Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.3.16 (calcineurin)
 17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ATM kinase, when activated postnatally, exerts multiple functions to prevent the onset of ataxia-telangiectasia (AT). Using freshly isolated thymocytes from Atm-/- mice that were under stress during postnatal differentiation, we noted that thiol redox activity, as indicated by reduction of the tetrazolium MTS, and DNA turnover activity, as indicated by incorporation of [(3)H]thymidine into DNA, were both greatly increased compared with activities in thymocytes from Atm+/+ mice. This increased thymidine incorporation could be suppressed by the thiol N-acetylcysteine. In primary noncycling splenocytes, mitogens proportionally increased both the rate of [(3)H]thymidine incorporation and the rate of reduction of MTS. The mitogen-induced activities in splenocytes were not affected by ATM but were suppressed by the calcineurin-dependent inhibitor FK-506, which has no effect on these activities in thymocytes. These findings suggest that increased [(3)H]thymidine incorporation and reducing power indicate increased cell cycling in mitogenically stimulated splenocytes, whereas these two indicators represent increased FK-506-independent DNA turnover activities in thymocytes. Thus, a primary function of ATM is to activate the redox-sensitive checkpoint required for down-regulation of DNA turnover activities in developing lymphocytes. Cell-cycling checkpoints in undamaged quiescent lymphocytes are not activated by ATM with mitogenic stimulation. ATM may suppress abnormal DNA turnover and the resultant oncogenesis by regulating cellular thiol redox pathways.
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PMID:The ataxia-telangiectasia gene product may modulate DNA turnover and control cell fate by regulating cellular redox in lymphocytes. 1134 81

Polo-like kinase 3 (Plk3, previously termed Prk) contributes to regulation of M phase of the cell cycle (Ouyang, B., Pan, H., Lu, L., Li, J., Stambrook, P., Li, B., and Dai, W. (1997) J. Biol. Chem. 272, 28646-28651). Plk3 physically interacts with Cdc25C and phosphorylates this protein phosphatase predominantly on serine 216 (Ouyang, B., Li, W., Pan, H., Meadows, J., Hoffmann, I., and Dai, W. (1999) Oncogene 18, 6029-6036), suggesting that the role of Plk3 in mitosis is mediated, at least in part, through direct regulation of Cdc25C. Here we show that ectopic expression of a kinase-active Plk3 (Plk3-A) induced apoptosis. In response to DNA damage, the kinase activity of Plk3 was rapidly increased in an ATM-dependent manner, whereas that of Plk1 was markedly inhibited. Recombinant Plk3 phosphorylated in vitro a glutathione S-transferase fusion protein containing p53, but not glutathione S-transferase alone. Recombinant Plk1 also phosphorylated p53 but on residues that differed from those targeted by Plk3. Co-immunoprecipitation and pull-down assays demonstrated that Plk3 physically interacted with p53 and that this interaction was enhanced upon DNA damage. In vitro kinase assays followed by immunoblotting showed that serine 20 of p53 was a target of Plk3. Furthermore, expression of a kinase-defective Plk3 mutant (Plk3(K52R)) resulted in significant reduction of p53 phosphorylation on serine 20, which was correlated with a decrease in the expression of p21 and with a concomitant increase in cell proliferation. These results strongly suggest that Plk3 functionally links DNA damage to cell cycle arrest and apoptosis via the p53 pathway.
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PMID:Plk3 functionally links DNA damage to cell cycle arrest and apoptosis at least in part via the p53 pathway. 1155 30

Ionizing radiation (IR) is known to activate multiple signaling pathways, resulting in diverse stress responses including apoptosis, cell cycle arrest, and gene induction. IR-activated cell cycle checkpoints are regulated by Ser/Thr phosphorylation, so we tested to see if protein phosphatases were targets of an IR-activated damage-sensing pathway. Jurkat cells were subjected to IR or sham radiation followed by brief (32)P metabolic labeling. Nuclear extracts were subjected to microcystin affinity chromatography to recover phosphatases, and the proteins were analyzed by two-dimensional gel electrophoresis. Protein sequencing revealed that the microcystin-bound proteins with the greatest reduction in (32)P intensity following IR were the alpha and delta isoforms of protein phosphatase 1 (PP1). Both of these PP1 isoforms contain an Arg-Pro-Ile/Val-Thr-Pro-Pro-Arg sequence near the C terminus, a known site of phosphorylation by Cdc/Cdk kinases, and phosphorylation attenuates phosphatase activity. In wild-type Jurkat cells or ataxia telangiectasia (AT) cells that are stably transfected with full-length ATM kinase, IR resulted in net dephosphorylation of this site in PP1 and produced activation of PP1. However, in AT cells that are deficient in ATM, IR failed to induce dephosphorylation or activation of PP1. IR-induced PP1 activation in the nucleus may be a critical component in an ATM-mediated pathway controlling checkpoint activation.
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PMID:Ionizing radiation activates nuclear protein phosphatase-1 by ATM-dependent dephosphorylation. 1220 91

