SKP1 Connects Cell Cycle Regulators to the Ubiquitin Proteolysis Machinery through a Novel Motif, the F-Box (original) (raw)
Developmental Cell, 2003
chromosome congression (den Elzen and Pines, 2001), Stanford University School of Medicine destruction of the anaphase inhibitor securin is required 300 Pasteur Drive for chromosome disjunction and the progression to ana-Stanford, California 94305 phase (Cohen-Fix et al., 1996), and destruction of cyclin B is required for mitotic exit (Murray et al., 1989). How is it that specific APC substrates are not destroyed in Summary S and G2 and yet are sequentially destroyed in M phase to coordinate specific mitotic events? Progression through mitosis occurs because cyclin Surveillance mechanisms or checkpoints that restrict B/Cdc2 activation induces the anaphase promoting specific mitotic events until the completion of earlier complex (APC) to cause cyclin B destruction and mievents contribute to the timing of APC substrate detotic exit. To ensure that cyclin B/Cdc2 does not prestruction. In the best-known example, the spindle asmaturely activate the APC in early mitosis, there must sembly checkpoint (Hoyt et al., 1991; Li and Murray, be a mechanism delaying APC activation. Emi1 is a 1991) restrains the activation of securin destruction to protein capable of inhibiting the APC in S and G2. We delay chromosome separation at anaphase (Cohen-Fix show here that Emi1 is phosphorylated by Cdc2, and et al., 1996) until all chromosomes have established a on a DSGxxS consensus site, is subsequently recogbipolar attachment to the mitotic spindle and congress nized by the SCF TrCP/Slimb ubiquitin ligase and destroyed, to the midline. In the case of the spindle assembly thus providing a delay for APC activation. Failure of checkpoint, the dynamic assembly of a series of factors TrCP-dependent Emi1 destruction stabilizes APC (including Mad1, Mad2, Bub1, and BubR1) that bind and substrates and results in mitotic catastrophe including inactivate Cdc20 (Fang et al., 1998; Tang et al., 2001) centrosome overduplication, potentially explaining delays securin destruction in either normal prometamitotic deficiencies in Drosophila Slimb/TrCP muphase (Geley et al., 2001) or when the spindle is distants. We hypothesize that Emi1 destruction relieves rupted by microtubule depolymerizing agents (Yu, 2002). a late prophase checkpoint for APC activation. Recent evidence shows that the protein Emi1 (Reimann et al., 2001a) inhibits APC Cdc20 activity in interphase Introduction in the Xenopus embryo to prevent destruction of mitotic cyclins. Emi1 can inhibit the APC by blocking the ability To successfully initiate mitosis and bipolar spindle asof the Cdc20 substrate adaptor to bind to its substrates sembly, cells must coordinate the maturation of spindle (Reimann et al., 2001b), although additional mechanisms poles with condensation of the chromosomes, nuclear are possible. Emi1 can also regulate the APC Cdh1 comenvelope breakdown, and prometaphase movements plex in somatic cells to cause the stabilization of S phase of condensed chromosomes on the assembling mitotic regulators including cyclin A at the G1-S transition (Hsu spindle. As the spindle matures, congression of chromoet al., 2002; Reimann et al., 2001b). A similar mechanism somes to the midzone at metaphase triggers chromofor stabilizing cyclin A by the Emi1 homolog Rca1 has some segregation, anaphase movements, and ultibeen demonstrated in Drosophila (Grosskortenhaus and mately cytokinesis. The timing of these complex events Sprenger, 2002). By inhibiting Cdh1 and Cdc20, Emi1 requires the sequential activation of cyclin-dependent defines an interval of cyclin stability and APC inactivity kinases (Cdks) by cyclin A and cyclin B (Minshull et al., from late G1 to early mitosis (Hsu et al., 2002). At the 1990), as well as activation of other mitotic regulatory beginning of this interval, Emi1 is upregulated in rekinases including members of the Aurora, Polo, and sponse to the E2F transcription factor (Hsu et al., 2002). NimA-related (Nek) kinase families. Cyclin B/Cdc2 also At the end of this interval, the Emi1 protein is ubiquitiactivates a critical E3 ubiquitin ligase, the anaphase nated and destroyed (Reimann et al., 2001a) close to promoting complex/cyclosome or APC (Irniger et al., the time when cyclin A is destroyed (Hsu et al., 2002). 1995; King et al., 1995; Lahav-Baratz et al., 1995; Tugen-The ubiquitination enzymes required for Emi1 destrucdreich et al., 1995; reviewed in Harper et al., 2002; Petion have not been identified, although destruction reters, 2002). The APC directs the assembly of multiubiquiquires Cdc2 and not the APC (Reimann et al., 2001a, tin chains on specific substrates and their subsequent 2001b)
