Negative regulation of mitosis by wee1+, a gene encoding a protein kinase homolog
TL;DR: Fission yeast wee1- mutants initiate mitosis at half the cell size of wild type and functions as a dose-dependent inhibitor of mitosis, the first such element to be specifically identified and cloned.
About: This article is published in Cell.The article was published on 1987-05-22. It has received 953 citations till now. The article focuses on the topics: Pom1 & Wee1.
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TL;DR: Phylogenetic mapping of the conserved protein kinase catalytic domains can serve as a useful first step in the functional characterization of these newly identified family members.
Abstract: In recent years, members of the protein kinase family have been discovered at an accelerated pace. Most were first described, not through the traditional biochemical approach of protein purification and enzyme assay, but as putative protein kinase amino acid sequences deduced from the nucleotide sequences of molecularly cloned genes or complementary DNAs. Phylogenetic mapping of the conserved protein kinase catalytic domains can serve as a useful first step in the functional characterization of these newly identified family members.
4,838Â citations
TL;DR: It appears that some checkpoints are eliminated during the early embryonic development of some organisms; this fact may pose special problems for the fidelity of embryonic cell division.
Abstract: The events of the cell cycle of most organisms are ordered into dependent pathways in which the initiation of late events is dependent on the completion of early events. In eukaryotes, for example, mitosis is dependent on the completion of DNA synthesis. Some dependencies can be relieved by mutation (mitosis may then occur before completion of DNA synthesis), suggesting that the dependency is due to a control mechanism and not an intrinsic feature of the events themselves. Control mechanisms enforcing dependency in the cell cycle are here called checkpoints. Elimination of checkpoints may result in cell death, infidelity in the distribution of chromosomes or other organelles, or increased susceptibility to environmental perturbations such as DNA damaging agents. It appears that some checkpoints are eliminated during the early embryonic development of some organisms; this fact may pose special problems for the fidelity of embryonic cell division.
3,048Â citations
TL;DR: The onset of M-phase is regulated by a mechanism common to all eukaryotic cells and requires p34cdc2 dephosphorylation and association with cyclin.
Abstract: The onset of M-phase is regulated by a mechanism common to all eukaryotic cells. Entry into M-phase is determined by activation of the p34cdc2 protein kinase which requires p34cdc2 dephosphorylation and association with cyclin.
2,798Â citations
TL;DR: An improved version of the yeast two-hybrid system is developed and used to isolate human cDNAs encoding proteins able to bind p110RB to demonstrate that PP-1 alpha isoforms preferentially bind the hypophosphorylated form of p110 RB.
Abstract: The retinoblastoma protein {pll0 nB) interacts with many cellular proteins in complexes potentially important for its growth-suppressing [unction. We have developed and used an improved version of the yeast two-hybrid system to isolate human cDNAs encoding proteins able to bind pll0 RB. One clone encodes a novel type 1 protein phosphatase catalytic subunit (PP-la2), which differs from the originally defined PP-lc~ by an amino-terminal l 1-amino-acid insert. In vitro-binding assays demonstrated that PP-lc~ isoforms preferentially bind the hypophosphorylated form of p ll0 RB. Moreover, similar pll0 RB sequences are required for binding PP-lc~2 and SV40 large T antigen. Cell cycle synchrony experiments revealed that this association occurs from mitosis to early Gv The implications of these findings on the regulation of both proteins are discussed.
1,470Â citations
Cites background from "Negative regulation of mitosis by w..."
...Progression through G1 would be determined by the balance of these opposing activities in much the same way that the wee1 and m i k l protein kinases and the cdc25 protein phosphatase antagonistically regulate the activity of the Cdc2/cyclin B complex for entry into mitosis (Russell and Nurse 1987}....
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01 Jan 1994
TL;DR: It is shown that the presence of phosphatase inhibitors can have major effects on the levels of phosphorylation and the activities of the kinase extracted from cells, and the combination of okadaic acid and sodium vanadate was most effective in protecting p34cdc2 against cellular phosphatases.
Abstract: 5 The activity of cyclin-dependent protein kinase p34cdc2 is regulated by phosphorylation. In this study, we show that the presence of phosphatase inhibitors can have major effects on the levels of phosphorylation and the activities of the kinase extracted from cells. The combination of okadaic acid and sodium vanadate was most effective in protecting p34cdc2 against cellular phosphatases. In the absence of these inhibitors, p34cdc2 was dephosphorylated with an altered activity, indicating that phosphatase activities remained high during extractions. In contrast to when both inhibitors were used, lower activity of the kinase was found when only sodium vanadate was used, whereas higher activity was found in the presence of okadaic acid. Other conventional phosphatase inhibitors such as NaP, NaRS03 and glycero12-phosphate, were not effective in preventing dephosphorylation from p34cdc2 in whole cell lysates.
1,308Â citations
References
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TL;DR: A method is described for the rapid generation and cloning of deletion derivatives well-suited for the sequencing of long stretches of DNA based on two useful features of exonuclease III: processive digestion at a very uniform rate and failure to initiate digestion at DNA ends with four-base 3'-protrusions.
3,849Â citations
TL;DR: The one-step gene disruption techniques described here are versatile in that a disruption can be made simply by the appropriate cloning experiment and the resultant chromosomal insertion is nonreverting and contains a genetically linked marker.
Abstract: The one-step gene disruption techniques described here are versatile in that a disruption can be made simply by the appropriate cloning experiment. The resultant chromosomal insertion is nonreverting and contains a genetically linked marker. Detailed knowledge of the restriction map of a fragment is not necessary. It is even possible to "probe" a fragment that is unmapped for genetic functions by constructing a series of insertions and testing each one for its phenotype.
2,848Â citations
TL;DR: The rapid degradation of injected alpha- and beta-casein as well as the inverse correlation of PEST regions with intracellular stability indicate that the presence of these regions can result in the rapid intrace cellular degradation of the proteins containing them.
Abstract: The amino acid sequences of ten proteins with intracellular half-lives less than 2 hours contain one or more regions rich in proline (P), glutamic acid (E), serine (S), and threonine (T). These PEST regions are generally, but not always, flanked by clusters containing several positively charged amino acids. Similar inspection of 35 proteins with intracellular half-lives between 20 and 220 hours revealed that only three contain a PEST region. On the basis of this information, it was anticipated that caseins, which contain several PEST sequences, would be rapidly degraded within eukaryotic cells. This expectation was confirmed by red blood cell-mediated microinjection of 125I-labeled caseins into HeLa cells where they exhibited half-lives of less than 2 hours. The rapid degradation of injected alpha- and beta-casein as well as the inverse correlation of PEST regions with intracellular stability indicate that the presence of these regions can result in the rapid intracellular degradation of the proteins containing them.
2,547Â citations
1,835Â citations
TL;DR: Evidence is described showing that cdc25+ functions to counteract the activity of the mitotic inhibitor wee1+, and indicating that both mitotic control elements act independently to regulate the initiation of mitosis.
942Â citations