The Plk3-Cdc25 Circuit

The Plk3-Cdc25 circuit

David L Myer1, El Mustapha Bahassi1 and Peter J Stambrook*,1

1Department of Cell Biology, Neurobiology and Anatomy, University of Cincinnati College of Medicine, 3125 Eden Avenue, Cincinnati, OH 45267, USA

Polo-like kinases (Plks) are key regulators of the cell between cell-cycle phases. The CDK is the catalytic cycle, especially in the G2 phase and mitosis. They are member of the complex, which associates with a incorporated into signaling networks that regulate many regulatory cyclin whose expression is dependent on the aspects of the cell cycle, including but not limited to cell-cycle phase. One of the more studied CDK/cyclin centrosome maturation and separation, mitotic entry, complexes is the Cdk1 (Cdc2)/cyclin B1 complex. The chromosome segregation, mitotic exit, and cytokinesis. initiation of CDK activity requires both phosphoryla- The Plks have well conserved 30-amino-acid elements, tion by the Cdk-activating kinase kinase on Thr161 designated polo boxes (PBs), located in their carboxyl- (Poon et al., 1993; Solomon et al., 1993) and removal of termini, which with their flanking regions constitute a inhibitory phosphates from Thr14 and Tyr15 on the functional Polo-box domain (PBD). Members of the Plk Cdk1. The Myt1 and Wee1 kinases phosphorylate family exist in a variety of organisms including Polo in Thr14 and Tyr15, respectively (Russell and Nurse, Drosophila melanogaster; Cdc5 in Saccharomyces cerevi- 1987; McGowan and Russell, 1993; Mueller et al., siae; Plo1 in Schizosaccharomyces pombe; Plx1 in 1995; Liu et al., 1997), and thereby block ATP binding Xenopus laevis; and Plk1, Snk/Plk2, Fnk/Prk/Plk3, to CDKs and inhibit phosphate transfer to CDK-bound and Sak in mammals. Polo, Cdc5, and Plo1 are essential substrates (Atherton-Fessler et al., 1993: Booher et al., for viability. The Plks can be separated into two groups 1997). according to their functions. The first group (Polo, Cdc5, The Cdc25C phosphatase, whose activity is central to plo1, Plx1, and Plk1) primarily performs mitotic func- mitotic entry, is a dual-specificity phosphatase that tions, whereas the second group (Plk2 and Plk3) appears directly dephosphorylates and activates the Cdk1/cyclin to have additional functions during the G1, S, and B kinase complex, leading to the initiation of mitosis. G2 phases of the cell cycle. Several contributions to this The Cdc25C phosphatase is activated at the G2/M issue will discuss different aspects of Plk involvement in transition by phosphorylation on serine and threonine cell-cycle regulation. This review, therefore, will focus on residues. One of the kinases capable of phosphorylating the role of Plk3 in regulating Cdc25 phosphatase function and activating Cdc25 is Cdk1, suggesting the existence and its effect on the cell cycle. of a positive feedback loop. Izumi and Maller (1995) Oncogene (2005) 24, 299–305. doi:10.1038/sj.onc.1208278 have shown that in interphase Xenopus egg extracts with no detectable Cdk1 and Cdk2, treatment with the Keywords: Polo-like kinases; Cdc25 phosphatases; phosphatase inhibitor microcystin activated a distinct phosphorylation; cell-cycle regulation kinase that could phosphorylate and activate Cdc25. Microcystin also induced other mitotic events such as chromosome condensation and nuclear envelope breakdown in extracts containing less than 5% of the mitotic level of Cdk1 kinase activity. These findings implicate a Introduction kinase other than Cdk1 and Cdk2 that can activate Cdc25 in vivo and suggest that this kinase may also The temporal and spatial order of cell division is phosphorylate M-phase substrates even in the absence maintained by the activity of cyclin-dependent kinases of a Cdk1 kinase (Izumi and Maller, 1995). The Xenopus (CDKs) (Nigg, 1993, 1995; Nurse et al., 1998; Zhou and polo-like kinase (Plk) Plx1 associates with and phos- Elledge, 2000) whose