Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tuberous sclerosis is an autosomal dominant disorder characterized by the development of benign growths in many tissues and organs. Linkage analysis revealed two disease-determining genes on chromosome 9 and chromosome 16. The TSC2 gene on chromosome 16 encodes a 1784-amino acid tumor suppressor protein, tuberin, that functions as a GTPase-activating protein for Rap1, a member of the superfamily of Ras-related proteins. By immunoblot analyses, we found TSC2 expression to be high in G0 as well as in early small G1 cells. Analyses after different cell synchronization procedures revealed that TSC2 mRNA and protein expression do not fluctuate throughout the cell cycle. Using inducible expression systems we further demonstrated that TSC2 expression is not affected by overexpression of the mitogenic transcription factor E2F-1 or c-Myc. Nevertheless, antisense inhibition of tuberin expression in logarithmically growing cells markedly decreased the percentage of cells in G1. Furthermore, we found that cells exposed to TSC2 antisense oligonucleotides did not undergo G0 arrest after serum withdrawal. Antisense inhibition of TSC2 expression also induced quiescent G0-arrested fibroblasts to reenter the cell cycle. Our data show for the first time that the absence of tuberin can both induce cells to pass through the G1/S transition of the eukaryotic cell cycle and prevent them from entering a quiescent state. These results have clear implications for the tumor suppressor function of TSC2. We further found that reentry into the cell cycle upon loss of TSC2 is dependent on the activity of the G1 cyclin-dependent kinases (CDKs), Cdk2 or Cdk4. Taken together with our finding that antisense inhibition of TSC2 causes up-regulation of cyclin D1 expression, these results provide the first evidence for a connection between tuberin/Rap1 and the G1 CDK-dependent regulation of the transition from G0/G1 to S phase.
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PMID:Role of the tuberous sclerosis gene-2 product in cell cycle control. Loss of the tuberous sclerosis gene-2 induces quiescent cells to enter S phase. 936 Oct 10

Tuberous sclerosis complex (TSC) is a multisystemic disorder characterized by systemic hamartomas. Although the disease-determining genes TSC1 and TSC2 have been isolated, the molecular pathogenesis of the disease is not understood. We examined cell cycle abnormalities in skin specimens and cultured cells derived from specific lesions of TSC patients with confirmed TSC1 or TSC2 mutations. None of the specimens used in this study showed loss of heterozygosity (LOH). We detected more cells positive for PCNA and fewer cells positive for MPP2 in the epidermis of TSC patients than in the epidermis of control patients without TSC. Incorporation of 5-bromo-2-deoxyuridine (BrdU) was similar in fibroblasts derived from TSC lesions and in normal human fibroblasts. These results suggest that the cell cycle of TSC cells shows a prolonged S phase. Flow cytometric analysis confirmed S phase prolongation in TSC cells. Many apoptotic cells were detected by a nick end labeling assay in both skin tissue and cultured fibroblasts derived from specific TSC lesions. Examination of cyclin levels showed increased nuclear cyclin A and cytoplasmic cyclin B and decreased nuclear cdc2 levels. We conclude that suppression of either TSC1 or TSC2 may change cyclin levels, prolong S phase and induce apoptotic cell death.
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PMID:Cells derived from tuberous sclerosis show a prolonged S phase of the cell cycle and increased apoptosis. 1175 89

