scholarly journals Dual Regulation of the Anaphase Promoting Complex in Human Cells by Cyclin A-Cdk2 and Cyclin A-Cdk1 Complexes

Cell Cycle ◽  
2006 ◽  
Vol 5 (6) ◽  
pp. 662-667 ◽  
Author(s):  
Jayashree Mitra ◽  
Greg H. Enders ◽  
Jane Azizkhan-Clifford ◽  
Kathleen L. Lengel
Keyword(s):  
Cyclin A ◽  
Human Cells ◽  
Anaphase Promoting Complex

Identifying Cyclin A/Cdk1 Substrates in Mitosis in Human Cells

2022 ◽  
pp. 175-182
Author(s):  
Ana Maria G. Dumitru ◽  
Duane A. Compton
Keyword(s):  
Cyclin A ◽  
Human Cells

scholarly journals Chk1 Phosphorylates Cdh1 to Promote SCFβTRCP-Dependent Degradation of Cdh1 During S-Phase

2019 ◽  
Author(s):  
Debjani Pal ◽  
Adrian E. Torres ◽  
Abbey L. Messina ◽  
Andrew Dickson ◽  
Kuntal De ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Feedback Loop ◽  
Cellular Proliferation ◽  
Genomic Stability ◽  
Cyclin A ◽  
S Phase ◽  
Anaphase Promoting Complex ◽  
Replication Machinery ◽  
Key Factor ◽  
Phase Entry

ABSTRACTThe interplay of the Anaphase-Promoting Complex/Cyclosome (APC/C) and Skp1-Cul1-F-box (SCF) E3 ubiquitin ligases is necessary for controlling cell cycle transitions and checkpoint responses, which are critical for maintaining genomic stability. Yet, the mechanisms underlying the coordinated activity of these enzymes are not completely understood. Recently, Cyclin A- and Plk1- mediated phosphorylation of Cdh1 was demonstrated to trigger its ubiquitination by SCFβTRCP at the G1/S transition. However, Cyclin A-Cdk and Plk1 activities peak in G2 so it is unclear why Cdh1 is targeted at G1/S but not in G2. Here, we show that phosphorylation of Cdh1 by Chk1 contributes to its recognition by SCFβTRCP, promotes efficient S-phase entry, and is important for cellular proliferation. Conversely, Chk1 activity in G2 inhibits Cdh1 accumulation. Overall, these data suggest a model whereby the rise and fall of Chk1 activity is a key factor in the feedback loop between APC/CCdh1 and the replication machinery that enhances the G1/S and S/G2 transitions, respectively.

Chromatin-bound Cdc6 persists in S and G2 phases in human cells, while soluble Cdc6 is destroyed in a cyclin A-cdk2 dependent process

2000 ◽  
Vol 113 (11) ◽  
pp. 1929-1938 ◽  
Author(s):  
D. Coverley ◽  
C. Pelizon ◽  
S. Trewick ◽  
R.A. Laskey
Keyword(s):  
Dna Replication ◽  
Protein Kinase ◽  
Mammalian Cell ◽  
Cyclin A ◽  
S Phase ◽  
Human Cells ◽  
Cell Extracts ◽  
In Vitro Replication ◽  
Initiation Of Dna Replication

Cdc6 is essential for the initiation of DNA replication in all organisms in which it has been studied. In addition, recombinant Cdc6 can stimulate initiation in G(1) nuclei in vitro. We have analysed the behaviour of recombinant Cdc6 in mammalian cell extracts under in vitro replication conditions. We find that Cdc6 is imported into the nucleus in G(1)phase, where it binds to chromatin and remains relatively stable. In S phase, exogenous Cdc6 is destroyed in a process that requires import into the nucleus and phosphorylation by a chromatin-bound protein kinase. Recombinant cyclin A-cdk2 can completely substitute for the nucleus in promoting destruction of soluble Xenopus and human Cdc6. Despite this regulated destruction, endogenous Cdc6 persists in the nucleus after initiation, although the amount falls. Cdc6 levels remain constant in G(2) then fall again before mitosis. We propose that cyclin A-cdk2 phosphorylation results in destruction of any Cdc6 not assembled into replication complexes, but that assembled proteins remain, in the phosphorylated state, in the nucleus. This process could contribute to the prevention of reinitiation in human cells by making free Cdc6 unavailable for re-assembly into replication complexes after G(1) phase.