Eukaryotic cells respond to DNA damage and stalled replication forks by activating protein kinase-mediated signaling pathways that promote cell cycle arrest and DNA repair. A central target of the cell cycle arrest program is the Cdc25A protein phosphatase. Cdc25A is required for S-phase entry and dephosphorylates tyrosine-15 phosphorylated Cdk1 (Cdc2) and Cdk2, positive regulators of cell division. Cdc25A is unstable during S-phase and is degraded through the ubiquitin-proteasome pathway, but its turnover is enhanced in response to DNA damage. Although basal and DNA-damage-induced turnover depends on the ATM-Chk2 and ATR-Chk1 pathways, how these kinases engage the ubiquitin ligase machinery is unknown. Here, we demonstrate a requirement for SCFbeta-TRCP in Cdc25A turnover during an unperturbed cell cycle and in response to DNA damage. Depletion of beta-TRCP stabilizes Cdc25A, leading to hyperactive Cdk2 activity. SCFbeta-TRCP promotes Chk1-dependent Cdc25A ubiquitination in vitro, and this involves serine 76, a known Chk1 phosphorylation site. However, recognition of Cdc25A by beta-TRCP occurs via a noncanonical phosphodegron in Cdc25A containing phosphoserine 79 and phosphoserine 82, sites that are not targeted by Chk1. These data indicate that Cdc25A turnover is more complex than previously appreciated and suggest roles for an additional kinase(s) in Chk1-dependent Cdc25A turnover.
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PMID:SCFbeta-TRCP links Chk1 signaling to degradation of the Cdc25A protein phosphatase. 1468 Dec 6

Phosphorylated histone H2AX (gamma-H2AX) forms foci over large chromatin domains surrounding double-stranded DNA breaks (DSB). These foci recruit DSB repair proteins and dissolve during or after repair is completed. How gamma-H2AX is removed from chromatin remains unknown. Here, we show that protein phosphatase 2A (PP2A) is involved in removing gamma-H2AX foci. The PP2A catalytic subunit [PP2A(C)] and gamma-H2AX coimmunoprecipitate and colocalize in DNA damage foci and PP2A dephosphorylates gamma-H2AX in vitro. The recruitment of PP2A(C) to DNA damage foci is H2AX dependent. When PP2A(C) is inhibited or silenced by RNA interference, gamma-H2AX foci persist, DNA repair is inefficient, and cells are hypersensitive to DNA damage. The effect of PP2A on gamma-H2AX levels is independent of ATM, ATR, or DNA-PK activity.
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PMID:gamma-H2AX dephosphorylation by protein phosphatase 2A facilitates DNA double-strand break repair. 1631 Mar 92

ATM (ataxia telangiectasia mutated) is required for the early response to DNA-damaging agents such as ionizing radiation (IR) that induce DNA double-strand breaks. Cells deficient in ATM are extremely sensitive to IR. It has been shown that IR induces immediate phosphorylation of ATM at Ser(1981), leading to catalytic activation of the protein. We recently isolated a novel BRCA1-associated protein, BAAT1 (BRCA1-associated protein required for ATM activation-1), by yeast two-hybrid screening and found that BAAT1 also binds to ATM, localizes to double-strand breaks, and is required for Ser(1981) phosphorylation of ATM. Small interfering RNA-mediated stable or transient reduction of BAAT1 resulted in decreased phosphorylation of both ATM at Ser(1981) and CHK2 at Thr(68). Treatment of BAAT1-depleted cells with okadaic acid greatly restored phosphorylation of ATM at Ser(1981), suggesting that BAAT1 is involved in the regulation of ATM phosphatase. Protein phosphatase 2A-mediated dephosphorylation of ATM was partially blocked by purified BAAT1 in vitro. Significantly, acute loss of BAAT1 resulted in increased p53, leading to apoptosis. These results demonstrate that DNA damage-induced ATM activation requires a coordinated assembly of BRCA1, BAAT1, and ATM.
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PMID:ATM activation by ionizing radiation requires BRCA1-associated BAAT1. 1645 82

Most of the known breast cancer susceptibility genes (BRCA1, BRCA2, CHEK2 and ATM) are involved in the damage response pathway. Other members of this pathway are therefore good candidates for additional breast cancer susceptibility genes. ATR, along with ATM, plays a central role in DNA damage recognition and Chk1 relays checkpoint signals from both ATR and ATM. PPP2R1B and PPP2R5B code for subunits of protein phosphatase 2A (PP2A), which regulates autophosphorylation of ATM. In addition, EIF2S6/Int-6, which was originally identified as a common integration site for the mouse mammary tumour virus in virally induced mouse mammary tumours, is a candidate breast cancer susceptibility gene because of its putative role in maintaining chromosome stability. To investigate the role of ATR, CHK1, PPP2R1B, PPP2R5B and EIF2S6/Int-6, we carried out mutation analysis of these genes in the index cases from non-BRCA1/BRCA2 breast cancer families. We also screened sporadic breast tumours for somatic mutations in PPP2R1B and PPP2R5B. Although we identified many novel variants, we found no evidence that highly penetrant germline mutations in these five genes contribute to familial breast cancer susceptibility.
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PMID:Mutation analysis of five candidate genes in familial breast cancer. 1718 32