The study of the molecular pathogenesis of epilepsy in tuberous sclerosis has taken on a new dimension with the identification of the TSC1 and TSC2 genes. While the development of seizures is ultimately related to mutations in one of the two genes, the mechanism underlying the genotype-phenotype relationship remains a puzzle. This chapter, presented arguments in favor of the hypothesis that abnormal cortical excitability originates in and around focal areas of structural malformations (i.e., cortical tubers and dysplasia) and that these "lesions" are the biologic consequences of tuberin and/or hamartin dysfunction. This model relies on the concept of a multistep process occurring early in cortical development whereby certain progenitor cells in the germinal layer of the ventricular zone destined for the cortex undergo inactivation of the TSC1 or TSC2 locus (Fig. 2). Immature neuroepithelial cells carrying "two-hit" mutations at either locus are believed to proliferate, migrate, and differentiate abnormally, resulting in the formation of "dysplastic" cells that are heterotopic in distribution. The pathology of the classic tuber suggests a clonal expansion of the bizarre-appearing giant cells that display incomplete, multilineage, and often ambiguous phenotype. Further, they infiltrate the six-layered structure of the cortex to form a poorly circumscribed area containing a mixture of cell types to create a highly disorganized region of a neuronal and glial network. Whether arising from the dysplastic "two-hit" target cells themselves or adjacent "innocent" bystander neurons as a result of aberrant cell-cell interaction, abnormal epileptic discharges originate from these structural abnormalities. The mechanism of how TSC1 and TSC2 inactivation causes tuber to develop is not known, but emerging experimental evidence suggests a disruption of the hamartin-tuberin "haloenzyme" in the regulation of cell size and number via the insulin signaling pathway and a p27/CDK-dependent mechanism. Biochemically, TSC1/TSC2 may associate with cytoskeletal components and vesicular adaptors in regulating sorting and trafficking of newly synthesized and recycling proteins in the post-Golgi compartments. As such, spatial and temporal localization of proteins may be affected in tuberin or hamartin-deficient neuronal cells where proper synaptic delivery of neurotransmitters plays an important role in normal cerebral function. We are in the earliest stages of understanding the role of TSC genes in epileptogenesis. To test the hypothesis outlined earlier, there is a need to create in vitro and in vivo models, as direct human experimentation is not feasible. To date, there are several rodent models of TSC, both spontaneous and recombinant strains. Unfortunately, none has consistently developed spontaneous cortical tubers, although one example was reported in an otherwise asymptomatic Eker rat (Mizuguchi et al., 2000). If the "two-hit" hypothesis is operational in tubers, as seen in other TSC lesions, it follows that radiation and chemical carcinogens should have a quantitative and qualitative effect on the development of these cerebral malformations. In preliminary experiments, we have found evidence of areas of cortical dysplasia in Eker rats irradiated early in life (Fig. 3). These dysplastic [figure: see text] cells stained positively with NeuN, consistent with the immunophenotype of cells in tubers. Alternatively, one can analyze the in vivo and in vitro characteristics of neuroprogenitor cells that are deficient of hamartin or tuberin. While homozygous mutants of TSC1 and TSC2 are lethal during midgestation, one of several techniques can be used to derive mutant neuroepithelial cells, including the procurement of -/- cells prior to embryonic deaths and subsequent cortical transplantation into syngeneic animals, development of conditional "knock outs," or chimeric mutants. These approaches, with their unique advantages and disadvantages, will be helpful in gaining insights into the development of cortical tubers and their electrophysiologic consequences.
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PMID:Tuberous sclerosis as an underlying basis for infantile spasm. 1204 Aug 99

TSC1 (tuberous sclerosis complex 1) encoding hamartin and TSC2 encoding tuberin are tumor suppressor genes responsible for the autosomal dominantly inherited disease tuberous sclerosis. These genes have been demonstrated to negatively regulate cell cycle progression, the activity of cdk2, and the degradation of the cyclin-dependent kinase inhibitor p27. To date, the underlying molecular mechanism remains elusive. Here, we show that tuberin binds to p27. Whereas tuberin also binds p27 in TSC1-negative cells, hamartin does not bind p27 without tuberin. p27 protein levels are regulated through ubiquitin-dependent degradation. Skp2 is the F-box protein, which, together with other proteins, forms an SCF (Skp1/cullin/F-box protein)-type E3 ubiquitin ligase complex whose task is to target p27 for degradation by the proteasome. We found that neither tuberin nor hamartin are in a complex with Skp2. Tuberin does not affect Skp2 protein levels, and the SCFSkp2 ubiquitin ligase does not regulate tuberin stability. But binding of tuberin to p27 sequesters p27 from Skp2 accompanied by an up-regulation of the p27 interaction with cdk2. Skp2-induced p27 degradation and cell cycle progression is abolished by tuberin's protective binding to p27. This work, the first description of the direct interaction of a tumor suppressor protein with p27, provides a molecular explanation for the effects of tuberous sclerosis complex genes on the cell cycle and demonstrates a new aspect of the SCFSkp2-mediated regulation of p27 stability.
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PMID:Tuberin binds p27 and negatively regulates its interaction with the SCF component Skp2. 1535 97