activity is modulated indirectly by phorylates the amino-terminal domain of Cdc25. When extracellular environmental signals and inhibited by recombinant Plx1 phosphorylates Cdc25 in a purified DNA damage checkpoint signals (Hartwell and system, Plx1 stimulates cdc25’s activity and results in a Weinert, 1989; Morgan, 1995; Zhou and Elledge, strong reaction with MPM-2, a monoclonal antibody 2000). Mammalian cells express at least six members directed at mitotic phosphoproteins. These studies of the CDK family, which form complexes with their indicate that Plx1 may participate in the control of respective cyclin partners to control the transitions mitotic progression through the regulation of Cdc25 activity (Kumagai and Dunphy, 1996). Subsequently, Maller’s group very elegantly demonstrated that Plx1 is *Correspondence: PJ Stambrook; E-mail: [email protected] activated in vivo during oocyte maturation with the same Plk3 and Cdc25 DL Myer et al 300 kinetics as Cdc25C. Microinjection of wild-type Plx1 APC to ubiquitinate cyclin B (Kotani et al., 1998). More into Xenopus oocytes accelerates the rate of activation of recently, studies have also demonstrated that polo kinase Cdc25C and Cdk1/cyclin B. Conversely, microinjection activity plays a pivotal role in the separation of sister of either an antibody against Plx1 or a kinase-dead chromatids during mitosis (Alexandru et al., 2001). The version of Plx1 significantly inhibits the activation of yeast Plk, Cdc5, is required to phosphorylate the cohesin Cdc25C and Cdk1/cyclin B. This effect can be reversed subunit Scc1, thereby facilitating its cleavage by separase by injection of active Cdc25C, indicating that Plx1 is at the onset of anaphase (Alexandru et al., 2001). upstream of Cdc25C (Qian et al., 1998). Consistent with this last finding, Plx1 is also a key Since these initial reports implicating Plx1 in Cdc25 regulator for the cleavage-independent dissociation of activation, other polo kinases, mainly human polo-like cohesin from chromosomes in Xenopus (Losada et al., kinase 1 and 3 (Plk1 and Plk3), have been shown to be 2002). In mammals, Plks promote spindle formation at involved in interactions with Cdc25 (Roshak et al., 2000; G2/M, activate APC as cells exit from mitosis, are Bahassi et al., 2004). The following review outlines our required for cytokinesis, and are part of the DNA view of the mammalian Plk3 and its role in mammalian damage checkpoint-signaling cascade in G2/M. Plk1 has Cdc25 phosphatases regulation. numerous mitotic functions including the regulation of cyclin B localization. Plk4, also designated Sak, is an essential Plk, which is required for the destruction of Plks and cell-cycle regulation cyclin B by the APC and for exit from mitosis in the postgastrulation embryo (Hudson et al., 2001). Plk2 and Progress through the various stages of the cell cycle is Plk3 are expressed as immediate-early transcripts; how- facilitated and controlled by multiple signaling pathways. ever, Plk3 is expressed throughout the cell cycle while Among the many regulatory enzymes that contribute to Plk2 is only detected in G1 (Simmons et al., 1992; these processes are the Plks, which are evolutionarily Donohue et al., 1995; Chase et al., 1998; Ma et al., 2003). conserved between yeast (Cdc5), Drosophila (Polo), In contrast to the other Plks, Plk3 cannot be over- Xenopus (Plx), and mammals (Plk), and which regulate expressed without inducing toxic effects, either through several mitotic events. Mammalian cells express four loss of cellular adhesion or induction of apoptosis (Conn members of the Plk family, Plk1, 2, 3, and 4. They are et al., 2000). Here, we propose that one or more Plks may important contributors to several cell-cycle events and play several additional roles in other phases of the cell are identified by their carboxy-terminal polo box (PB) cycle. Given the role of Cdc25A in regulating the G1/S