scholarly journals The RZZ complex requires the N-terminus of KNL1 to mediate optimal Mad1 kinetochore localization in human cells

Open Biology ◽  
2015 ◽  
Vol 5 (11) ◽  
pp. 150160 ◽  
Author(s):  
Gina V. Caldas ◽  
Tina R. Lynch ◽  
Ryan Anderson ◽  
Sana Afreen ◽  
Dileep Varma ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Spindle Assembly Checkpoint ◽  
Protein S ◽  
Human Cells ◽  
Anaphase Promoting Complex ◽  
Anaphase Onset ◽  
N Terminus ◽  
Independent Mechanism ◽  
Direct Linkage ◽  
Surveillance Mechanism

The spindle assembly checkpoint is a surveillance mechanism that blocks anaphase onset until all chromosomes are properly attached to microtubules of the mitotic spindle. Checkpoint activity requires kinetochore localization of Mad1/Mad2 to inhibit activation of the anaphase promoting complex/cyclosome in the presence of unattached kinetochores. In budding yeast and Caenorhabditis elegans , Bub1, recruited to kinetochores through KNL1, recruits Mad1/Mad2 by direct linkage with Mad1. However, in human cells it is not yet established which kinetochore protein(s) function as the Mad1/Mad2 receptor. Both Bub1 and the RZZ complex have been implicated in Mad1/Mad2 kinetochore recruitment; however, their specific roles remain unclear. Here, we investigate the contributions of Bub1, RZZ and KNL1 to Mad1/Mad2 kinetochore recruitment. We find that the RZZ complex localizes to the N-terminus of KNL1, downstream of Bub1, to mediate robust Mad1/Mad2 kinetochore localization. Our data also point to the existence of a KNL1-, Bub1-independent mechanism for RZZ and Mad1/Mad2 kinetochore recruitment. Based on our results, we propose that in humans, the primary mediator for Mad1/Mad2 kinetochore localization is the RZZ complex.

scholarly journals Differential expression, localization and activity of two alternatively spliced isoforms of human APC regulator CDH1

2003 ◽  
Vol 374 (2) ◽  
pp. 349-358 ◽  
Author(s):  
Yuan ZHOU ◽  
Yick-Pang CHING ◽  
Raymond W. M. NG ◽  
Dong-Yan JIN
Keyword(s):  
Cell Cycle ◽  
Differential Expression ◽  
Cultured Cells ◽  
Cyclin A ◽  
Cyclin B1 ◽  
Anaphase Promoting Complex ◽  
Cycle Arrest ◽  
Localization Signal ◽  
Alternatively Spliced

The timely destruction of key regulators through ubiquitin-mediated proteolysis ensures the orderly progression of the cell cycle. The APC (anaphase-promoting complex) is a major component of this degradation machinery and its activation is required for the execution of critical events. Recent studies have just begun to reveal the complex control of the APC through a regulatory network involving WD40 repeat proteins CDC20 and CDH1. In the present paper, we report on the identification and characterization of human CDH1β, a novel alternatively spliced isoform of CDH1. Both CDH1α and CDH1β can bind to the APC and stimulate the degradation of cyclin B1, but they are differentially expressed in human tissues and cells. CDH1α contains a nuclear localization signal which is absent in CDH1β. Intracellularly, CDH1α appears in the nucleus whereas CDH1β is a predominantly cytoplasmic protein. The forced overexpression of CDH1α in cultured cells correlates with the reduction of nuclear cyclin A, but the steady-state amount of cyclin A does not change noticeably in CDH1β-overexpressed cells. In Xenopus embryos, ectopic overexpression of human CDH1α, but not of CDH1β, induces cell-cycle arrest during the first G1 phase at the mid-blastula transition. Taken together, our findings document the differential expression, subcellular localization and cell-cycle-regulatory activity of human CDH1 isoforms.