Overexpression of human IGF-1 with the bovine keratin 5 (BK5) promoter (BK5.IGF-1 transgenic mice) induces persistent epidermal hyperplasia and leads to spontaneous skin tumor formation. In previous work, PI3K and Akt activities were found to be elevated in the epidermis of BK5.IGF-1 transgenic mice compared to nontransgenic littermates. In the present study, we examined the importance of the PI3K/Akt signaling pathway in mediating the skin phenotype and the skin tumor promoting action of IGF-1 in these mice. Western blot analyses with epidermal lysates showed that signaling components downstream of PI3K/Akt were altered in epidermis of BK5.IGF-1 mice. Increased phosphorylation of GSK-3 (Ser(9/21)), TSC2(Thr(1462)), and mTOR(Ser(2448)) was observed. In addition, hypophosphorylation and increased protein levels of beta-catenin were observed in the epidermis of BK5.IGF-1 mice. These data suggested that components downstream of Akt might be affected, including cell cycle machinery in the epidermis of BK5.IGF-1 mice. Protein levels of cyclins (D1, E, A), E2F1, and E2F4 were all elevated in the epidermis of BK5.IGF-1 mice. Also, immunoprecipitation experiments demonstrated an increase in cdk4/cyclin D1 and cdk2/cyclin E complex formation, suggesting increased cdk activity in the epidermis of transgenic mice. In further studies, the PI3K inhibitor, LY294002, significantly blocked IGF-1-mediated epidermal proliferation and skin tumor promotion in DMBA-initiated BK5.IGF-1 mice. In addition, inhibition of PI3K/Akt with LY294002 reversed many of the cell cycle related changes observed in untreated transgenic animals. Collectively, the current results supported the hypothesis that elevated PI3K/Akt activity and subsequent activation of one or more downstream effector pathways contributed significantly to the tumor promoting action of IGF-1 in the epidermis of BK5.IGF-1 mice.
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PMID:Role of PI3K/Akt signaling in insulin-like growth factor-1 (IGF-1) skin tumor promotion. 1608 73

p27(Kip1) plays an important role in cell cycle regulation by inhibiting cyclin-CDK complex activity in the nucleus. p27(Kip1) is regulated by its concentration as well as by its subcellular localization. Tuberin, encoded by the tuberous sclerosis tumor suppressor gene TSC2, is a potent negative cell cycle regulator. We show herein, that tuberin induces nuclear p27 localization by inhibiting its 14-3-3-mediated cytoplasmic retention. Tuberin interferes with 14-3-3's counteracting effects on p27-mediated cell cycle arrest. Akt-mediated phosphorylation of p27, but not of tuberin, negatively regulates tuberin's potential to trigger p27 nuclear localization. In G0 cells, tuberin binds p27 triggering downregulation of p27's binding to 14-3-3 and of its cytoplasmic retention. At transition to S phase p27 is phosphorylated by Akt, tuberin/p27 complex levels are downregulated and binding of p27 to 14-3-3 increases triggering cytoplasmic retention of p27. These findings demonstrate p27 localization during the mammalian cell cycle to be under the control of the tumor suppressor tuberin.
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PMID:p27Kip1 localization depends on the tumor suppressor protein tuberin. 1747 Apr 59

The calcium/calmodulin-dependent kinase that phosphorylates and inactivates eukaryotic elongation factor 2 (eEF2 kinase; eEF2K) is subject to multisite phosphorylation, which regulates its activity. Phosphorylation at Ser359 inhibits eEF2K activity even at high calcium concentrations. To identify the kinase that phosphorylates Ser359 in eEF2K, we developed an extensive purification protocol. Tryptic mass fingerprint analysis identified it as cdc2 (cyclin-dependent kinase 1). cdc2 co-purifies with Ser359 kinase activity and cdc2-cyclin B complexes phosphorylate eEF2K at Ser359. We demonstrate that cdc2 contributes to controlling eEF2 phosphorylation in cells. cdc2 is activated early in mitosis. Kinase activity against Ser359 in eEF2K also peaks at this stage of the cell cycle and eEF2 phosphorylation is low in mitotic cells. Inactivation of eEF2K by cdc2 may serve to keep eEF2 active during mitosis (where calcium levels rise) and thereby permit protein synthesis to proceed in mitotic cells. Amino-acid starvation decreases cdc2's activity against eEF2K, whereas loss of TSC2 (a negative regulator of mammalian target of rapamycin complex 1(mTORC1)) increases it. These data closely match the control of Ser359 phosphorylation and indicate that cdc2 may be regulated by mTORC1.
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PMID:cdc2-cyclin B regulates eEF2 kinase activity in a cell cycle- and amino acid-dependent manner. 1833 51