motifs, which are responsible for subcellular localization transition, and recent studies on the function of Cdc25A and substrate recognition (Nigg, 1998; Roshak et al., in S phase, we expect that understanding how Plk3 2000; Toyoshima-Morimoto et al., 2001; Golan et al., regulates these phosphatases will soon reveal an ex- 2002; Sutterlin et al., 2001; Elia et al., 2003; Cheng et al., panded role for Plk3 and the other Plks. 2003). Genetic and biochemical approaches with several organisms indicate that Plks regulate diverse cellular events at various cell-cycle stages. The Xenopus Plx1 is a The Plk3 critical trigger that initiates the onset of the G2/M transition Kumagai and Dunphy (1996), and appears to In addition to its various functions during the entry into be part of the Cdk1/cyclin B amplification loop in meiotic and exit from mitosis, Plk3 also appears to be associated maturation of Xenopus oocytes (Karaiskou et al., 2004). with Golgi breakdown since ectopic expression of Plk3 Plx1 also participates in the destruction of targets of the induces fragmentation of Golgi stacks (Ruan et al., anaphase-promoting complex (APC) to mediate mitotic 2004). Xie et al. (2004) examined functional as well as exit in the early embryonic cell cycle (Brassac et al., 2000). physical interactions between Plk3 and MEK1/ERK Genetic studies in Drosophila and yeast indicate that Plks and suggested that Plk3 may be a downstream mediator function in centrosome assembly and separation during of MEK1 in the signaling cascade leading to Golgi the formation of the bipolar spindle. Polo mutants in fragmentation. Furthermore, Wang et al. (2002) re- Drosophila display hypercondensed chromosomes, ported that Plk3 may play an important role in monopolar spindles, disorganized spindle poles, and they regulating microtubule dynamics and centrosomal manifest abnormal chromosome segregation (Llamazares function, and that deregulated expression of Plk3 results et al., 1991). The Schizosaccharomyces pombe plo1À in cell-cycle arrest and apoptosis. This observation is mutant displays similar phenotypes, such as the forma- consistent with a previous report showing that over- tion of monopolar spindles and a failure in septum expression of Plk3 in mammalian cells suppresses formation after nuclear division (Ohkura et al., 1995), proliferation and inhibits colony formation, and that and the budding yeast Plk homolog, Cdc5, seems to play overexpression of Plk3 induces chromatin condensation an important role in actin ring formation and cytokinesis and apoptosis (Conn et al., 2000). In addition to its (Song and Lee, 2001). Other studies in yeast show that nuclear functions, Plk3 clearly has cytoplasmic roles the APC-dependent destruction of Clb2 (cyclin B) given its involvement in Golgi breakdown (Ruan et al., depends on Cdc5 activity (Charles et al., 1998; Jaspersen 2004). Furthermore, Holtrich et al. (2000) have shown et al., 1998; Shirayama et al., 1998). In mammalian cells, that deregulated Plk3 expression disrupts the cellular F- Plk1 phosphorylates at least three components of the actin network and induces a spherical morphology. In APC, and in an in vitro-reconstituted system, it activates addition, Plk3 binds to the Ca2 þ /integrin-binding

Oncogene Plk3 and Cdc25 DL Myer et al 301 protein Cib in two-hybrid analyses and in co-immuno- which dephosphorylates and activates the Cdk1/cyclin B precipitation assays (Kauselmann et al., 1999), and both complex (Gautier and Maller, 1991; Izumi et al., 1992; proteins colocalize in mammalian cells. The homology Kumagai and Dunphy, 1992; Hoffmann et al., 1993). of Cib with calmodulin and with calcineurin B suggests Cdc25C is predominantly cytoplasmic during the G1 that Cib might be a Plk regulatory subunit (Holtrich and S phases in a variety of cell lines (Dalal et al., 1999; et al., 2000). Lastly, plk3 is an early inducible gene since Graves et al., 