scholarly journals Defining pathways of spindle checkpoint silencing: functional redundancy between Cdc20 ubiquitination and p31comet

2011 ◽  
Vol 22 (22) ◽  
pp. 4227-4235 ◽  
Author(s):  
Luying Jia ◽  
Bing Li ◽  
Ross T. Warrington ◽  
Xing Hao ◽  
Shixuan Wang ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Spindle Checkpoint ◽  
Functional Redundancy ◽  
Mitotic Checkpoint ◽  
Human Cells ◽  
Unresolved Issue ◽  
Anaphase Promoting Complex ◽  
Anaphase Onset ◽  
Mitotic Checkpoint Complex

The spindle checkpoint senses unattached or improperly attached kinetochores during mitosis, inhibits the anaphase-promoting complex or cyclosome (APC/C), and delays anaphase onset to prevent aneuploidy. The mitotic checkpoint complex (MCC) consisting of BubR1, Bub3, Mad2, and Cdc20 is a critical APC/C-inhibitory checkpoint complex in human cells. At the metaphase–anaphase transition, the spindle checkpoint turns off, and MCC disassembles to allow anaphase onset. The molecular mechanisms of checkpoint inactivation are poorly understood. A major unresolved issue is the role of Cdc20 autoubiquitination in this process. Although Cdc20 autoubiquitination can promote Mad2 dissociation from Cdc20, a nonubiquitinatable Cdc20 mutant still dissociates from Mad2 during checkpoint inactivation. Here, we show that depletion of p31comet delays Mad2 dissociation from Cdc20 mutants that cannot undergo autoubiquitination. Thus both p31comet and ubiquitination of Cdc20 are critical mechanisms of checkpoint inactivation. They act redundantly to promote Mad2 dissociation from Cdc20.

scholarly journals Preventing the Degradation of Mps1 at Centrosomes Is Sufficient to Cause Centrosome Reduplication in Human Cells

2007 ◽  
Vol 18 (11) ◽  
pp. 4457-4469 ◽  
Author(s):  
Christopher Kasbek ◽  
Ching-Hui Yang ◽  
Adlina Mohd Yusof ◽  
Heather M. Chapman ◽  
Mark Winey ◽  
...  
Keyword(s):  
Phosphorylation Site ◽  
Proteasome Inhibition ◽  
Cyclin A ◽  
S Phase ◽  
Human Cells ◽  
Centrosome Duplication ◽  
Cyclin Dependent Kinase ◽  
Mitotic Spindles ◽  
Cdk2 Activity ◽  
Necessary And Sufficient

Supernumerary centrosomes promote the assembly of abnormal mitotic spindles in many human tumors. In human cells, overexpression of the cyclin-dependent kinase (Cdk)2 partner cyclin A during a prolonged S phase produces extra centrosomes, called centrosome reduplication. Cdk2 activity protects the Mps1 protein kinase from proteasome-mediated degradation, and we demonstrate here that Mps1 mediates cyclin A-dependent centrosome reduplication. Overexpression of cyclin A or a brief proteasome inhibition increases the centrosomal levels of Mps1, whereas depletion of Cdk2 leads to the proteasome-dependent loss of Mps1 from centrosomes only. When a Cdk2 phosphorylation site within Mps1 (T468) is mutated to alanine, Mps1 cannot accumulate at centrosomes or participate in centrosome duplication. In contrast, phosphomimetic mutations at T468 or deletion of the region surrounding T468 prevent the proteasome-dependent removal of Mps1 from centrosomes in the absence of Cdk2 activity. Moreover, cyclin A-dependent centrosome reduplication requires Mps1, and these stabilizing Mps1 mutations cause centrosome reduplication, bypassing cyclin A. Together, our data demonstrate that the region surrounding T468 contains a motif that regulates the accumulation of Mps1 at centrosomes. We suggest that phosphorylation of T468 attenuates the degradation of Mps1 at centrosomes and that preventing this degradation is necessary and sufficient to cause centrosome reduplication in human cells.

