Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P43146 (tumour suppressor)
 5,935 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The tumour suppressor p53 protein integrates multiple signals regulating cell cycle progression and apoptosis. This regulation is mediated by several kinases that phosphorylate specific residues in the different functional domains of the p53 molecule. The human VRK1 protein is a new kinase related to a poxvirus kinase, and more distantly to the casein kinase 1 family. We have characterized the biochemical properties of human VRK1 from HeLa cells. VRK1 has a strong autophosphorylating activity in several Ser and Thr residues. VRK-1 phosphorylates acidic proteins, such as phosvitin and casein, and basic proteins such as histone 2b and myelin basic protein. Because some transcription factors are regulated by phosphorylation, we tested as substrates the N-transactivation domains of p53 and c-Jun fused to GST. Human c-Jun is not phosphorylated by VRK1. VRK1 phosphorylates murine p53 in threonine 18. This threonine is within the p53 hydrophobic loop (residues 13-23) required for the interaction of p53 with the cleft of its inhibitor mdm-2. The VRK1 C-terminus domain (residues 268-396) that contains a nuclear localization signal targets the protein to the nucleus, as determined by using fusion proteins with the green fluorescent protein. We conclude that VRK1 is an upstream regulator of p53 that belongs to a new signalling pathway.
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PMID:The human vaccinia-related kinase 1 (VRK1) phosphorylates threonine-18 within the mdm-2 binding site of the p53 tumour suppressor protein. 1095 72

The tumour suppressor protein, PTEN (phosphatase and tensin homologue deleted on chromosome 10) is a member of the mixed function, serine/threonine/tyrosine phosphatase subfamily of protein phosphatases. Its physiological substrates, however, are primarily 3-phosphorylated inositol phospholipids, which are products of phosphoinositide 3-kinases. PTEN thus antagonizes PI 3-kinase-dependent signalling pathways, which explains to a large extent its tumour suppressor status. We have examined the kinetic behaviour, substrate specificity and regulation of PTEN both in vitro and in a variety of cellular models. Although PTEN can utilize both phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P(3)] and its water-soluble headgroup, inositol 1,3,4,5-tetrakisphosphate, as substrates, it displays classical features of interfacial catalysis, which greatly favour the lipid substrate (by as much as 1000-fold as judged by K(cat)/K(m) values). Expression of PTEN in U87 cells (which lack endogenous PTEN) and measuring the levels of all known 3-phosphorylated lipids suggests that phosphatidylinositol 3,4-bisphosphate and PtdIns(3,4,5)P(3) are both substrates, but that phosphatidylinositol 3-phosphate and phosphatidylinositol 3,5-bisphosphate are not. PTEN binds to several PDZ-domain-containing proteins via a consensus sequence at its extreme C-terminus. Disruption of targeting to PDZ-domain proteins selectively blocks some PTEN functions, but not others, suggesting the existence of spatially localized, functionally dedicated pools of signalling lipids. We have also shown recently that PTEN expression is controlled at the transcriptional level and is profoundly upregulated by peroxisome proliferator activated receptor gamma agonists, thereby providing possible implications for these drugs in diabetes, inflammation and cancer.
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PMID:Antagonism of PI 3-kinase-dependent signalling pathways by the tumour suppressor protein, PTEN. 1170 86

Peutz-Jeghers syndrome is an inherited cancer syndrome, which results in a greatly increased risk of developing tumours in those affected. The causative gene encodes a nuclear-localized protein kinase, termed LKB1, which is predicted to function as a tumour suppressor. The mechanism by which LKB1 is regulated in cells is not known, and nor have any of its physiological substrates been identified. Recent studies have demonstrated that LKB1 is phosphorylated in cells. As a first step towards identifying the roles that phosphorylation of LKB1 play, we have mapped the residues that are phosphorylated in human embryonic kidney (HEK)-293 cells, as well as the major in vitro autophosphorylation sites. We demonstrate that LKB1 expressed in HEK-293 cells, in addition to being phosphorylated at Ser(431), a previously characterized phosphorylation site, is also phosphorylated at Ser(31), Ser(325) and Thr(366). Incubation of wild-type LKB1, but not a catalytically inactive mutant, with manganese-ATP in vitro resulted in the phosphorylation of LKB1 at Thr(336) as well as at Thr(366). We were unable to detect autophosphorylation at Thr(189), a site previously claimed to be an LKB1 autophosphorylation site. A catalytically inactive mutant of LKB1 was phosphorylated at Ser(31) and Ser(325) in HEK-293 cells to the same extent as the wild-type enzyme, indicating that LKB1 does not phosphorylate itself at these residues. We show that phosphorylation of LKB1 does not directly affect its nuclear localization or its catalytic activity in vitro, but that its phosphorylation at Thr(336), and perhaps to a lesser extent at Thr(366), inhibits LKB1 from suppressing cell growth.
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PMID:Identification and characterization of four novel phosphorylation sites (Ser31, Ser325, Thr336 and Thr366) on LKB1/STK11, the protein kinase mutated in Peutz-Jeghers cancer syndrome. 1185 58