2000). Its exclusion from the nucleus is plk3 mRNA expression in cultured cell lines is rapidly dependent on the export protein CRM1 as suggested by activated by mitogens (Li et al., 1996). Clinically, plk3 its nuclear accumulation following treatment with mRNA expression appears to be downregulated in a leptomycin B (Graves et al., 2001). Human Cdc25C is majority (26 of 35) of primary head and neck squamous phosphorylated on serine 216 (S216) and is bound to 14- cell carcinomas investigated compared with adjacent 3-3 proteins throughout interphase (Peng et al., 1997). uninvolved tissues from the same patients (Dai et al., Phosphorylation of S216 is required for 14-3-3 binding, 2000). It is apparent that Plk3 has multiple functions in and mutation of this serine to alanine abrogates its cellular homeostasis. One key function is the role of Plk3 interaction with 14-3-3 (Peng et al., 1997). Intrinsic in the cell cycle and its participation in activating the nuclear import and export signals, together with 14-3-3 Cdc25 phosphatases. In this review, we highlight the binding, appear to regulate coordinately the intracellu- role of Plk3 in Cdc25 phosphatase regulation. lar trafficking of Cdc25C (Graves et al., 2001). When Cdc25C enters the nucleus, it is bound by Pin1, which inactivates Cdc25C phosphatase activity. When Cdc25C The Cdc25 phosphatases is phosphorylated by Xenopus Plx and Cdk1, however, the association with Pin1 increases the phosphatase The cdc25 gene was first identified as a regulator of the activity of Cdc25C (Stukenberg et al., 2001). cell cycle in yeast (Hartwell et al., 1974; Nurse, 1975). Subsequently, homologues of the protein have been identified in most eukaryotic unicellular and multi- Plks in Cdc25C regulation cellular organisms examined. There are three cdc25 genes, cdc25A, B, and C, in the human genome (Gould The interaction between Plks and the Cdc25 phospha- et al., 1990; Galaktionov and Beach, 1991), which tases was initially described in Xenopus. The Xenopus encode proteins 470–566 residues in length, each of Plk, Plx, is required for small oocytes to re-enter meiosis which is composed of two main domains. The amino- in response to progesterone (Karaiskou et al., 2004). terminal portion appears to be a regulatory domain Plx was the first Plk shown to be required for the involved in the modulation of enzymatic activity. It has G2/M transition. The Cdk1/cyclin B1 complex several phosphorylation sites, binding sites for protein increases Plx activation to promote Cdc25 activation partners, alternatively spliced domains, and functionally and create a positive feedback loop (Abrieu et al., 1998). important domains such as nuclear localization se- Anti-Plx1 antibodies block Cdc25 activity and the quences and nuclear exclusion sequences (NES). The consequent Tyr15 dephosphorylation of cdk1/cyclin B three CDC25 proteins are more highly related in the (Qian et al., 1998). Work in starfish embryos indicates carboxy-terminal half that contains the catalytic domain that Plk1 may be more important for the inhibition of than in the amino-half of the protein. This includes a Myt1, a cdk1/cyclin B complex inhibitor. The starfish Cys-(X)5-Arg motif that is conserved in all dual- embryo Plk1 is activated downstream of cdk1/cyclin B specificity phosphatases, and the CH2A and CH2B in order to inhibit the activity of Myt1 in meiosis II and homology motifs that are also found in MAP kinase meiotic re-entry (Okano-Uchida et al., 2003). The phosphatases (Keyse and Ginsburg, 1993). Crystal- budding yeast polo kinase Cdc5 binds and phosphor- lographic data, however, show conformational differ- ylates the Wee1 kinase homolog, Swe1 (Bartholomew ences in the carboxy-terminal regions of the three et al., 2001). CDC25 proteins, suggesting that their catalytic proper- We have shown that Plk3 phosphorylates serine 191, ties may not be identical (Fauman et al., 