scholarly journals How cyclin A destruction escapes the spindle assembly checkpoint

2010 ◽  
Vol 190 (4) ◽  
pp. 501-509 ◽  
Author(s):  
Barbara Di Fiore ◽  
Jonathon Pines
Keyword(s):  
Ubiquitin Ligase ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin A ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
N Terminus ◽  
Specific Proteins ◽  
Necessary And Sufficient

The anaphase-promoting complex/cyclosome (APC/C) is the ubiquitin ligase essential to mitosis, which ensures that specific proteins are degraded at specific times to control the order of mitotic events. The APC/C coactivator, Cdc20, is targeted by the spindle assembly checkpoint (SAC) to restrict APC/C activity until metaphase, yet early substrates, such as cyclin A, are degraded in the presence of the active checkpoint. Cdc20 and the cyclin-dependent kinase cofactor, Cks, are required for cyclin A destruction, but how they enable checkpoint-resistant destruction has not been elucidated. In this study, we answer this problem: we show that the N terminus of cyclin A binds directly to Cdc20 and with sufficient affinity that it can outcompete the SAC proteins. Subsequently, the Cks protein is necessary and sufficient to promote cyclin A degradation in the presence of an active checkpoint by binding cyclin A–Cdc20 to the APC/C.

scholarly journals Cyclin A/Cdk2 complexes regulate activation of Cdk1 and Cdc25 phosphatases in human cells

Oncogene ◽  
2004 ◽  
Vol 23 (19) ◽  
pp. 3361-3367 ◽  
Author(s):  
Jayashree Mitra ◽  
Greg H Enders
Keyword(s):  
Cyclin A ◽  
Human Cells

scholarly journals Characterization of Vertebrate Cohesin Complexes and Their Regulation in Prophase

2000 ◽  
Vol 151 (4) ◽  
pp. 749-762 ◽  
Author(s):  
Izabela Sumara ◽  
Elisabeth Vorlaufer ◽  
Christian Gieffers ◽  
Beate H. Peters ◽  
Jan-Michael Peters
Keyword(s):  
Budding Yeast ◽  
Human Cells ◽  
Cyclin B ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Sister Chromatids ◽  
A Subunit ◽  
Mitotic Regulation

In eukaryotes, sister chromatids remain connected from the time of their synthesis until they are separated in anaphase. This cohesion depends on a complex of proteins called cohesins. In budding yeast, the anaphase-promoting complex (APC) pathway initiates anaphase by removing cohesins from chromosomes. In vertebrates, cohesins dissociate from chromosomes already in prophase. To study their mitotic regulation we have purified two 14S cohesin complexes from human cells. Both complexes contain SMC1, SMC3, SCC1, and either one of the yeast Scc3p orthologs SA1 and SA2. SA1 is also a subunit of 14S cohesin in Xenopus. These complexes interact with PDS5, a protein whose fungal orthologs have been implicated in chromosome cohesion, condensation, and recombination. The bulk of SA1- and SA2-containing complexes and PDS5 are chromatin-associated until they become soluble from prophase to telophase. Reconstitution of this process in mitotic Xenopus extracts shows that cohesin dissociation does neither depend on cyclin B proteolysis nor on the presence of the APC. Cohesins can also dissociate from chromatin in the absence of cyclin-dependent kinase 1 activity. These results suggest that vertebrate cohesins are regulated by a novel prophase pathway which is distinct from the APC pathway that controls cohesins in yeast.

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