Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.27 (AMPK)
 6,299 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

StubSNF1 is a potato cDNA that encodes a protein kinase similar to the yeast SNF1 gene involved in transcriptional regulation of glucose-repressible genes. The yeast SNF1 functions in a complex with GAL83/SIP1/SIP2 and SNF4 proteins. We have used StubSNF1 as bait in a yeast two-hybrid system to screen for potato cDNAs encoding proteins that bind to StubSNF1. Three overlapping cDNAs, two different in size, were isolated. DNA sequence analysis revealed that they were orthologues of the yeast GAL83/SIP1/SIP2 genes and their mammalian counterparts, AMPK beta-subunits. The direct interaction between the potato proteins StubGAL83 and StubSNF1 was shown by an in vitro binding assay. Southern and Northern hybridisations revealed that StubGAL83 exists in a low copy number in the potato genome and is highly (but organ-specifically) expressed in potato. In contrast, StubSNF1 possesses low transcript levels in each organ, except in flowers where high amounts of StubSNF1 mRNA could be detected. We demonstrate here that StubGAL83 can also interact with yeast SNF4 in a yeast two-hybrid system suggesting that plant SNF1 kinases may function in complexes similar to those detected in yeast and mammals.
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PMID:Potato StubSNF1 interacts with StubGAL83: a plant protein kinase complex with yeast and mammalian counterparts. 1020 10

Previous findings indicate that the protein c-KIT and its ligand, stem cell factor (SCF) play a crucial role in the development of melanocytes from their precursors in the embryonic neural crest cells. Using a monoclonal anti-c-KIT antibody, ACK2, which is an antagonistic blocker of c-KIT function, we and colleagues demonstrated that mouse melanocytes disappeared with the injection of ACK2 during certain periods of embryonic and postnatal life. The precise mechanisms of this disappearance, however, remain unclear. Because melanocytes disappeared without any inflammation in these in vivo studies, we suspect that apoptosis was a main cause of their disappearance. In this study, to clarify the underlying mechanism, we studied whether ACK2 induces apoptosis in c-KIT-positive melanoblasts, which appear in mouse neural crest cells cultured with SCF from 9.5 d old mouse embryos. With an in situ apoptosis detection kit, a significant increase in apoptosis was detected after the removal of SCF, which further increased with the addition of ACK2 during SCF-dependent periods. The occurrence of apoptosis in the cultured cells was also demonstrated by a DNA analysis and electron microscopy. Immunohistochemical double staining confirmed that the apoptotic cells were c-KIT positive, and the electron microscopy showed that these apoptotic cells were melanocyte precursors. It was therefore demonstrated that apoptosis was induced in the SCF-dependent c-KIT-positive melanocytes in vitro when the SCF/c-KIT interaction was obstructed. These findings elucidate the mechanism of the regulation of melanocyte development, and the survival and proliferation of these precursor cells, by SCF/c-KIT interaction.
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PMID:Removal of stem cell factor or addition of monoclonal anti-c-KIT antibody induces apoptosis in murine melanocyte precursors. 1023 74

The myocyte enhancer factor (MEF)2 transcription factor is important for development of differentiated skeletal muscle. We investigated the regulation of MEF2 DNA binding in differentiated primary human skeletal muscle cells and isolated rat skeletal muscle after exposure to various stimuli. MEF2 DNA binding activity in nonstimulated (basal) muscle cultures was almost undetectable. Exposure of cells for 20 min to 120 nM insulin, 0.1 and 1.0 mM hydrogen peroxide, osmotic stress (400 mM mannitol), or 1.0 mM 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR) led to a profound increase in MEF2 DNA binding. To study signaling pathways mediating MEF2 activity, we preincubated human skeletal muscle cell cultures or isolated rat epitrochlearis muscles with inhibitors of p38 mitogen-activated protein kinase (MAPK) (10 microM SB-203580), MEK1 (50 microM PD-98059), PKC (1 and 10 microM GF109203X), phosphatidylinositol (PI) 3-kinase (10 microM LY-294002), or AMP-activated protein kinase (AMPK; 20 microM compound C). All stimuli resulted primarily in activation of MEF2D DNA binding. Exposure of cells to osmotic or oxidative stress increased MEF2 DNA binding via pathways that were completely blocked by MAPK inhibitors and partially blocked by inhibitors of PKC, PI 3-kinase, and AMPK. In epitrochlearis muscle, MAPK inhibitors blocked contraction but not AICAR-mediated MEF2 DNA binding. Thus activation of MEF2 in skeletal muscle is regulated via parallel intracellular signaling pathways in response to insulin, cellular stress, or activation of AMPK.
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PMID:MEF2 activation in differentiated primary human skeletal muscle cultures requires coordinated involvement of parallel pathways. 1496 Apr 15

A novel member of the human AMPK family, ARK5, was recently discovered to be a key molecule in mediating cancer cell migration activity in human pancreas cancer cell line PANC-1, and its activation was found to be induced by Akt-dependent phosphorylation at Ser 600. DNA array analysis with 241 paired cDNAs from 13 different types of tumors and corresponding normal tissues derived from cancer patients revealed ARK5 overexpression in the samples of colorectal cancer. ARK5 expression was measured and an in vitro invasion assay was performed in six human colorectal cancer cell lines, WiDr, HCT-15, DLD-1, SW620, LoVo, and SW480, and since high invasion activity was concordant with higher ARK5 expression, ARK5 expression was examined in relation to tumor progression and metastatic activity in clinical samples. In 56 clinical samples of primary colorectal cancers and their liver metastases, higher ARK5 expression was observed in the samples from more advanced cases, and much higher expression was observed in the liver metastases. In situ hybridization analysis showed ARK5 overexpression in tumor cells. Based on these findings, we propose that ARK5 overexpression is involved in tumor progression of colon cancer clinically.
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PMID:ARK5 expression in colorectal cancer and its implications for tumor progression. 1498 52

