scispace - formally typeset
Search or ask a question
Journal ArticleDOI

A PIM-CHK1 signaling pathway regulates PLK1 phosphorylation and function during mitosis.

TL;DR: A new PIM-CHK1-PLK1 phosphorylation cascade that regulates different mitotic steps independently of the CHK1 DNA damage response function is identified.
Abstract: Although the kinase CHK1 is a key player in the DNA damage response (DDR), several studies have recently provided evidence of DDR-independent roles of CHK1, in particular following phosphorylation of its S280 residue. Here, we demonstrate that CHK1 S280 phosphorylation is cell cycle-dependent and peaks during mitosis. We found that this phosphorylation was catalyzed by the kinase PIM2, whose protein expression was also increased during mitosis. Importantly, we identified polo-like kinase 1 (PLK1) as a direct target of CHK1 during mitosis. Genetic or pharmacological inhibition of CHK1 reduced the activating phosphorylation of PLK1 on T210, and recombinant CHK1 was able to phosphorylate T210 of PLK1 in vitro Accordingly, S280-phosphorylated CHK1 and PLK1 exhibited similar specific mitotic localizations, and PLK1 was co-immunoprecipitated with S280-phosphorylated CHK1 from mitotic cell extracts. Moreover, CHK1-mediated phosphorylation of PLK1 was dependent on S280 phosphorylation by PIM2. Inhibition of PIM proteins reduced cell proliferation and mitotic entry, which was rescued by expressing a T210D phosphomimetic mutant of PLK1. Altogether, these data identify a new PIM-CHK1-PLK1 phosphorylation cascade that regulates different mitotic steps independently of the CHK1 DDR function.This article has an associated First Person interview with the first author of the paper.

Content maybe subject to copyright    Report

Citations
More filters
Journal ArticleDOI
01 Feb 2021-Leukemia
TL;DR: It is demonstrated that deubiquitinylase USP7 is both a marker of resistance to chemotherapy and a potential therapeutic target in overcoming resistance to treatment.
Abstract: Resistance of acute myeloid leukemia (AML) to therapeutic agents is frequent. Consequently, the mechanisms leading to this resistance must be understood and addressed. In this paper, we demonstrate that inhibition of deubiquitinylase USP7 significantly reduces cell proliferation in vitro and in vivo, blocks DNA replication progression and increases cell death in AML. Transcriptomic dataset analyses reveal that a USP7 gene signature is highly enriched in cells from AML patients at relapse, as well as in residual blasts from patient-derived xenograft (PDX) models treated with clinically relevant doses of cytarabine, which indicates a relationship between USP7 expression and resistance to therapy. Accordingly, single-cell analysis of AML patient samples at relapse versus at diagnosis showed that a gene signature of the pre-existing subpopulation responsible for relapse is enriched in transcriptomes of patients with a high USP7 level. Furthermore, we found that USP7 interacts and modulates CHK1 protein levels and functions in AML. Finally, we demonstrated that USP7 inhibition acts in synergy with cytarabine to kill AML cell lines and primary cells of patients with high USP7 levels. Altogether, these data demonstrate that USP7 is both a marker of resistance to chemotherapy and a potential therapeutic target in overcoming resistance to treatment.

17 citations

Journal ArticleDOI
TL;DR: In this paper, the authors show that PLK1 is enriched at double strand breaks (DSBs) within seconds of UV laser irradiation in a PARP-1dependent manner and then disperses within 10 min in aPARG-dependent manner.
Abstract: Polo-like kinase 1 (PLK1) is a master kinase that regulates cell cycle progression. How its enzymatic activity is regulated in response to DNA damage is not fully understood. We show that PLK1 is enriched at double strand breaks (DSBs) within seconds of UV laser irradiation in a PARP-1-dependent manner and then disperses within 10 min in a PARG-dependent manner. Poly(ADP-)ribose (PAR) chains directly bind to PLK1 in vitro and inhibit its enzymatic activity. CHK1-mediated PLK1 phosphorylation at S137 prevents its binding to PAR and recruitment to DSBs but ensures PLK1 phosphorylation at T210 and its enzymatic activity toward RAD51 at S14. This subsequent phosphorylation event at S14 primes RAD51 for CHK1-mediated phosphorylation at T309, which is essential for full RAD51 activation. This CHK1-PLK1-RAD51 axis ultimately promotes homologous recombination (HR)-mediated repair and ensures chromosome stability and cellular radiosensitivity. These findings provide biological insight for combined cancer therapy using inhibitors of PARG and CHK1.

