Reciprocal Regulation between 53BP1 and the Anaphase-Promoting Complex/Cyclosome Is Required for Genomic Stability during Mitotic Stress

Abstract

The anaphase-promoting complex/cyclosome (APC/C) is an E3 ubiquitin ligase that targets substrates for degradation to promote mitotic progression. Here, we show that the DNA damage response protein 53BP1 contains conserved KEN boxes that are required for APC/C-dependent degradation in early mitosis. Mutation of the 53BP1 KEN boxes stabilized the protein and extended mitotic duration, whereas 53BP1 knockdown resulted in a shorter and delayed mitosis. Loss of 53BP1 increased APC/C activity, and we show that 53BP1 is a direct APC/C inhibitor. Although 53BP1 function is not absolutely required for normal cell cycle progression, knockdown was highly toxic in combination with mitotic spindle poisons. Moreover, chemical inhibition of the APC/C was able to rescue the lethality of 53BP1 loss. Our findings reveal a reciprocal regulation between 53BP1 and APC/C that is required for response to mitotic stress and may contribute to the tumor-suppressor functions of 53BP1.

DNA damage response proteins regulating mitotic cell division: double agents preserving genome stability

“…Other DNA damage response proteins also have roles in mitotic progression: Loss of ATM or BRCA2 function reduces the time from nuclear envelop breakdown to anaphase onset in agreement with a role for these proteins in spindle checkpoint signaling . Also, 53BP1‐depleted cells spent less time in mitosis compared with controls . In contrast, Chk2‐deficient cells exhibit a prometaphase delay compared with controls, which is consistent with a role for Chk2 in mitotic spindle formation .…”

“…53BP1 was also initially proposed to participate in the mitotic spindle checkpoint based on its localization in the outer kinetochore from prophase until early anaphase and on its hyperphosphorylation in mitotic cell extracts . However, 53BP1 is absent from prometaphase kinetochores after prolonged mitotic delay by centrosome loss or inhibition of Eg5 kinesin that activate the mitotic spindle checkpoint, suggesting 53BP1 is not a typical spindle checkpoint component . Aurora B phosphorylates 53BP1 at serine 1342 (S1342) in vitro and in mitotic HeLa cells; furthermore, Aurora B inhibition by a small‐molecule inhibitor or expression of nonphosphorylatable mutant 53BP1‐S1342A protein reduces 53BP1 kinetochore staining compared with control cells, suggesting Aurora B phosphorylates S1342 to promote 53BP1 localization to kinetochores .…”

“…Aurora B phosphorylates 53BP1 at serine 1342 (S1342) in vitro and in mitotic HeLa cells; furthermore, Aurora B inhibition by a small‐molecule inhibitor or expression of nonphosphorylatable mutant 53BP1‐S1342A protein reduces 53BP1 kinetochore staining compared with control cells, suggesting Aurora B phosphorylates S1342 to promote 53BP1 localization to kinetochores . Depletion of 53BP1 or expression of mutant 53BP1‐S1342A increases the frequency of lagging chromosomes compared with control cells, indicating that 53BP1 is required for optimal chromosome segregation . 53BP1 colocalizes with the inner kinetochore marker ACA in merotelic kinetochores; furthermore, 53BP1 associates with MCAK by co‐immunoprecipitation experiments in mitotic cell extracts, suggesting a potential role for 53BP1 in merotelic error correction (Fig.…”

“…However, a direct correlation between loss of 53BP1 kinetochore localization and increased frequency of merotelic attachments remains to be established. Also, 53BP1 inhibits APC/C activity in cell culture and in vitro ; furthermore, 53BP1‐deficient cells are often delayed entering mitosis and transit through mitosis faster than controls .…”

Activation of the Anaphase Promoting Complex reverses multiple drug resistant cancer

“…More recently, accumulating evidence implicates the APC in cell cycle-independent functions such as neurogenesis, longevity, stress response and genome stability [23][24][25][26]. The idea that APC function is critical to ward off cancer is gaining traction, as the APC has been shown to be important for DNA repair decisions [27,28], to preserve genomic stability in humans and yeast [29,30], and APC impairment is associated with onset of drug resistance [31,32]. Our results, therefore, identify the APC as a potential therapeutic target that protects the genome, thereby delaying the development of aggressive treatment resistant tumors.…”