The direct mechanism by which the serine/threonine kinase Akt (also known as protein kinase B (PKB)) regulates cell growth is unknown. Here, we report that Drosophila melanogaster Akt/PKB stimulates growth by phosphorylating the tuberous sclerosis complex 2 (Tsc2) tumour suppressor and inhibiting formation of a Tsc1-Tsc2 complex. We show that Akt/PKB directly phosphorylates Drosophila Tsc2 in vitro at the conserved residues, Ser 924 and Thr 1518. Mutation of these sites renders Tsc2 insensitive to Akt/PKB signalling, increasing the stability of the Tsc1-Tsc2 complex within the cell. Stimulating Akt/PKB signalling in vivo markedly increases cell growth/size, disrupts the Tsc1-Tsc2 complex and disturbs the distinct subcellular localization of Tsc1 and Tsc2. Furthermore, all Akt/PKB growth signals are blocked by expression of a Tsc2 mutant lacking Akt phosphorylation sites. Thus, Tsc2 seems to be the critical target of Akt in mediating growth signals for the insulin signalling pathway.
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PMID:Akt regulates growth by directly phosphorylating Tsc2. 1222 18

The serine/threonine protein kinase LKB1 functions as a tumour suppressor, and mutations in this enzyme lead to the inherited Peutz-Jeghers cancer syndrome. We previously found that LKB1 was phosphorylated at Thr-366 in vivo, a residue conserved in mammalian, Xenopus and Drosophila LKB1, located on a C-terminal non-catalytic moiety of the enzyme. Mutation of Thr-366 to Ala or Asp partially inhibited the ability of LKB1 to suppress growth of G361 melanoma cells, but did not affect LKB1 activity in vitro or LKB1 localization in vivo. As a first step in exploring the role of this phosphorylation further, we have generated a phosphospecific antibody specifically recognizing LKB1 phosphorylated at Thr-366 and demonstrate that exposure of cells to ionizing radiation (IR) induced a marked phosphorylation of LKB1 at Thr-366 in the nucleus. Thr-366 lies in an optimal phosphorylation motif for the phosphoinositide 3-kinase-like kinases DNA-dependent protein kinase (DNA-PK), ataxia telangiectasia mutated kinase (ATM) and ataxia telangiectasia-related kinase (ATR), which function as sensors for DNA damage in cells and mediate cellular responses to DNA damage. We demonstrate that both DNA-PK and ATM efficiently phosphorylate LKB1 at Thr-366 in vitro and provide evidence that ATM mediates this phosphorylation in vivo. This is based on the finding that LKB1 is not phosphorylated in a cell line lacking ATM in response to IR, and that agents which induce cellular responses via ATR in preference to ATM poorly induce phosphorylation of LKB1 at Thr-366. These observations provide the first link between ATM and LKB1 and suggest that ATM could regulate LKB1.
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PMID:Ionizing radiation induces ataxia telangiectasia mutated kinase (ATM)-mediated phosphorylation of LKB1/STK11 at Thr-366. 1223 50

The tumour suppressor p53 is important in the cell decision to either arrest cell cycle progression or induce apoptosis in response to a variety of stimuli. p53 post-translational modifications and association with other proteins have been implicated in the regulation of its stability and transcriptional activities. Here we report that, on DNA damage, p53 interacts with Pin1, a peptidyl-prolyl isomerase, which regulates the function of many proteins involved in cell cycle control and apoptosis. The interaction is strictly dependent on p53 phosphorylation, and requires Ser 33, Thr 81 and Ser 315. On binding, Pin1 generates conformational changes in p53, enhancing its transactivation activity. Stabilization of p53 is impaired in UV-treated Pin1(-/-) cells owing to its inability to efficiently dissociate from Mdm2. As a consequence, a reduced p53-dependent response was detected in Pin1(-/-) cells, and this correlates with a diminished transcriptional activation of some p53-regulated genes. Our results suggest that, following stress-induced phosphorylation, p53 needs to form a complex with Pin1 and to undergo a conformational change to fulfil its biological roles.
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PMID:The prolyl isomerase Pin1 reveals a mechanism to control p53 functions after genotoxic insults. 1239 40

The tumour suppressor p53 is a tetrameric protein that is phosphorylated in its BOX-I transactivation domain by checkpoint kinase 2 (CHK2) in response to DNA damage. CHK2 cannot phosphorylate small peptide fragments of p53 containing the BOX-I motif, indicating that undefined determinants in the p53 tetramer mediate CHK2 recognition. Two peptides derived from the DNA-binding domain of p53 bind to CHK2 and stimulate phosphorylation of full-length p53 at Thr 18 and Ser 20, thus identifying CHK2-docking sites. CHK2 can be fully activated in trans by the two p53 DNA-binding-domain peptides, and can phosphorylate BOX-I transactivation-domain fragments of p53 at Thr 18 and Ser 20. Although CHK2 has a basal Ser 20 kinase activity that is predominantly activated towards Thr 18, CHK1 has constitutive Thr 18 kinase activity that is predominantly activated in trans towards Ser 20. Cell division cycle 25C (CDC25C) phosphorylation by CHK2 is unaffected by the p53 DNA-binding-domain peptides. The CHK2-docking site in the BOX-V motif is the smallest of the two CHK2 binding sites, and mutating certain amino acids in the BOX-V peptide prevents CHK2 activation. A database search identified a p53 BOX-I-homology motif in p21(WAF1) and although CHK2 is inactive towards this protein, the p53 DNA-binding-domain peptides induce phosphorylation of p21(WAF1) at Ser 146. This provides evidence that CHK2 can be activated allosterically towards some substrates by a novel docking interaction, and identify a potential regulatory switch that may channel CHK2 into distinct signalling pathways in vivo.
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PMID:Allosteric effects mediate CHK2 phosphorylation of the p53 transactivation domain. 1289 1

