Characterisation of DNA-protein crosslink proteolysis repair in response to chemotherapy

Primary Supervisor: Professor Kristijan Ramadan

Project Overview:

DNA-protein crosslinks (DPCs) are under-investigated DNA lesions caused by the covalent attachment of proteins to DNA. DPCs are induced by endogenous aldehydes, chemotherapeutic drugs (such as PARP1 or Topoisomerase inhibitors) and ionising radiation. However, little is known about DPC repair and how do cells acquire resistance to DPC-induced chemo- or radiotherapy. We discovered that the SPRTN metalloprotease is a specialised and essential enzyme for DNA-protein crosslink repair during DNA replication fork progression (Lessel et al. Nature Genetics 2014, Vaz et al. Molecular Cell, 2016). Mutations in SPRTN causes Ruijs-Aalfs Syndrome, characterised by premature ageing and liver cancer (Figure 1). 

Figure 1 - Mutations in SPRTN causes Ruijs-Aalfs Syndrome, characterised by premature ageing and liver cancer.

Importantly, we have just discovered that SPRTN and its partner - the ATPase p97 - process two chemotherapy induced DPC lesions;  topoisomerase 1-cleavage complex and trapped PARP-1. These two DPC lesions are essential for therapeutic effect of topoisomerase and PARP inhibitors in colorectal and breast cancer, respectively (Fielden et al. Nature Communications, 2020 and Krastev et al., Nature Cell Biology, 2022).

The enzymes involved in DNA repair are promising druggable targets for cancer therapy. Our discovery of the SPRTN metalloprotease and ATPase p97 as the essential components of DNA repair have been recognised as the promising druggable targets for cancer therapy.

DPhil project aims to characterise SPRTN and p97 enzymatic activities in the context of DNA-protein crosslink repair pathway, with the special focus on trapped-PARP1. You will use standard biochemical and cell biological techniques coupled to state-of-the-art technologies such as CRISPR/Cas9 gene editing, mass-spectrometry and super-resolution microscopy to address urgent questions to be answered in DNA-protein crosslink proteolysis repair and cancer therapy fields. Your work will directly help us to elucidate the mechanism of DNA-protein crosslink proteolysis and trapped-PARP1 (DPC-like lesion) repair in genome stability and how chemical inhibition of the SPRTN protease and/or p97 ATPase could overcome chemotherapy resistance, especially in the context of hypoxic tumours. Here generated knowledge will not only elucidate some basic mechanisms of DNA-protein crosslink proteolysis repair but it will also help us to improve chemotherapy outcome of PARP inhibitors.

Training Opportunities: 

We have a dynamic and vibrant environment in our laboratories (Ramadan and D’Angiolella). Our laboratories have an extensive expertise in biochemistry, molecular and cell biology, ionizing radiation, genome stability, the ubiquitin system and protein degradation in cancer research. We use state-of-the-art technologies such as Quantitative Mass-Spectrometry, CRISPR/Cas9 gene editing, Confocal Microscopy, FACS, DNA combing, DNA repair and High-Throughput live cell imaging and analysis. You, as a PhD/DPhil student will get an excellent training in the aforementioned laboratory technologies.  Most importantly, once you obtain your PhD/DPhil under our supervision, you will become an independent and competitive scientist on the market. All our students have so far secured their next-destination jobs at the most prestigious scientific institutions worldwide, such as Oxford, Cambridge, Harvard or pharma companies.

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Relevant Publications:

Krastev, D.B., Li, S., Sun, Y., Wicks, A.J., Hoslett, G., Weekes, D., Badder, L.M., Knight, E.G., Marlow, R., Pardo, M.C. and Yu, L., 2022. The ubiquitin-dependent ATPase p97 removes cytotoxic trapped PARP1 from chromatin. Nature cell biology, 24(1), pp.62-73.

Vaz, B., Popovic, M., Newman, J.A., Fielden, J., Aitkenhead, H., Halder, S., Singh, A.N., Vendrell, I., Fischer, R., Torrecilla, I. and Drobnitzky, N., 2016. Metalloprotease SPRTN/DVC1 orchestrates replication-coupled DNA-protein crosslink repair. Molecular cell, 64(4), pp.704-719.

Lessel, D., Vaz, B., Halder, S., Lockhart, P.J., Marinovic-Terzic, I., Lopez-Mosqueda, J., Philipp, M., Sim, J.C., Smith, K.R., Oehler, J. and Cabrera, E., 2014. Mutations in SPRTN cause early onset hepatocellular carcinoma, genomic instability and progeroid features. Nature genetics, 46(11), pp.1239-1244.