1998; Reynolds and to a lesser extent serine 198, within the NES to et al., 1999; Wilborn et al., 2001). The Cdc25 promote the nuclear accumulation of Cdc25C, and that phosphatases are essential regulators of cell-cycle substitution of serine 191 with an alanine abrogates transitions. In lower eukaryotes, Cdc25 is an activator Cdc25C’s nuclear localization (Bahassi et al., 2004). of the G2/M transition by virtue of its ability to Further compelling evidence linking Plk3 to Cdc25C dephosphorylate the Cdk1 protein kinase (see Draetta accumulation in the nucleus derives from overexpression and Eckstein, 1997 for a review), while Cdc25 B and C of Plk3. Transient transfection of cells with wild-type appear to regulate progression from G2 to M phase Plk3 but not with a kinase dead Plk3 results in a marked (Honda et al., 1993; Mailand et al., 2002). increase of Cdc25C in the nuclear fraction following cell fractionation. This nuclear accumulation is inhibited when Plk3 is suppressed by siRNA, implicating the Cdc25C phosphatase involvement of Plk3 in the nuclear accumulation of Cdc25C (Bahassi et al., 2004). Toyoshima-Morimoto Entry into mitosis depends on phosphorylation and et al. (2002) reported that Plk1 phosphorylates Cdc25C activation of the dual-specificity phosphatase Cdc25C, on serine 198 to promote its nuclear localization.

Oncogene Plk3 and Cdc25 DL Myer et al 302 Replacement of serine 198 with either an alanine or a 1999). Although it is expressed continuously, Cdc25A phosphomimetic aspartic acid, however, does not result undergoes frequent turnover throughout the cell cycle. in loss of Cdc25C from the nucleus (Bahassi et al., 2004). Its abundance is regulated by post-translational mod- It is likely that disruption of the NES, which contains ification, which either stabilizes it or promotes its serine 198, may be sufficient to retain Cdc25C in the degradation (Mailand et al., 2002). Consequently, nucleus. Since the NES is located in close proximity to Cdc25A has two major peaks of accumulation during the 14-3-3/cyclin B binding domain on Cdc25C, another cell-cycle progression, one in G1 and the other in S/G2 possibility is that phosphorylation of serines 191 and (Bernardi et al., 2000). 198 within the NES inhibits 14-3-3 binding and During S phase and G2, the level of Cdc25A is promotes cyclin B binding. A previous report had regulated by the ubiquitin ligase SCF (Skp1/Cullin/F- argued that Plk3 phosphorylates Cdc25C on serine 216, box) (Donzelli et al., 2002). The Cdc25A is phosphory- a result which is inconsistent with the ability of Plk3 to lated on Ser82 and Ser88, which mediate binding to the complement a cdc5ts mutant (Ouyang et al., 1997), but SCF/beta-TrCP complex to promote ubiquitin ligation which attributed to a possible role for Plk3 in down- and Cdc25A degradation (Busino et al., 2003). Another regulating cdk1 (p34cdc2/cdk1) activity during late mitosis ubiquitin ligase APC/C (anaphase promoting complex/ since considerable Plk3 activity remains during mitosis cyclosome), in complex with the cdh1 subunit, regulates (Ouyang et al., 1997). the level of Cdc25A at mitotic exit and during G1 An additional relationship between the Plks and the (Donzelli et al., 2002). Residual Cdc25A remaining from Cdk1/cyclin B complex was described by Toyoshima- mitosis is degraded in G1 in response to APC-mediated Morimoto et al. (2001). This group reported that Plk1 ubiquitylation. To promote the G1/S transition, tran- phosphorylates serine 147 on cyclin B in the middle of scription of cdc25A is enhanced, and the APC is its NES and targets it to the nucleus during prophase. inactivated. They suggested that polo kinases might play a central In addition to the role of Cdc25A to maintain proper role in coordinating the nuclear translocation of both cell-cycle progression, its levels are acutely responsive to Cdc25C and its target Cdk1/cyclin B to the nucleus. The