Cell numbers are regulated by a balance between processes of proliferation and apoptosis (programmed cell death). Proper regulation in a cell requires an accurate co-ordination between these two processes. Indeed, it has recently been found that dysregulation of cell cycle progression is essential for the initiation of apoptosis. Retinoblastoma protein (RB) is an important tumour suppressor and a cell cycle regulator. Most recent studies suggest that RB also plays a regulatory role in the process of apoptosis. During the onset of apoptosis, the hyperphosphorylated form of RB (p120/hyper) is converted to a hypophosphorylated form (p115/hypo), which is mediated by a specific protein-serine/ threonine phosphatase activity. The p115/hypo/RB may play an active role in the regulation of apoptosis. Accompanied by the endonucleosomal fragmentation of DNA, the newly formed p115/hypo/RB is immediately cleaved by a protease that has properties of the interleukin-1beta-converting enzyme family. By contrast, the unphosphorylated form of RB (p110/unphos) remains uncleaved during apoptosis. Further studies suggest that p110/unphos/RB functions as an inhibitor of apoptosis. Therefore, a balance between RB phosphatases and kinases and consequent RB phosphorylation status may be important for the determination of cellular fate.
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PMID:Putative roles of retinoblastoma protein in apoptosis. 1469 99

We recently demonstrated that the LKB1 tumour suppressor kinase, in complex with the pseudokinase STRAD and the scaffolding protein MO25, phosphorylates and activates AMP-activated protein kinase (AMPK). A total of 12 human kinases (NUAK1, NUAK2, BRSK1, BRSK2, QIK, QSK, SIK, MARK1, MARK2, MARK3, MARK4 and MELK) are related to AMPK. Here we demonstrate that LKB1 can phosphorylate the T-loop of all the members of this subfamily, apart from MELK, increasing their activity >50-fold. LKB1 catalytic activity and the presence of MO25 and STRAD are required for activation. Mutation of the T-loop Thr phosphorylated by LKB1 to Ala prevented activation, while mutation to glutamate produced active forms of many of the AMPK-related kinases. Activities of endogenous NUAK2, QIK, QSK, SIK, MARK1, MARK2/3 and MARK4 were markedly reduced in LKB1-deficient cells. Neither LKB1 activity nor that of AMPK-related kinases was stimulated by phenformin or AICAR, which activate AMPK. Our results show that LKB1 functions as a master upstream protein kinase, regulating AMPK-related kinases as well as AMPK. Between them, these kinases may mediate the physiological effects of LKB1, including its tumour suppressor function.
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PMID:LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1. 1497 52

Mutations in the LKB1 tumour suppressor threonine kinase cause the inherited Peutz-Jeghers cancer syndrome and are also observed in some sporadic cancers. Recent work indicates that LKB1 exerts effects on metabolism, polarity and proliferation by phosphorylating and activating protein kinases belonging to the AMPK subfamily. In vivo, LKB1 forms a complex with STRAD, an inactive pseudokinase, and MO25, an armadillo repeat scaffolding-like protein. Binding of LKB1 to STRAD-MO25 activates LKB1 and re-localises it from the nucleus to the cytoplasm. To learn more about the inherent properties of the LKB1-STRAD-MO25 complex, we first investigated the activity of 34 point mutants of LKB1 found in human cancers and their ability to interact with STRAD and MO25. Interestingly, 12 of these mutants failed to interact with STRAD-MO25. Performing mutagenesis analysis, we defined two binding sites located on opposite surfaces of MO25alpha, which are required for the assembly of MO25alpha into a complex with STRADalpha and LKB1. In addition, we demonstrate that LKB1 does not require phosphorylation of its own T-loop to be activated by STRADalpha-MO25alpha, and discuss the possibility that this unusual mechanism of regulation arises from LKB1 functioning as an upstream kinase. Finally, we establish that STRADalpha, despite being catalytically inactive, is still capable of binding ATP with high affinity, but that this is not required for activation of LKB1. Taken together, our findings reinforce the functional importance of the binding of LKB1 to STRAD, and provide a greater understanding of the mechanism by which LKB1 is regulated and activated through its interaction with STRAD and MO25.
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PMID:Analysis of the LKB1-STRAD-MO25 complex. 1556 63


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