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

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 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 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 Aberrant activation of cyclin A-CDK induces G2/M-phase checkpoint in human cells

Cell Cycle ◽  
2019 ◽  
Vol 19 (1) ◽  
pp. 84-96 ◽  
Author(s):  
Yasunori Akaike ◽  
Taku Chibazakura
Keyword(s):  
Cyclin A ◽  
Human Cells ◽  
M Phase

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

scholarly journals Cell Cycle-Dependent Regulation of Human DNA Polymerase α-Primase Activity by Phosphorylation

1999 ◽  
Vol 19 (1) ◽  
pp. 646-656 ◽  
Author(s):  
Christian Voitenleitner ◽  
Christoph Rehfuess ◽  
Melissa Hilmes ◽  
Lynda O’Rear ◽  
Pao-Chi Liao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Polymerase ◽  
Cyclin E ◽  
Cyclin A ◽  
Human Cells ◽  
Dependent Manner ◽  
Dna Polymerase Α ◽  
A Cell ◽  
Cycle Dependent

ABSTRACT DNA polymerase α-primase is known to be phosphorylated in human and yeast cells in a cell cycle-dependent manner on the p180 and p68 subunits. Here we show that phosphorylation of purified human DNA polymerase α-primase by purified cyclin A/cdk2 in vitro reduced its ability to initiate simian virus 40 (SV40) DNA replication in vitro, while phosphorylation by cyclin E/cdk2 stimulated its initiation activity. Tryptic phosphopeptide mapping revealed a family of p68 peptides that was modified well by cyclin A/cdk2 and poorly by cyclin E/cdk2. The p180 phosphopeptides were identical with both kinases. By mass spectrometry, the p68 peptide family was identified as residues 141 to 160. Cyclin A/cdk2- and cyclin A/cdc2-modified p68 also displayed a phosphorylation-dependent shift to slower electrophoretic mobility. Mutation of the four putative phosphorylation sites within p68 peptide residues 141 to 160 prevented its phosphorylation by cyclin A/cdk2 and the inhibition of replication activity. Phosphopeptide maps of the p68 subunit of DNA polymerase α-primase from human cells, synchronized and labeled in G1/S and in G2, revealed a cyclin E/cdk2-like pattern in G1/S and a cyclin A/cdk2-like pattern in G2. The slower-electrophoretic-mobility form of p68 was absent in human cells in G1/S and appeared as the cells entered G2/M. Consistent with this, the ability of DNA polymerase α-primase isolated from synchronized human cells to initiate SV40 replication was maximal in G1/S, decreased as the cells completed S phase, and reached a minimum in G2/M. These results suggest that the replication activity of DNA polymerase α-primase in human cells is regulated by phosphorylation in a cell cycle-dependent manner.

New alternative phosphorylation sites on the cyclin dependent kinase 1/cyclin a complex in p53-deficient human cells treated with etoposide: possible association with etoposide-induced apoptosis

APOPTOSIS ◽  
2007 ◽  
Vol 12 (10) ◽  
pp. 1847-1855 ◽  
Author(s):  
Karen Higginbottom ◽  
Ulrike Jahnke ◽  
Adrian C. Newland ◽  
Finbarr E. Cotter ◽  
Paul David Allen
Keyword(s):  
Cyclin A ◽  
Human Cells ◽  
Phosphorylation Sites ◽  
Cyclin Dependent Kinase ◽  
Induced Apoptosis

scholarly journals G2 delay induced by nitrogen mustard in human cells affects cyclin A/cdk2 and cyclin B1/cdc2-kinase complexes differently.

1993 ◽  
Vol 268 (11) ◽  
pp. 8298-8308
Author(s):  
P.M. O'Connor ◽  
D.K. Ferris ◽  
M. Pagano ◽  
G. Draetta ◽  
J. Pines ◽  
...  
Keyword(s):  
Nitrogen Mustard ◽  
Cyclin A ◽  
Cyclin B1 ◽  
Human Cells ◽  
Cdc2 Kinase ◽  
G2 Delay

Alpha-Tocopherol Protects Cultured Human Cells from the Acute Lethal Cytotoxicity of Dioxin

2002 ◽  
Vol 72 (3) ◽  
pp. 147-153 ◽  
Author(s):  
Kei-Ichi Hirai ◽  
Jie-Hong Pan ◽  
Ying-Bo Shui ◽  
Eriko Simamura ◽  
Hiroki Shimada ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Damage ◽  
Smooth Endoplasmic Reticulum ◽  
Dehydroascorbic Acid ◽  
Human Cells ◽  
Alpha Tocopherol ◽  
Cervical Cancer Cells ◽  
Cultured Human Cells ◽  
Nuclear Damage ◽  
Conjunctival Epithelial Cells

The possible protection of cultured human cells from acute dioxin injury by antioxidants was investigated. The most potent dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), caused vacuolization of the smooth endoplasmic reticulum and Golgi apparatus in cultured human conjunctival epithelial cells and cervical cancer cells. Subsequent nuclear damage included a deep irregular indentation resulting in cell death. A dosage of 30–40 ng/mL TCDD induced maximal intracellular production of H2O2 at 30 minutes and led to severe cell death (0–31% survival) at two hours. A dose of 1.7 mM alpha-tocopherol or 1 mM L-dehydroascorbic acid significantly protected human cells against acute TCDD injuries (78–97% survivals), but vitamin C did not provide this protection. These results indicate that accidental exposure to fatal doses of TCDD causes cytoplasmic free radical production within the smooth endoplasmic reticular systems, resulting in severe cytotoxicity, and that vitamin E and dehydroascorbic acid can protect against TCDD-induced cell damage.

Chromium oxidation state mapping in human cells

2003 ◽  
Vol 104 ◽  
pp. 289-292 ◽  
Author(s):  
R. Ortega ◽  
B. Fayard ◽  
M. Salomé ◽  
G. Devès ◽  
J. Susini
Keyword(s):  
Oxidation State ◽  
Human Cells ◽  
Chromium Oxidation
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