Distinct functions of PAXX and MRI during chromosomal end joining

A key step of canonical-nonhomologous end joining is synapsis of DNA double-strand break (DSB) ends, which appears to be mediated by both the DNA-PKcs dimer and XLF homodimer. We have examined this process by monitoring end joining (EJ) of blunt Cas9 DSBs, focusing on the Ku-binding factors PAXX and MRI. We found that PAXX and/or MRI are dispensable for such EJ. However, PAXX becomes important for blunt DSB EJ with disruption of either DNA-PKcs or XLF, whereas MRI becomes important only with XLF disruption. In contrast, while DNA-PKcs also suppresses short deletion mutations with microhomology, this effect is not magnified with PAXX loss. Finally, XLF loss causes an increase in larger deletions compared to DNA-PKcs inhibition, which is magnified with loss of MRI. We suggest that PAXX promotes DSB end synapsis in a manner that is partially redundant with DNA-PKcs and XLF, whereas MRI is a backup factor for XLF-mediated EJ.