The Cdc14B-Cdh1-Plk1 Axis Controls the G2 DNA-Damage-Response Checkpoint

Abstract

Summary In response to DNA damage in G2, mammalian cells must avoid entry into mitosis and instead initiate DNA repair. Here we show that in response to genotoxic stress in G2, the phosphatase Cdc14B translocates from the nucleolus to the nucleoplasm and induces the activation of the ubiquitin ligase APC/CCdh1, with the consequent degradation of Plk1, a prominent mitotic kinase. This process induces the stabilization of Claspin and Wee1, as the proteolysis of these two proteins requires phosphorylation by Plk1, and allows an efficient G2 checkpoint. As a by-product of APC/CCdh1 reactivation in DNA-damaged G2 cells, Claspin, which we show to be a novel substrate of APC/CCdh1 in G1, is targeted for degradation. However, this process is counteracted by the deubiquitylating enzyme Usp28 to permit Claspin-mediated activation of Chk1 in response to DNA damage. These findings define a novel pathway that is crucial for the G2 DNA damage response checkpoint.

p53- and p21-dependent premature APC/C–Cdh1 activation in G2 is part of the long-term response to genotoxic stress

“…Our data are in good agreement with two recent studies that also used HCT116 cells to show p21-dependent destabilization of cyclin-B1 (Gillis et al, 2009;Lee et al, 2009), cyclin-A, Cdc20, Securin and Emi1 ) after induction of DNA damage. We did not, however, find experimental support for the rapid induction of APC/C activity that has been reported for X-irradiated HeLa cells (Sudo et al, 2001) and doxorubicin-treated U2OS cells (Bassermann et al, 2008). Possibly, the chemical inhibitors of DNA replication that were used in both studies of cell-cycle synchronization before induction of DNA damage particularly prime cells for rapid APC/C activation.…”

“…The same factor was previously shown to be required for premature APC/C activation in doxorubicin-treated U2OS cells, by using an short interfering RNA-based knockdown approach (Bassermann et al, 2008). Surprisingly, Cdc14BÀ/À cells were still capable of activating the APC/C after doxorubicin treatment to the same extent as wt cells (Figure 7e).…”

“…In principle, the anaphase-promoting complex or cyclosome (APC/C) would represent an attractive target for such a p53-dependent regulation, as this ubiquitin ligase functions as a master regulator for the stability of several rate-limiting proteins controlling replication and cell division (Peters, 2006). Indeed, two reports have shown that in response to DNA damage the APC/C, which is normally kept inactive from the late G 1 to early M-phase, is prematurely activated in G2 (Sudo et al, 2001;Bassermann et al, 2008). However, this activation followed DNA damage with short-term kinetics (as early as 90 min), making a p53-driven mechanism involving de novo protein synthesis unlikely.…”

BimEL is phosphorylated at mitosis by Aurora A and targeted for degradation by βTrCP1

“…The siRNA oligonucleotide sequence targeting bTRCP1/2 has been described previously 54 and corresponds to nt 515-535 of human bTRCP1 and nt 262-282 of human bTRCP2. The following siRNA sequences were used to knock down Aurora A: 55 5 0 -725 AUGCCC UGUCUUACUGUCA 743 -3 0 and 5 0 -155 AUUCUUCCCAGCGCGUUCC 173 -3 0 .…”

Un nouvel axe du mal tumoral ?

“…Par exemple, Cdc14B pourrait participer à l'activation de l'APC Cdh1 en fin de mitose et en G1, comme cela a été démontré chez la levure S. cerevisiae [7]. En accord avec cette idée, on a récemment observé que Cdc14B prend part à l'activation de Cdh1 en réponse au stress génotoxique [8]. De plus, il est possible qu'en déphosphorylant activement p27, Cdc14B la stabilise et favorise l'établissement et/ou le maintien de la phase G1.…”