Autophosphorylation of Polo-like Kinase 4 and Its Role in Centriole Duplication (original) (raw)
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Plk1 relieves centriole block to reduplication by promoting daughter centriole maturation
Nature communications, 2015
Centrosome overduplication promotes mitotic abnormalities, invasion and tumorigenesis. Cells regulate the number of centrosomes by limiting centriole duplication to once per cell cycle. The orthogonal orientation between a mother and a daughter centriole, established at the time of centriole duplication, is thought to block further duplication of the mother centriole. Loss of orthogonal orientation (disengagement) between two centrioles during anaphase is considered a licensing event for the next round of centriole duplication. Disengagement requires the activity of Polo-like kinase 1 (Plk1), but how Plk1 drives this process is not clear. Here we employ correlative live/electron microscopy and demonstrate that Plk1 induces maturation and distancing of the daughter centriole, allowing reduplication of the mother centriole even if the original daughter centriole is still orthogonal to it. We find that mother centrioles can undergo reduplication when original daughter centrioles are on...
Current Biology, 2010
Supernumerary centrioles lead to abnormal mitosis [1,2] which in turn promotes tumorigenesis [3,4]. Thus, centriole duplication must be coordinated with the cell cycle to ensure that the number of centrioles in the cell doubles precisely during each cell cycle [5]. However, in some transformed cells centrioles undergo multiple rounds of duplication (reduplication) during prolonged interphase [6-8]. Mechanisms responsible for centriole reduplication are poorly understood. Here, we report that centrioles reduplicate consistently in cancerous and nontransformed human cells during G2 arrests and this reduplication requires the activity of Polo-like kinase 1 (Plk1). We also find that cell's ability to reduplicate centrioles during S-arrests depends on the presence of activated (T210-phosphorylated) Plk1 at the centrosome. In the absence of activated Plk1, nascent procentrioles remain associated with mother centrioles, which prevent centriole reduplication. In contrast, if Plk1(pT210) appears at the centrosome, procentrioles mature, disengage from mother centrioles, and ultimately duplicate. Plk1 activity is not required for the assembly of procentrioles, however. Thus, the role of Plk1 is to coordinate centriole duplication cycle with the cell cycle. Activation of Plk1 during late-S-G2 induces procentriole maturation and after this point the centriole cycle can be completed autonomously, even in the absence of cell cycle progression. Results and Discussion Centrioles Reduplicate in G2-arrested Cells Normally, new centrioles (procentrioles) assemble at G1/S transition and they remain associated with mother centrioles until the ensuing mitosis [9]. If the cell cycle is arrested with hydroxyurea, in many cell types (e.g., HeLa) procentrioles form once and they remain associated with mother centrioles indefinitely. Interestingly, these procentrioles also remain significantly shorter than their mothers [7] and they lack hPOC5 (Figure S1, A-E), a protein normally recruited to the distal parts of procentrioles during G2 [10]. In contrast, some transformed cells (e.g., U2-OS) continuously reduplicate centrioles upon HU arrest [6,11]. In these cell types procentrioles develop to full length and recruit hPOC5 prior to disengaging from their mothers, and ultimately reduplicating (Figure S1, F-J). These differences prompted us to speculate that HU-arrested U2-OS cells progress farther toward
Cyclin-dependent kinase control of centrosome duplication
Proceedings of the National Academy of Sciences, 1999
Centrosomes nucleate microtubules and duplicate once per cell cycle. This duplication and subsequent segregation in mitosis results in maintenance of the one centrosome͞cell ratio. Centrosome duplication occurs during the G 1 ͞S transition in somatic cells and must be coupled to the events of the nuclear cell cycle; failure to coordinate duplication and mitosis results in abnormal numbers of centrosomes and aberrant mitoses. Using both in vivo and in vitro assays, we show that centrosome duplication in Xenopus laevis embryos requires cyclin͞cdk2 kinase activity. Injection of the cdk (cyclin-dependent kinase) inhibitor p21 into one blastomere of a dividing embryo blocks centrosome duplication in that blastomere; the related cdk inhibitor p27 has a similar effect. An in vitro system using Xenopus extracts carries out separation of the paired centrioles within the centrosome. This centriole separation activity is dependent on cyclin͞cdk2 activity; depletion of either cdk2 or of the two activating cyclins, cyclin A and cyclin E, eliminates centriole separation activity. In addition, centriole separation is inhibited by the mitotic state, suggesting a mechanism of linking the cell cycle to periodic duplication of the centrosome.