A Late Mitotic Regulatory Network Controlling Cyclin Destruction inSaccharomyces cerevisiae
Molecular Biology of the Cell, 1998
Exit from mitosis requires the inactivation of mitotic cyclin-dependent kinase–cyclin complexes, primarily by ubiquitin-dependent cyclin proteolysis. Cyclin destruction is regulated by a ubiquitin ligase known as the anaphase-promoting complex (APC). In the budding yeast Saccharomyces cerevisiae, members of a large class of late mitotic mutants, including cdc15, cdc5, cdc14, dbf2, and tem1, arrest in anaphase with a phenotype similar to that of cells expressing nondegradable forms of mitotic cyclins. We addressed the possibility that the products of these genes are components of a regulatory network that governs cyclin proteolysis. We identified a complex array of genetic interactions among these mutants and found that the growth defect in most of the mutants is suppressed by overexpression of SPO12, YAK1, and SIC1 and is exacerbated by overproduction of the mitotic cyclin Clb2. When arrested in late mitosis, the mutants exhibit a defect in cyclin-specific APC activity that is accompanied by high Clb2 levels and low levels of the anaphase inhibitor Pds1. Mutant cells arrested in G1 contain normal APC activity. We conclude that Cdc15, Cdc5, Cdc14, Dbf2, and Tem1 cooperate in the activation of the APC in late mitosis but are not required for maintenance of that activity in G1.
Defective in Mitotic Arrest 1 (Dma1) Ubiquitin Ligase Controls G1 Cyclin Degradation
Journal of Biological Chemistry, 2012
Background: Dma ubiquitin ligases control the cell cycle in diverse organisms. In humans, these enzymes act as tumor suppressors that prevent aberrant mitosis. Results: Dma1 targets the cyclin Pcl1 for destruction. Conclusion: Dma1 ubiquitin ligase activity controls stability of G 1 cyclins. Significance: Pcl1 is the second reported substrate for Dma1 enzymes. Uncovering new Dma1 substrates could help to elucidate cellular functions of these enzymes. * This work was supported by Spanish Ministry of Science and Innovation Grant BFU 2009-09278, now known as the Ministry of Economy and Competitiveness. □ S This article contains supplemental data. 1 Supported by a postgraduate Junior Faculty fellowship from the Universitat Internacional de Catalunya and l'Obra Social la Caixa.
Biochemical Journal, 2007
The ubiquitin-conjugating enzyme Cdc34 (cell division cycle 34) plays an essential role in promoting the G 1 -S-phase transition of the eukaryotic cell cycle and is phosphorylated in vivo. In the present study, we investigated if phosphorylation regulates Cdc34 function. We mapped the in vivo phosphorylation sites on budding yeast Cdc34 (yCdc34; Ser 207 and Ser 216 ) and human Cdc34 (hCdc34 Ser 203 , Ser 222 and Ser 231 ) to serine residues in the acidic tail domain, a region that is critical for Cdc34's cell cycle function. CK2 (protein kinase CK2) phosphorylates both yCdc34 and hCdc34 on these sites in vitro. CK2-mediated phosphorylation increased yCdc34 ubiquitination activity towards the yeast Saccharomyces cerevisiae Sic1 in vitro, when assayed in the presence of its cognate SCF Cdc4 E3 ligase [where SCF is Skp1 (S-phase kinase-associated protein 1)/cullin/F-box]. Similarly, mutation of the yCdc34 phosphorylation sites to alanine, aspartate or glutamate residues altered Cdc34-SCF Cdc4 -mediated Sic1 ubiquitination activity. Similar results were obtained when yCdc34's ubiquitination activity was assayed in the absence of SCF Cdc4 , indicating that phosphorylation regulates the intrinsic catalytic activity of Cdc34. To evaluate the in vivo consequences of altered Cdc34 activity, wild-type yCdc34 and the phosphosite mutants were introduced into an S. cerevisiae cdc34 deletion strain and, following synchronization in G 1 -phase, progression through the cell cycle was monitored. Consistent with the increased ubiquitination activity in vitro, cells expressing the phosphosite mutants with higher catalytic activity exhibited accelerated cell cycle progression and Sic1 degradation. These studies demonstrate that CK2-mediated phosphorylation of Cdc34 on the acidic tail domain stimulates Cdc34-SCF Cdc4 ubiquitination activity and cell cycle progression.
