from aberrant replication, or DNA repair itself. DNA lesions threaten essential processes such as transcription and replication and can lead apoptosis and cancer. For example, accumulation of protein-linked DNA breaks (PDBs) cause various neurological diseases, and on the other hand, have been exploited to treat cancer.
Although we know much about DNA-repair pathways from studies in cultured cells, we know little about the extent of functional redundancy at the organismal level. This is important since harnessing this knowledge is rapidly emerging as a powerful approach to treat diseases such as neurodegeneration and cancer. For example, we recently reported, in Nature Neuroscience, a novel mechanism by which PDBs and DNA/RNA hybrids cause motor neuron disease.
Through BBSRC funding and a joint studentship, we established CRISPR mutants in tdp1, brca2, atm, rad52, rad51, which all act in DNA repair, but are mostly viable and often have only mild/no defects as embryos. The objective of the PhD project is to identify backup pathways that can protect the organism if the primary PDB repair pathway is absent. As DNA-repair pathways are often redundant, homozygous mutants provide an excellent background for chemical/genetic modifier screens. Our primary focus will be on tdp1 mutants, these embryos are - surprisingly- as resistant to DNA damage, as their siblings. We will use CRISPR/CRISPRi technology, and have various chemical libraries available and will screen for defects after induction of DNA damage. We developed an in vivo GFP-reporter system, that uses destruction of a sentinel-repressor to show GFP activation after defective DNA repair. This provides a simple readout in embryos to quantify DNA repair. Importantly, identification of mechanisms behind redundancy may suggest clinical strategies to treat the human disease that results from mutation of tdp1, SCAN1. Moreover, it will provide a platform to stratify cancer patients receiving TDP1 inhibitors currently under development in our labs in collaboration with CRUK technology arm (CRT).

Publications

10 25 50

Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M011151/1 30/09/2015 29/09/2023
2109800 Studentship BB/M011151/1 30/09/2018 31/12/2022 Ruth Thomas