5'AMP-activated protein kinase (AMPK) and the mammalian target of rapamycin (mTOR) are two serine/threonine protein kinases responsible for cellular energy homeostasis and translational control, respectively. Evidence suggests that these two kniases are potential targets for cancer chemotherapy against hepatocellular carcinoma (HCC). Antroquinonol that is isolated from Antrodia camphorate, a well-known Traditional Chinese Medicine for treatment of liver diseases, displayed effective anticancer activity against both HBV DNA-positive and -negative HCC cell lines. The rank order of potency against HCCs is HepG2>HepG2.2.15>Mahlavu>PLC/PRF/5>SK-Hep1>Hep3B. Antroquinonol completely abolished cell-cycle progression released from double-thymidine-block synchronization and caused a subsequent apoptosis. The data were supported by down-regulation and reduced nuclear translocation of G1-regulator proteins, including cyclin D1, cyclin E, Cdk4 and Cdk2. Further analysis showed that the mRNA expressions of the G1-regulator proteins were not modified by antroquinonol, indicating an inhibition of translational but not transcriptional levels. Antroquinonol induced the assembly of tuberous sclerosis complex (TSC)-1/TSC2, leading to the blockade of cellular protein synthesis through inhibition of protein phosphorylation including mTOR (Ser(2448)), p70(S6K) (Thr(421)/Ser(424) and Thr(389)) and 4E-BP1 (Thr(37)/Thr(46) and Thr(70)). Furthermore, the AMPK activity was elevated by antroquinonol. Compound C, a selective AMPK inhibitor, significantly reversed antroquinonol-mediated effects suggesting the crucial role of AMPK. Besides, the loss of mitochondrial membrane potential and depletion of mitochondrial content indicated the mitochondrial stress caused by antroquinonol. In summary, the data suggest that antroquinonol displays anticancer activity against HCCs through AMPK activation and inhibition of mTOR translational pathway, leading to G1 arrest of the cell-cycle and subsequent cell apoptosis.
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PMID:Antroquinonol displays anticancer potential against human hepatocellular carcinoma cells: a crucial role of AMPK and mTOR pathways. 1972 12

The mammalian target of rapamycin (mTOR) is one target of BCR-ABL fusion gene of chronic myeloid leukemia (CML). Moreover, it drives a compensatory route to Imatinib mesylate (IM) possibly involved in the progression of leukemic progenitors towards a drug-resistant phenotype. Accordingly, mTOR inhibitors are proposed for combined therapeutic strategies in CML. The major caveat in the use of mTOR inhibitors for cancer therapy comes from the induction of an mTOR-phosphatidylinositol 3 kinase (PI3k) feedback loop driving the retrograde activation of Akt. Here we show that the rapamycin derivative RAD 001 (everolimus, Novartis Institutes for Biomedical Research) inhibits mTOR and, more importantly, revokes mTOR late re-activation in response to IM. RAD 001 interferes with the assembly of both mTOR complexes: mTORC1 and mTORC2. The inhibition of mTORC2 results in the de-phosphorylation of Akt at Ser(473) in the hydrophobic motif of C-terminal tail required for Akt full activation and precludes Akt re-phosphorylation in response to IM. Moreover, RAD 001-induced inhibition of Akt causes the de-phosphorylation of tuberous sclerosis tumor suppressor protein TSC2 at 14-3-3 binding sites, TSC2 release from 14-3-3 sigma (restoring its inhibitory function on mTORC1) and nuclear import (promoting the nuclear translocation of cyclin-dependent kinase [CDK] inhibitor p27(Kip1), the stabilization of p27(Kip1) ligand with CDK2, and the G(0)/G(1) arrest). RAD 001 cytotoxicity on cells not expressing the BCR-ABL fusion gene or its p210 protein tyrosine kinase (TK) activity suggests that the inhibition of normal hematopoiesis may represent a drug side effect.
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PMID:RAD 001 (everolimus) prevents mTOR and Akt late re-activation in response to imatinib in chronic myeloid leukemia. 2001 66

Rheb is a small GTPase primarily known for activating mammalian target of rapamycin complex 1 (mTORC1) and promoting cell growth in response to insulin and nutrients (amino acids, glucose). Shortage of glucose activates adenosine 5'-monophosphate-activated protein kinase (AMPK), which induces catabolic processes that produce ATP and suppresses energy-consuming anabolic reactions. As part of the latter response, AMPK activates the TSC1-TSC2 tumor suppressor complex, which in turn inhibits Rheb, thereby reducing mTORC1 activity and consequently suppressing protein synthesis. We recently identified an mTORC1-independent Rheb-to-AMPK feedback signaling loop in Tsc2-null in vitro models of Tuberous Sclerosis Complex (TSC). In addition to activating AMPK, Rheb reduced the nuclear levels of the cyclin-dependent kinase inhibitor p27(KIP1) (p27). Importantly, Rheb-mediated repression of p27 correlated with activation of Cdk2 and cell proliferation, and with tumor formation by TSC cells. Considering that AMPK was previously reported to regulate stability and subcellular localization of p27, we hypothesize that Rheb regulates p27 in TSC cells by activating AMPK. This article discusses how Rheb-to-AMPK, and p27 signaling may impact on disease progression and treatment of TSC, including sporadic lymphangioleiomyomatosis (S-LAM) and malignancies.
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PMID:Consequences of interrupted Rheb-to-AMPK feedback signaling in tuberous sclerosis complex and cancer. 2214 93


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