Role for cyclin-dependent kinase 2 in mitosis exit
Current Biology, 2001
Activity of the cyclin-dependent kinase (cdk) 1-cyclin B 1-cyclin B activity [1]. Exit from mitosis depends on the inactivation of the complex by the complex, also called M phase-promoting factor (MPF), is crucial for mitosis. MPF is required for spindle assembly degradation of cyclin B [2]. Cdk2 is also active during mitosis [3, 4]. In Xenopus egg extracts, cdk2 and chromosome condensation, whereas MPF downregulation by ubiquitin-mediated proteolysis of cyclin B is is primarily in complex with cyclin E, which is stable [5]. At the end of mitosis, downregulation of cdk2-needed to complete mitosis [1, 2]. Sister chromatid separation also requires ubiquitin-mediated proteolysis of securcyclin E activity is accompanied by inhibitory phosphorylation of cdk2 [6]. Here, we show that ins, which inhibit anaphase. The degradation of securins and mitotic cyclins relies upon the cell cycle-regulated cdk2-cyclin E activity maintains cdk1-cyclin B during mitosis. At mitosis exit, cdk2 is inactivated activity of the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C). This appears to be stimulated prior to cdk1. The loss of cdk2 activity follows and depends upon an increase in protein kinase A (PKA) by cdk1-dependent phosphorylation [2, 7-9]. In addition, at least two WD-40 repeat-containing proteins bind APC/C activity. Prematurely inactivating cdk2 advances the time of cyclin B degradation and cdk1 and are required for its action [2]. In Xenopus egg extracts, the Cdc20/Fizzy homolog is required for APC/C-mediated
Cell Cycle, 2013
For ordered mitotic progression, various proteins have to be regulated by an ubiquitin ligase, the anaphase-promoting complex or cyclosome (ApC/C) with appropriate timing. Recent studies have implied that the activity of ApC/C also contributes to release of mitotic checkpoint complexes (MCCs) from its target Cdc20 in the process of silencing the spindle assembly checkpoint (SAC). Here we describe a temperature-sensitive mutant (ubc11-P93L) in which cell cycle progression is arrested at mitosis. the mutant grows normally at the restrictive temperature when SAC is inactivated, suggesting that the arrest is not due to abnormal spindle assembly, but rather due to prolonged activation of SAC. Supporting this notion, MCCs remain bound to ApC/C even when SAC is satisfied. the ubc11 + gene encodes one of the two e2 enzymes required for progression through mitosis in fission yeast. Remarkably, Slp1 (a fission yeast homolog of Cdc20), which is degraded in an ApC/C-dependent manner, stays stable throughout the cell cycle in the ubc11-P93L mutant lacking the functional SAC. other ApC/C substrates, in contrast, were degraded on schedule. We have also found that a loss of Ubc4, the other e2 required for progression through mitosis, does not affect the stability of Slp1. We propose that each of the two e2 enzymes is responsible for collaborating with ApC/C for a specific set of substrates, and that Ubc11 is responsible for regulating Slp1 with ApC/C for silencing the SAC.
The anaphase-promoting complex: a key factor in the regulation of cell cycle
Oncogene, 2005
Events controlling cell division are governed by the degradation of different regulatory proteins by the ubiquitin-dependent pathway. In this pathway, the attachment of a polyubiquitin chain to a substrate by an ubiquitin-ligase targets this substrate for degradation by the 26S proteasome. Two different ubiquitin ligases play an important role in the cell cycle: the SCF (Skp1/Cullin/ F-box) and the anaphase-promoting complex (APC). In this review, we describe the present knowledge about the APC. We pay particular attention to the latest results concerning APC structure, APC regulation and substrate recognition, and we discuss the implication of these findings in the understanding the APC function.
Current Biology, 1998
Background: Following chromosome segregation in anaphase, ubiquitindependent degradation of mitotic cyclins contributes to the exit from mitosis. A key step in this process is catalyzed by a ubiquitin-protein ligase known as the anaphase-promoting complex (APC), the regulation of which is poorly understood. The Polo-related protein kinase Cdc5 in Saccharomyces cerevisiae might encode a regulator of the APC, because cdc5 mutant cells arrest with a late mitotic phenotype similar to that observed in cells with defective cyclin destruction.