Genomic instability is an enabling hallmark of cancer, often triggered by abnormal expression and behaviour of DNA damage repair (DDR) proteins [1] High levels of mutations and aberrant posttranslational modifications (PTMs) can lead to changes in interacting partners and/or subcellular location, affecting the efficiency of DDR and causing subsequent mutagenesis [2,3]. These deficiencies in DDR can sensitise cancer cells to DNA damaging agents, such as chemotherapeutics and radiotherapy. Despite this, cancer cells use alternative DDR pathways to bypass cell death and become resistant to such therapeutics. Therefore, contributing genes and proteins to these reliant pathways have become prime candidates for oncology drug design and diagnostics. The prime examples are PARP1 and BRCA1/2, , targeted by the therapeutic inhibitor olaparib and diagnostic markers for breast and ovarian cancers, respectively [4].
Ionising radiation (IR) is detrimental for DNA and causes prolific cellular changes, such as cell morphology and activation of multiple signalling cascades. Whether response to IR is pro or antiapoptotic, trafficking between organelles is required for a proportion of proteins to function, as it is well documented that some proteins have localisationdependent roles and ‘moonlight’ between subcellular compartments [3,5]. Previous work studying relocalisation have been performed using lowthroughput techniques, which rely on effective fluorescent antibodies or GFPtagging [6]. To investigate this trafficking, we applied a holistic and novel spatial proteomics workflow, LOPITDC. This technique uses the combination of differential ultracentrifugation for organelle separation, multiplexed quantitative mass spectrometry and machine learning to produce subcellular proteome maps [7]. Currently, we have successfully produced extremely reproducible, high-resolution spatial proteome maps of dynamic protein re-localisation in a lung carcinoma cell line (A549) in response to ionising radiation. We hope this will give us a deeper understanding of these proteins’ roles and behaviour to this stimulus and to aid DDR-targeting drug design.