Dephosphorylation of Plk1 occurs through PP2A-B55/ENSA/Greatwall pathway during mitotic DNA damage recovery

Abstract

Recovery from DNA damage is critical for cell survival. However, serious damage cannot be repaired, leading to cell death for prevention of abnormal cell growth. Previously, we demonstrated that 4N-DNA accumulates via the initiation of an abnormal interphase without cytokinesis and that re-replication occurs during a prolonged recovery period in the presence of severe DNA damage in mitotic cells. Mitotic phosphorylated Plk1 is typically degraded during mitotic exit. However, Plk1 has unusually found to be dephosphorylated in mitotic slippage without cytokinesis during recovery from mitotic DNA damage. Here, we investigated how Plk1 dephosphorylation is established during recovery from mitotic DNA damage. Mitotic DNA damage activated ATM and Chk1/2 and repressed Cdk1 and Greatwall protein kinase, followed by PP2A activation through the dissociation of ENSA and PP2A-B55. Interaction between Plk1 and PP2A-B55α or PP2A-B55δ was strongly induced during recovery from mitotic DNA damage. Moreover, the depletion of PP2A-B55α and/or PP2A-B55δ by siRNA transfection led to the recovery of Plk1 phosphorylation and progression of the cell cycle into the G 1 phase. Therefore, to adapt to severe DNA damage, the activated Greatwall/ENSA signaling pathway was repressed by ATM/Chk1/2, even in mitotic cells. Activation of the PP2A-B55 holoenzyme complex induced the dephosphorylation of Plk1 and Cdk1, and finally, mitotic slippage occurred without normal chromosome segregation and cytokinesis.

Kinetochore phosphatases suppress autonomous Polo-like kinase 1 activity to control the mitotic checkpoint

“…These small differences could reflect modest effects of PP2A on MELT dephosphorylation, but given the strength of PP2A inhibition under these conditions, and the fact that 43% of phospho-peptides are regulated by knockdown of PP2A subunits alone ( Kauko et al, 2020 ), we prefer the interpretation that these effects are either off-target or due to elevated kinase activity. For example, PLK1 and/or MPS1 activity may be increased—both of these are known targets of PP2A ( Hayward et al, 2019 ; Kim et al, 2019 ; Wang et al, 2015 )—or other phospho-epitopes may be elevated that react with the pMELT antibody.…”

PP2A and tumor radiotherapy

“…This greatwall kinase/PP2A signaling pathway is thought to be a primary regulator of normal Cdk1 functionality in the context of mitosis [ 42 ]. PP2A can also act on other mitotic mediators such as the mitosis-specific kinases PLK1, which is a key marker of G2/M phase arrest following PP2A inhibition and which interacts with centrosomes during mitosis [ 43 , 44 ]. PP2A is also involved in the negative feedback inhibition of PLK1 and Aurora B, thereby regulating the spindle collection checkpoint in order to ensure that microtubules are properly connected to the centromere [ 45 ].…”

“…Following drug- or radiation-induced DNA damage, PP2A dephosphorylation can inhibit PLK1, which phosphorylates and activates Cdc25 and cyclins involved in the G2/M checkpoint, thereby facilitating cell cycle progression. PP2A may thus prevent cells from dividing by inhibiting PLK1 [ 44 ]. Moreover, inhibition of PP2A showed that radiation-induced inactivation of ATR and Chk1 kinase, phosphorylation of Cdc2-Tyr15, and inactivation of G2/M phase checkpoints, which attenuated radiation-induced G2/M arrest, thereby enabling tumor cells to enter into mitosis via reducing DNA damage repair efficiency and aggravating cellular mitotic disorders [ 51 ].…”

Cell Cycle and DNA Repair Regulation in the Damage Response: Protein Phosphatases Take Over the Reins

“…The interaction between PLK1 and PP2A-B55α or PP2A-B55δ is stimulated during recovery from DNA damage. In addition, the disruption of PP2A activity promotes PLK1 phosphorylation and progression into mitosis [151]. In line with these findings, MASTL, the human homologue of Greatwall, is not required either for DDR activation nor DNA repair, but it is required for controlling the timing of mitosis re-entry and the subsequent fate of the recovering cells [152].…”