ATM and Rad3-related (ATR) is a regulatory kinase that, when activated by hydroxyurea, UV, or human immunodeficiency virus-1 Vpr, causes cell cycle arrest through Chk1-Ser(345) phosphorylation. We demonstrate here that of these three agents only Vpr requires protein phosphatase type 2A (PP2A) to activate ATR for Chk1-Ser(345) phosphorylation. A requirement for PP2A by Vpr was first shown with the PP2A-specific inhibitor okadaic acid, which reduced Vpr-induced G(2) arrest and Cdk1-Tyr(15) phosphorylation. Using small interference RNA to down-regulate specific subunits of PP2A indicated that the catalytic beta-isoform PP2A(Cbeta) and the A regulatory alpha-isoform PP2A(Aalpha) are involved in the G(2) induction, and these downregulations decreased the Vpr-induced, ATR-dependent phosphorylations of Cdk1-Tyr(15) and Chk1-Ser(345). In contrast, the same down-regulations had no effect on hydroxyurea- or UV-activated ATR-dependent Chk1-Ser(345) phosphorylation. Vpr and hydroxyurea/UV all induce ATR-mediated gammaH2AX-Ser(139) phosphorylation and foci formation, but down-regulation of PP2A(Aalpha) or PP2A(Cbeta) did not decrease gammaH2AX-Ser(139) phosphorylation by any of these agents or foci formation by Vpr. Conversely, H2AX down-regulation had little effect on PP2A(Aalpha/Cbeta)-mediated G(2) arrest and Chk1-Ser(345) phosphorylation by Vpr. The expression of vpr increases the amount and phosphorylation of Claspin, an activator of Chk1 phosphorylation. Down-regulation of either PP2A(Cbeta) or PP2A(Aalpha) had little effect on Claspin phosphorylation, but the amount of Claspin was reduced. Claspin may then be one of the phosphoproteins through which PP2A(Aalpha/Cbeta) affects Chk1 phosphorylation when ATR is activated by human immunodeficiency virus-1 Vpr.
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PMID:Phosphatase type 2A-dependent and -independent pathways for ATR phosphorylation of Chk1. 1721 May 76

ATM is a central mediator of the cellular response to the DNA damage produced by ionizing radiation. We recently showed that protein phosphatase 1 (PP1) is activated by ATM. Because Nek2 is activated by autophosphorylation, and because its dephosphorylation is catalyzed by PP1, we asked if the radiation damage signal to Nek2 was mediated by PP1. Overexpression of Nek2 induces premature centrosome splitting probably by phosphorylating centrosome cohesion proteins C-Nap1 and Rootletin. In this study, we show isoform specificity of PP1 binding and regulation of Nek2. Although both PP1alpha and PP1gamma coimmunoprecipitated with Nek2, only PP1alpha regulated Nek2 function. Ionizing radiation inhibited Nek2 activity, and this response was dependent on ATM and on PP1 binding to Nek2 and coincident with Thr(320) dephosphorylation of PP1. Radiation-induced inhibition of centrosome splitting was abrogated in cells expressing Nek2 mutated in the PP1-binding motif outside the kinase domain. Conversely, cells depleted of PP1alpha by small interfering RNA showed enhanced centrosome splitting and loss of radiation-induced inhibition of centrosome splitting. The identification of a PP1-specific isoform mediating a checkpoint response opens up the possibility of selectively targeting phosphatases as novel radiation sensitizers.
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PMID:Protein phosphatase-1alpha regulates centrosome splitting through Nek2. 1728 41

The phosphorylation state of the tumor suppressor protein BRCA1 is tightly associated with its functions including cell cycle control and DNA repair. Protein kinases involved in the DNA damage checkpoint control, such as ATM, ATR, and hCds1/Chk2, have been shown to phosphorylate and activate BRCA1 upon DNA damage. We reported previously that protein phosphatase 1alpha (PP1alpha) interacts with and dephosphorylates hCds1/Chk2-phosphorylated BRCA1. This study demonstrates the identification of a PP1-binding motif 898KVTF901 in BRCA1. Mutation or deletion of critical residues in this PP1-binding motif substantially reduces the interaction between BRCA1 and PP1alpha. PP1alpha can also dephosphorylate ATM and ATR phosphorylation sites in BRCA1 and may serve as a general regulator for BRCA1 phosphorylation. Unlike wild-type BRCA1, expression of the PP1 non-binding mutant BRCA1 protein in BRCA1-deficient cells failed to enhance survival after DNA damage. Taken together, these results suggest that interaction with PP1alpha is important for BRCA1 function.
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PMID:Identification and functional characterization of a PP1-binding site in BRCA1. 1760 99


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