DNA damage. Damage to DNA promotes Cdc25A timing of this translocation may be essential for degradation, which in turn arrests the cells at the G1/S coordinating M-phase events. transition to allow DNA repair. In response to DNA Despite recent progress, our understanding of the damage, Chk1 and Chk2 kinases phosphorylate Ser123 overlapping and unique roles of the Cdc25 phosphatases on Cdc25A to initiate its ubiquitylation and proteo- and their activating kinases at various cell-cycle transi- some-mediated degradation (Mailand et al., 2000; Falck tions remains incomplete. It is somewhat enigmatic, et al., 2001). In the absence of DNA damage, Chk1 however, that homozygous disruption of Cdc25C in phosphorylates Thr507 on Cdc25A to facilitate Cdc25A mice yields healthy animals whose cells suffer no mitotic interaction with 14-3-3 (Chen et al., 2003). It is likely defect, have no alteration of Cdk1 phosphorylation and that this interaction maintains Cdc25A in an inactive respond normally to DNA damage (Chen et al., 2001). form until it is needed to promote the full activation of It is possible that Cdc25A may compensate for the Cdk1/cyclin B. absence of Cdc25C in these animals since Cdc25A can Once it was learned that Cdc25A was stabilized bind and activate Cdk1-cyclin B, and can accelerate cell during mitosis, the kinases that target Cdc25A and its division when overexpressed (Chen et al., 2001). role in mitosis became a focus of research interest. Prior Conversely, when Cdc25A is underexpressed, mitotic to mitosis, the Cdk1/cyclin B complex is activated by entry is delayed (Mailand et al., 2002). Cdc25B (Lammer et al., 1998). This activation allows Cdk1/cyclin B to promote Cdk1-mediated phosphorylation of Ser17 and Ser115 of Cdc25A, which stabilizes Cdc25A phosphatase Cdc25A and prevents its degradation (Mailand et al, 2002). Cdk1, however, is not alone in its capacity to Cdc25A regulates entry of cells into S phase by depho- phosphorylate and potentially stabilize Cdc25A at sphorylating Thr14 and Tyr15 on Cdk2/cyclin E and is mitosis. When the drug roscovitine, a Cdk1 and Cdk2 required for mitotic progression. Overexpression of kinase inhibitor, was used to inhibit Cdk1 and Cdk2 Cdc25A promotes a more rapid S-phase entry, while during mitosis, mutants of Cdc25A with altered Cdk1 microinjection of anti-Cdc25A antibodies and transfec- phosphorylation sites were still phosphorylated. This tion of Cdc25A antisense oligonucleotides both inhibit finding led the authors to speculate that a Plk that was S-phase entry (Hoffmann et al., 1994). Similarly, insensitive to the drug might be responsible for Cdc25A antisense oligonucleotides directed at cdc25a mRNA stability in mitosis (Mailand et al., 2002). inhibited Cdk1/cyclin B activation and mitotic entry (Mailand et al., 2002). Cdc25A is regulated at transcriptional and post- Plks in Cdc25A regulation translational levels. The transcription factors E2F1, E2F4, and c-Myc promote the expression of Cdc25A in While most Plks are upregulated and activated at the mid- to late G1 (Galaktionov et al., 1996; Vigo et al., onset of mitosis, the pattern of expression and activation 1999), and its activity progressively increases from the of Plk3 remains unchanged throughout cell cycle G1/S transition until mitosis (Blomberg and Hoffmann, (Bahassi et al., 2002). This observation led us to