17 citations

Journal ArticleDOI
TL;DR: In vivo, the therapeutic nanoparticle-mediated delivery of PIM2 siRNA led to profound anti-tumor effects in a liver cancer xenograft mouse model, and the potential of targeted therapies based on the specific inhibition of Pim2 in liver cancer is emphasized.
Abstract: Liver cancer is the fourth leading cause of cancer‑related mortality worldwide with limited therapeutic options. Thus, novel treatment strategies are urgently required. While the oncogenic kinase, proviral integration site for Moloney murine leukemia virus 2 (PIM2), has been shown to be overexpressed in liver cancer, little is known about the role of PIM2 in this tumor entity. In this study, we explored the functional relevance and therapeutic potential of PIM2 in liver cancer. Using PIM2‑specific siRNAs, we examined the effects of PIM2 knockdown on proliferation (WST‑1 assays and spheroid assays), 3D‑colony formation and colony spread, apoptosis (flow cytometry and caspase 3/caspase 7 activity), as well as cell cycle progression (flow cytometry, RT‑qPCR and western blot analysis) in the two liver cancer cell lines, HepG2 and Huh‑7. In subcutaneous liver cancer xenografts, we assessed the effects of PIM2 knockdown on tumor growth via the systemic delivery of polyethylenimine (PEI)‑complexed siRNA. The knockdown of PIM2 resulted in potent anti‑proliferative effects in cells grown on plastic dishes, as well as in spheroids. This was due to G0/G1 cell cycle blockade and the subsequent downregulation of genes related to the S phase as well as the G2/M phase of the cell cycle, whereas the apoptotic rates remained unaltered. Furthermore, colony formation and colony spread were markedly inhibited by PIM2 knockdown. Notably, we found that HepG2 cells were more sensitive to PIM2 knockdown than the Huh‑7 cells. In vivo, the therapeutic nanoparticle‑mediated delivery of PIM2 siRNA led to profound anti‑tumor effects in a liver cancer xenograft mouse model. On the whole, the findings of this study underscore the oncogenic role of PIM2 and emphasize the potential of targeted therapies based on the specific inhibition of PIM2 in liver cancer.

16 citations

Journal ArticleDOI
TL;DR: In this paper , the authors discuss the DNA damage checkpoint, the mitotic networks it inhibits to prevent segregation of damaged chromosomes and the strategies cells employ to quench the checkpoint controls to override the G2/M arrest.
Abstract: Chromosomes are susceptible to damage during their duplication and segregation or when exposed to genotoxic stresses. Left uncorrected, these lesions can result in genomic instability, leading to cells' diminished fitness, unbridled proliferation or death. To prevent such fates, checkpoint controls transiently halt cell cycle progression to allow time for the implementation of corrective measures. Prominent among these is the DNA damage checkpoint which operates at G2/M transition to ensure that cells with damaged chromosomes do not enter the mitotic phase. The execution and maintenance of cell cycle arrest are essential aspects of G2/M checkpoint and have been studied in detail. Equally critical is cells' ability to switch-off the checkpoint controls after a successful completion of corrective actions and to recommence cell cycle progression. Interestingly, when corrective measures fail, cells can mount an unusual cellular response, termed adaptation, where they escape checkpoint arrest and resume cell cycle progression with damaged chromosomes at the cost of genome instability or even death. Here, we discuss the DNA damage checkpoint, the mitotic networks it inhibits to prevent segregation of damaged chromosomes and the strategies cells employ to quench the checkpoint controls to override the G2/M arrest.

4 citations

Journal ArticleDOI
TL;DR: Zhang et al. as discussed by the authors found that TIG1 can regulate the tumorigenesis of colorectal cancer cells by regulating the activity of the Polo-like kinases (Plks).

3 citations

References
More filters
Journal ArticleDOI
TL;DR: It is shown that in human cells, Chk1 is phosphorylated on serine 345 (S345) in response to UV, IR, and hydroxyurea (HU).
Abstract: Chk1, an evolutionarily conserved protein kinase, has been implicated in cell cycle checkpoint control in lower eukaryotes. By gene disruption, we show that CHK1 deficiency results in a severe proliferation defect and death in embryonic stem (ES) cells, and peri-implantation embryonic lethality in mice. Through analysis of a conditional CHK1-deficient cell line, we demonstrate that ES cells lacking Chk1 have a defective G(2)/M DNA damage checkpoint in response to gamma-irradiation (IR). CHK1 heterozygosity modestly enhances the tumorigenesis phenotype of WNT-1 transgenic mice. We show that in human cells, Chk1 is phosphorylated on serine 345 (S345) in response to UV, IR, and hydroxyurea (HU). Overexpression of wild-type Atr enhances, whereas overexpression of the kinase-defective mutant Atr inhibits S345 phosphorylation of Chk1 induced by UV treatment. Taken together, these data indicate that Chk1 plays an essential role in the mammalian DNA damage checkpoint, embryonic development, and tumor suppression, and that Atr regulates Chk1.

1,707 citations


"A PIM-CHK1 signaling pathway regula..." refers background in this paper

  • ...CHK1 knockout mice are embryonic lethal (Lam et al., 2004; Liu et al., 2000), and Chk1+/– mice exhibit hematopoietic defects (Boles et al., 2010)....

    [...]

  • ...CHK1deficient blastocysts and embryonic stem cells also show severe proliferation defects as well as an impaired cell cycle checkpoint response (Liu et al., 2000; Takai et al., 2000)....

    [...]

  • ...CHK1 knockout mice are embryonic lethal (Lam et al., 2004; Liu et al., 2000), and Chk1 mice exhibit hematopoietic defects (Boles et al....

    [...]

  • ...CHK1deficient blastocysts and embryonic stem cells also show severe proliferation defects as well as an impaired cell cycle checkpoint response (Liu et al., 2000; Takai et al., 2000)....

    [...]

Journal ArticleDOI
04 Sep 2008-Nature
TL;DR: It is demonstrated that the initial activation of PLK1 is a primary function of aurora A, and that Bora/aurora-A-dependent phosphorylation is a prerequisite for PLK 1 to promote mitotic entry after a checkpoint-dependent arrest.
Abstract: Polo-like kinase-1 (PLK1) is an essential mitotic kinase regulating multiple aspects of the cell division process. Activation of PLK1 requires phosphorylation of a conserved threonine residue (Thr 210) in the T-loop of the PLK1 kinase domain, but the kinase responsible for this has not yet been affirmatively identified. Here we show that in human cells PLK1 activation occurs several hours before entry into mitosis, and requires aurora A (AURKA, also known as STK6)-dependent phosphorylation of Thr 210. We find that aurora A can directly phosphorylate PLK1 on Thr 210, and that activity of aurora A towards PLK1 is greatly enhanced by Bora (also known as C13orf34 and FLJ22624), a known cofactor for aurora A (ref. 7). We show that Bora/aurora-A-dependent phosphorylation is a prerequisite for PLK1 to promote mitotic entry after a checkpoint-dependent arrest. Importantly, expression of a PLK1-T210D phospho-mimicking mutant partially overcomes the requirement for aurora A in checkpoint recovery. Taken together, these data demonstrate that the initial activation of PLK1 is a primary function of aurora A.

664 citations


"A PIM-CHK1 signaling pathway regula..." refers background in this paper

  • ...Mac urek, L., Lindqvist, A., Lim, D., Lampson, M. A., Klompmaker, R., Freire, R., Clouin, C., Taylor, S. S., Yaffe, M. B. and Medema, R. H. (2008)....

    [...]

  • ...Since STK10, SLK and Aurora A have also been previously characterized as direct mediators of PLK1 phosphorylation at T210 (Walter et al., 2003; Mac urek et al., 2008; Seki et al., 2008), it remains to be clarified how these different kinases coordinate their activities to phosphorylate and activate PLK1....

    [...]

  • ...Since STK10, SLK and Aurora A have also been previously characterized as direct mediators of PLK1 phosphorylation at T210 (Walter et al., 2003; Mac urek et al., 2008; Seki et al., 2008), it remains to be clarified how these different kinases coordinate their activities to phosphorylate and activate…...

    [...]

