DNA-PK promotes DNA end resection at DNA double strand breaks in G cells.

DNA double-strand break (DSB) repair by homologous recombination is confined to the S and G phases of the cell cycle partly due to 53BP1 antagonizing DNA end resection in G phase and non-cycling quiescent (G) cells where DSBs are predominately repaired by non-homologous end joining (NHEJ). Unexpectedly, we uncovered extensive MRE11- and CtIP-dependent DNA end resection at DSBs in G murine and human cells. A whole genome CRISPR/Cas9 screen revealed the DNA-dependent kinase (DNA-PK) complex as a key factor in promoting DNA end resection in G cells.

A Proximity Ligation Method to Detect Proteins Bound to Single-Stranded DNA after DNA End Resection at DNA Double-Strand Breaks.

After a DNA double-strand break, cells utilize either non-homologous end joining or homologous recombination to repair the broken DNA ends. Homologous recombination requires extensive nucleolytic processing of one of the DNA strands, resulting in long stretches of 3' single-strand DNA overhangs. Typically, single-stranded DNA is measured using immunofluorescence microscopy to image the foci of replication protein A, a single-stranded DNA-binding protein.

LIN37-DREAM prevents DNA end resection and homologous recombination at DNA double-strand breaks in quiescent cells.

DNA double-strand break (DSB) repair by homologous recombination (HR) is thought to be restricted to the S- and G- phases of the cell cycle in part due to 53BP1 antagonizing DNA end resection in G-phase and non-cycling quiescent (G) cells. Here, we show that LIN37, a component of the DREAM transcriptional repressor, functions in a 53BP1-independent manner to prevent DNA end resection and HR in G cells. Loss of LIN37 leads to the expression of HR proteins, including BRCA1, BRCA2, PALB2, and RAD51, and promotes DNA end resection in G cells even in the presence of 53BP1.

Proteomic identification of histone post-translational modifications and proteins enriched at a DNA double-strand break.

Here, we use ChAP-MS (chromatin affinity purification with mass spectrometry), for the affinity purification of a sequence-specific single-copy endogenous chromosomal locus containing a DNA double-strand break (DSB). We found multiple new histone post-translational modifications enriched on chromatin bearing a DSB from budding yeast. One of these, methylation of histone H3 on lysine 125, has not previously been reported.