Oncogene Plk3 and Cdc25 DL Myer et al 303 speculate about a possible role for Plk3 in regulating Cdc25A or Cdc25C remains unknown. One model other stages of the cell cycle, mainly the G1/S transition suggests that Plk3 may mediate binding of cyclin B to through the control of Cdc25A function. In this context, the carboxy-terminus of Cdc25A with consequent we have identified two Plk3-mediated phosphorylation displacement of 14-3-3, and a similar mechanism may sites within the carboxy-terminus of Cdc25A (unpub- apply to Cdc25B since it also acts as a substrate for Plk3 lished data). Both sites overlap with the cyclin B binding (Ouyang et al., 1999). site on Cdc25A. The potential physiological relevance of phosphorylating these sites includes modulation of Cdc25A association with 14-3-3 or cyclin B, and Conclusions possible regulation of Cdc25A stability. The protein levels of Cdc25A are stabilized in prometaphase during Based on a BLAST search (Altschul et al., 1990), the which a high level of active Cdk1/cyclin B complex is Polo-box domain (PBD) appears to be a domain that is present (Mailand et al., 2002). Subsequently, Cdk1/ unique to Plks and absent from other proteins. Since it is cyclin B activity decreases concomitant with a decrease not a modular domain represented in other proteins, its in Cdc25A (Mailand et al., 2002). These observations function may be involved in activities such as intracel- suggest that Cdc25A is required to maintain active lular localization and the intramolecular regulation of Cdk1/cyclin B in the presence of the Myt1 and Wee1 Plk kinase activity. Recent resolution of the structure of inhibitory kinases until the onset of metaphase and that the PBD (Leung et al., 2002; Cheng et al., 2003) opens the activity of Plk3 may be required to facilitate the up the possibility of defining targeted mutations to interaction between cyclin B and Cdc25A until that probe cellular function and the opportunity to design time. peptidomimics that might inhibit the function of Plks in The phosphorylation of Cdc25A by Plk3 may play yet tumor cells. It is worth noting that microinjection of another role. The cyclin B binding site on Cdc25A is in anti-Plk1 antibodies severely impairs the ability of cells close proximity to a 14-3-3 binding site for which the to divide, but distinguishes between transformed and residue T507 is essential. Chen et al. (2003) have nontransformed cells (Lane and Nigg, 1996). HeLa cells speculated that Chk1 phosphorylates T507 on Cdc25A responded to Plk1 inhibition by mitotic catastrophe throughout the cell cycle. This phosphorylation main- (Cogswell et al., 2000). In contrast, nontransformed tains 14-3-3 binding to Cdc25A and keeps Cdc25A in an Hs68 cells arrest primarily as single mononucleated cells inactive state. This type of inhibitory mechanism in G2 (Cogswell et al., 2000). Presumably, these involving 14-3-3 has been studied for Cdc25B. The differences in cell behavior reflect a difference in Cdc25A and Cdc25B phosphatases share the same stringency for the requirement of Plk1 function at the residues responsible for binding cyclin B and 14-3-3 to G2/M transition. This differential response between their carboxy-termini. The 14-3-3 protein binds as a transformed and nontransformed cells to Plk1 inhibition dimer to sites within the amino-terminus and the is suggestive of Plk1 and other Plks as selective targets carboxy-terminus of Cdc25B, pulling the two termini for potential anticancer therapeutic purposes. together and preventing access to Cdc25B’s catalytic domain and nuclear export signal (Giles et al., 2003). When 14-3-3 is not bound to the amino-terminus, the Acknowledgements catalytic site is open and the nuclear export signal is also We apologize to the many authors whose original work could not be cited owing to space limitations. We are grateful to the exposed. The binding of 14-3-3 to Ser323 affects the National Institutes of Health, NIH Grants, R01 CA90934, activity and localization of Cdc25B (Giles et al., 2003). UO1 ES011038, and P30 ES05652 and The Center for Other 14-3-3 isoforms have also been shown to affect the Environmental Genetics at the University of Cincinnati function of Cdc25A (Chen et al., 2003) and Cdc25C (CEG Center Grant P30-ES06096) for research support. DM (Margolis et al., 2003), but how 14-3-3 is displaced from was a trainee on NIH training grant T32 ES07250.

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