Journal ArticleDOI
20 Jun 2008-Science
TL;DR: It is reported that the synergistic action of Bora and the kinase Aurora A (Aur-A) controls the G2-M transition and provides a mechanism for one of the most important yet ill-defined events in the cell cycle.
Abstract: A central question in the study of cell proliferation is, what controls cell-cycle transitions? Although the accumulation of mitotic cyclins drives the transition from the G2 phase to the M phase in embryonic cells, the trigger for mitotic entry in somatic cells remains unknown. We report that the synergistic action of Bora and the kinase Aurora A (Aur-A) controls the G2-M transition. Bora accumulates in the G2 phase and promotes Aur-A-mediated activation of Polo-like kinase 1 (Plk1), leading to the activation of cyclin-dependent kinase 1 and mitotic entry. Mechanistically, Bora interacts with Plk1 and controls the accessibility of its activation loop for phosphorylation and activation by Aur-A. Thus, Bora and Aur-A control mitotic entry, which provides a mechanism for one of the most important yet ill-defined events in the cell cycle.

574 citations


"A PIM-CHK1 signaling pathway regula..." refers background in this paper

  • ...…STK10, SLK and Aurora A have also been previously characterized as direct mediators of PLK1 phosphorylation at T210 (Walter et al., 2003; Mac urek et al., 2008; Seki et al., 2008), it remains to be clarified how these different kinases coordinate their activities to phosphorylate and activate PLK1....

    [...]

Journal ArticleDOI
TL;DR: Recent insights into the biology of PLKs will be reviewed, with an emphasis on their role in malignant transformation, and progress in the development of small-molecule PLK1 inhibitors will be examined.
Abstract: Polo-like kinase 1 (PLK1), which helps orchestrate key events in the cell cycle, has been preclinically validated as an anticancer target, whereas it seems that the closely related kinases PLK2 and PLK3 might act as tumour suppressors. Strebhardt reviews recent insights into the biology of PLKs, with emphasis on their role in malignant transformation, and examines progress in the development of small-molecule PLK1 inhibitors as anticancer drugs. The polo-like kinase 1 (PLK1) acts in concert with cyclin-dependent kinase 1–cyclin B1 and Aurora kinases to orchestrate a wide range of critical cell cycle events. Because PLK1 has been preclinically validated as a cancer target, small-molecule inhibitors of PLK1 have become attractive candidates for anticancer drug development. Although the roles of the closely related PLK2, PLK3 and PLK4 in cancer are less well understood, there is evidence showing that PLK2 and PLK3 act as tumour suppressors through their functions in the p53 signalling network, which guards the cell against various stress signals. In this article, recent insights into the biology of PLKs will be reviewed, with an emphasis on their role in malignant transformation, and progress in the development of small-molecule PLK1 inhibitors will be examined.

572 citations


"A PIM-CHK1 signaling pathway regula..." refers methods in this paper

  • ...For these experiments, costaining was performed with an antibody against PLK1, a wellestablished marker of the different mitotic steps (Strebhardt, 2010)....

    [...]

Journal ArticleDOI
TL;DR: Targeted disruption of Chk1 in mice showed that ChK1(-/-) embryos exhibit gross morphologic abnormalities in nuclei as early as the blastocyst stage, which may indicate that Chk 1 is indispensable for cell proliferation and survival through maintaining the G(2) checkpoint in mammals.
Abstract: The recent discovery of checkpoint kinases has suggested the conservation of checkpoint mechanisms between yeast and mammals. In yeast, the protein kinase Chk1 is thought to mediate signaling associated with the DNA damage checkpoint of the cell cycle. However, the function of Chk1 in mammals has remained unknown. Targeted disruption of Chk1 in mice showed that Chk1(-/-) embryos exhibit gross morphologic abnormalities in nuclei as early as the blastocyst stage. In culture, Chk1(-/-) blastocysts showed a severe defect in outgrowth of the inner cell mass and died of apoptosis. DNA replication block and DNA damage failed to arrest the cell cycle before initiation of mitosis in Chk1(-/-) embryos. These results may indicate that Chk1 is indispensable for cell proliferation and survival through maintaining the G(2) checkpoint in mammals.

521 citations


"A PIM-CHK1 signaling pathway regula..." refers background in this paper

  • ...CHK1deficient blastocysts and embryonic stem cells also show severe proliferation defects as well as an impaired cell cycle checkpoint response (Liu et al., 2000; Takai et al., 2000)....

    [...]

  • ...CHK1deficient blastocysts and embryonic stem cells also show severe proliferation defects as well as an impaired cell cycle checkpoint response (Liu et al., 2000; Takai et al., 2000)....

    [...]