CDK1 Prevents Unscheduled PLK4-STIL Complex Assembly in Centriole Biogenesis
Current biology : CB, 2016
Centrioles are essential for the assembly of both centrosomes and cilia. Centriole biogenesis occurs once and only once per cell cycle and is temporally coordinated with cell-cycle progression, ensuring the formation of the right number of centrioles at the right time. The formation of new daughter centrioles is guided by a pre-existing, mother centriole. The proximity between mother and daughter centrioles was proposed to restrict new centriole formation until they separate beyond a critical distance. Paradoxically, mother and daughter centrioles overcome this distance in early mitosis, at a time when triggers for centriole biogenesis Polo-like kinase 4 (PLK4) and its substrate STIL are abundant. Here we show that in mitosis, the mitotic kinase CDK1-CyclinB binds STIL and prevents formation of the PLK4-STIL complex and STIL phosphorylation by PLK4, thus inhibiting untimely onset of centriole biogenesis. After CDK1-CyclinB inactivation upon mitotic exit, PLK4 can bind and phosphoryl...
SAK/PLK4 Is Required for Centriole Duplication and Flagella Development
Current Biology, 2005
Background: SAK/PLK4 is a distinct member of the polo-like kinase family. SAK 2/2 mice die during embryogenesis, whereas SAK +/2 mice develop liver and lung tumors and SAK +/2 MEFs show mitotic abnormalities. However, the mechanism underlying these phenotypes is still not known. Results: Here, we show that downregulation of SAK in Drosophila cells, by mutation or RNAi, leads to loss of centrioles, the core structures of centrosomes. Such cells are able to undergo repeated rounds of cell division, but display broad disorganized mitotic spindle poles. We also show that SAK mutants lose their centrioles during the mitotic divisions preceding male meiosis but still produce cysts of 16 primary spermatocytes as in the wild-type. Mathematical modeling of the stereotyped cell divisions of spermatogenesis can account for such loss by defective centriole duplication. The majority of spermatids in SAK mutants lack centrioles and so are unable to make sperm axonemes. Finally, we show that depletion of SAK in human cells also prevents centriole duplication and gives rise to mitotic abnormalities. Conclusions: SAK/PLK4 is necessary for centriole duplication both in Drosophila and human cells. Drosophila cells tolerate the lack of centrioles and undertake mitosis but cannot form basal bodies and hence flagella. Human cells depleted of SAK show error-prone mitosis, likely to underlie its tumor-suppressor role.
Nature communications, 2017
Centrosome number is tightly controlled during the cell cycle to ensure proper spindle assembly and cell division. However, the underlying mechanism that controls centrosome number remains largely unclear. We show herein that the DNA replication licensing factor Cdc6 is recruited to the proximal side of the centrioles via cyclin A to negatively regulate centrosome duplication by binding and inhibiting the cartwheel protein Sas-6 from forming a stable complex with another centriole duplication core protein, STIL. We further demonstrate that Cdc6 colocalizes with Plk4 at the centrosome, and interacts with Plk4 during S phase. Plk4 disrupts the interaction between Sas-6 and Cdc6, and suppresses the inhibitory role of Cdc6 on Sas-6 by phosphorylating Cdc6. Overexpressing wild-type Cdc6 or Plk4-unphosphorylatable Cdc6 mutant 2A reduces centrosome over-duplication caused by Plk4 overexpression or hydroxyurea treatment. Taken together, our data demonstrate that Cdc6 and Plk4 antagonistical...
Plk4 triggers autonomous de novo centriole biogenesis and maturation
2020
Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell-types, centrioles assemble de novo, yet by poorly understood mechanisms. Here, we established a controlled system to investigate de novo centriole biogenesis, usingDrosophila melanogasteregg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at high Plk4 concentration, centrioles form de novo, mature and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the kinetics of centriole assembly. Moreover, our results suggest Plk4 operates in a switch-like manner to control the onset of de novo centriole formation, and that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigate...