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Repair of double-stranded DNA breaks (DSBs) in mammalian cells primarily occurs by the non-homologous end-joining (NHEJ) pathway, which requires seven core proteins (Ku70/Ku86, DNA-PKcs (DNA-dependent protein kinase catalytic subunit), Artemis, XRCC4-like factor (XLF), XRCC4 and DNA ligase IV). Here we show using combined affinity purification and mass spectrometry that DNA-PKcs co-purifies with all known core NHEJ factors. Furthermore, we have identified a novel evolutionary conserved protein associated with DNA-PKcs-c9orf142. Computer-based modelling of c9orf142 predicted a structure very similar to XRCC4, hence we have named c9orf142-XLS (XRCC4-like small protein). Depletion of c9orf142/XLS in cells impaired DSB repair consistent with a defect in NHEJ. Furthermore, c9orf142/XLS interacted with other core NHEJ factors. These results demonstrate the existence of a new component of the NHEJ DNA repair pathway in mammalian cells. [ABSTRACT FROM AUTHOR]

Post-transcriptional control of gene expression is mediated by the interaction of RNA-binding proteins with their cognate mRNAs that specifically regulate their stability, localization and translation. mRNA-binding proteins are multifunctional and it has been proposed therefore that a combinatorial RNA-binding protein code exists that allows specific protein sub-complexes to control cytoplasmic gene expression under a range of pathophysiological conditions. We show that polypyrimidine tract-binding protein (PTB) is central to one such complex that forms in apoptotic cells. Thus, during apoptosis initiated by TNF-related apoptosis inducing ligand there is a change in the repertoire of RNA-binding proteins with which PTB interacts. We show that altering the cellular levels of PTB and its binding partners, either singly or in combination, is sufficient to directly change the rates of apoptosis with increased expression of PTB, YBX1, PSF and NONO/p54nrb accelerating this process. Mechanistically, we show that these proteins post-transcriptionally regulate gene expression, and therefore apoptotic rates, by interacting with and stimulating the activity of RNA elements (internal ribosome entry segments) found in mRNAs that are translated during apoptosis. Taken together, our data show that PTB function is controlled by a set of co-recruited proteins and importantly provide further evidence that it is possible to dictate cell fate by modulating cytoplasmic gene expression pathways alone. [ABSTRACT FROM AUTHOR]