Understanding PRMT5 activity in the DNA damage response in the context to MTAP deficiency

Faithful DNA damage repair (DDR) is essential for maintaining genome stability and the prevention of disease such as cancer and neurodegeneration. The protein methyltransferase PRMT5 has become associated with DDR and splicing with multiple studies indicating a role in double strand break (DSB) repair and the replication stress response.
Targeting PRMT5 therapeutically is a growing research area. First generation PRMT5 inhibitors have entered Phase I clinical trials but are limited clinically due to adverse off-target effects. In contrast, second generation inhibitors targeting PRMT5 in an MTAP null background have shown promising efficacy with reduced toxicities. MTAP is a gene that is proximal to the tumour suppressor gene CDKN2A and is regularly co-deleted in cancer. MTAP deficiency leads to the accumulation of a metabolite called MTA that is an endogenous inhibitor of PRMT5 catalytic activity. MTAP null cells are viable, whilst full PRMT5 inhibition tends to be lethal, this synthetic lethal relationship therefore implies that PRMT5 substrates in this genetic background are essential for cell survival and thus critical PRMT5 targets.
As PRMT5 has this known role in DDR, this PhD project will explore which PRMT5-mediated DNA damage repair pathway are vital for the survival of MTAP null cells. Using a panel of isogenic cell lines where the only genetic difference is MTAP status, a variety of methodologies will be used to address this question including protein biochemistry, immunoblotting, appropriate cell biology assays, RNA sequencing, microscopy, flow cytometry, DNA damaging agent screen and relevant DDR assays. To elucidate the consequences of MTAP depletion on PRMT5's interactome, a mass spectrometry approach will be utilised within the isogenic pairs after DNA damage to identify key PRMT5 substrates. After which potential novel interacting proteins will be assessed to identify PRMT5 methylation events and the functional relevance of this methylation to DDR.