Regulatory cross-talk determines the cellular levels of 53BP1 protein, a critical factor in DNA repair

Abstract

DNA oubletrand reaks (DSBs) severely disrupt DNA integrity. 53BP1 plays critical roles in determining DSB repair. Whereas the recruitment of 53BP1 to the DSB site is key for its function, recent evidence suggests that 53BP1's abundance also plays an important role in DSB repair because recruitment to damage sites will be influenced by protein availability. Initial evidence has pointed to three proteins, the ubiquitin-conjugating enzyme UbcH7, the cysteine protease cathepsin L (CTSL), and the nuclear structure protein lamin A/C, that may impact 53BP1 levels, but the roles of each protein and any interplay between them were unclear. Here we report that UbcH7-dependent degradation plays a major role in controlling 53BP1 levels both under normal growth conditions and during DNA damage. CTSL influenced 53BP1 degradation during DNA damage while having little effect under normal growth conditions. Interestingly, both the protein and the mRNA levels of CTSL were reduced in UbcH7-depleted cells. Lamin A/C interacted with 53BP1 under normal conditions. DNA damage disrupted the lamin A/C-53BP1 interaction, which preceded the degradation of 53BP1 in soluble, but not chromatin-enriched, cellular fractions. Inhibition of 53BP1 degradation by a proteasome inhibitor or by UbcH7 depletion restored the 53BP1-lamin A/C interaction. Depletion of lamin A/C, but not CTSL, caused a similar enhancement in cell sensitivity to DNA damage as UbcH7 depletion. These data suggest that multiple pathways collectively fine-tune the cellular levels of 53BP1 protein to ensure proper DSB repair and cell survival.

The nuclear structural protein NuMA is a negative regulator of 53BP1 in DNA double-strand break repair

“…Hence, NuMA does not regulate 53BP1 expression but may rather sequestrate 53BP1 in the absence of damage to keep this master DDR effector in check. We note that other studies have shown degradation of 53BP1 by the proteasome after DNA damage (60,61). This effect was not observed in U2OS or HMT-3522 S1 cells in our experimental conditions, possibly due to cell line-specific expression and compartmentalization of the protein.…”

How cells ensure correct repair of DNA double-strand breaks

“…The ability of 53BP1 to bind to H2AK15ub is also limited by phosphorylation or acetylation of key residues within the ubiquitin-dependent recruitment region (13,55,56). In addition to its recruitment, the stability of 53BP1 is also regulated by proteasome-mediated degradation dependent on the E2 enzyme, UBCH7 (57). UBCH7 activity is up-regulated upon replication stress, leading to increased degradation of 53BP1 and therefore higher levels of HR.…”

A role of the 53BP1 protein in genome protection: structural and functional characteristics of 53BP1-dependent DNA repair

“…Gibbs-Seymour et al [155] also documented that lamins A/C interact with the 53BP1 protein under normal physiological conditions; however, DNA damage weakens this protein-protein interaction. In this case, the 53BP1 protein is degraded in the 26S proteasome, but the depletion of the ubiquitin-conjugating enzyme UbcH7 restores the lamin A/C-53BP1 complex [159]. Interestingly, the lamin precursor pre-lamin A interferes with damaged chromatin [160].…”