Glucose transport into muscle is important for the maintenance of normoglycemia. Thus, understanding mechanisms that regulate expression of GLUT4, the main glucose transporter in skeletal muscle, is important to identify targets for the treatment of diabetes. Exercise increases the expression of GLUT4 mRNA and protein, and we have been investigating the mechanisms involved. Transcription of the GLUT4 gene is transiently activated after an acute bout of exercise and GLUT4 protein can be increased as much as two- to threefold after a few days of repeated exercise bouts. Studies of the GLUT4 promoter have identified two sets of DNA sequences that are important for metabolic regulation and also for increased transcription of the gene in response to exercise. These DNA elements have been shown to bind the transcription factors myocyte enhancer factor 2 (MEF2) and GLUT4 enhancer factor (GEF). The mechanisms that activate these proteins remain one of the important areas of research in this field. Signals that link muscle contraction to the activation of transcription factors (MEF2, GEF) involved in increased expression of GLUT4 during exercise is another area needing further research. Two signals that show promise are changes in the energy charge (acting through AMP activated kinase [AMPK]) and changes in intracellular calcium (acting through calcineurin [a calcium-calmodulin activated phosphatase] and calcium-calmodulin activated kinase [CAMK]). There is good evidence that both increased AMPK activity and increased CAMK activity cause increased transcription of the GLUT4 gene. It remains to be demonstrated that exercise is acting through one or both of these mechanisms.
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PMID:Regulation of GLUT4 gene expression during exercise. 1523 26

Stem cell factor (SCF) and its receptor, KIT, are essential to the migration and differentiation of melanocytes during embryogenesis. We previously demonstrated that apoptosis is induced by blocking survival function of the SCF/KIT interaction in a mouse neural crest cell (NCC) primary culture. Using the NCCmelb4 cell line, we investigated the occurrence of apoptosis in the cultured cells when KIT receptors were blocked by the monoclonal anti-KIT antibody (ACK2). Apoptosis following treatment with ACK2 was detected by DNA fragmentation assay, in situ apoptosis detection, and electron microscopy. We noted a decrease in extracellular signal-related kinase (ERK) and ribosomal S6 kinase (RSK) protein expression following ACK2 incubation. Western blot analysis and real-time quantitative RT-PCR revealed an apparent time-dependent reduction in Bcl-2 protein levels with respect to ACK2 within the NCCmelb4 cells. In terms of Bax expression, a difference was not found. Fas and caspase8 proteins increased time-dependently in proportion to ACK2 incubation. We noted apoptotic cell death upon addition of ACK2, with evidence of possible involvement of Bcl-2 and Fas in the induction of apoptosis. In contrast, no significant correlation between Fas ligand (Fas-L) expression and ACK2 was found. Fas activation appears to occur independent of Fas-L during ACK2-induced cell death. Therefore, we propose that Fas-L expression in NCCmelb4 cells does not play a major role in facilitating apoptosis. Furthermore, we hypothesize that these molecules combined with SCF/KIT play an important role in regulating the induction of vertebrate NCC apoptosis during embryogenesis.
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PMID:Bcl-2 reduced and fas activated by the inhibition of stem cell factor/KIT signaling in murine melanocyte precursors. 1565 78

In addition to their ligand-mediated activation, nuclear receptor activity is finely tuned by their phosphorylation status. PPARs are phosphorylated by several kinases (PKA, PKC, MAPKs, and AMPK), which affect their activity in a ligand-dependent or -independent manner according to the isoform and cellular context. Molecular consequences are multiple, including changes in ligand affinity, DNA binding, recruitment of transcriptional cofactors, proteasome degradation... Finally, the physiological relevance of PPAR phosphorylation is discussed.
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PMID:Phosphorylation of PPARs: from molecular characterization to physiological relevance. 1573 34

AMPK acts as a cellular fuel gauge and responds to decreased cellular energy status by inhibiting ATP-consuming pathways and increasing ATP-synthesis. The aim of this study was to examine the role of AMPK in modulating poly(ADP-ribose) polymerase (PARP), a nuclear enzyme involved in maintaining chromatin structure and DNA repair. HT-29 cells infected with constitutively active AMPK demonstrated increased PARP automodification and an increase in bioNAD incorporation. AMPK and PARP co-immunoprecipitated under basal conditions and in response to H(2)O(2), suggesting a physical interaction under both resting and stress-induced conditions. Incubation of PARP with purified AMPK resulted in the phosphorylation of PARP; and the inclusion of AMP as an AMPK activator potentiated PARP phosphorylation. Using immobilized PARP, the incorporation of bioNAD by PARP was dramatically increased following the addition of AMPK. These data suggest a novel role for AMPK in regulating PARP activity through a direct interaction involving phosphorylation.
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PMID:AMP-activated protein kinase is a positive regulator of poly(ADP-ribose) polymerase. 1648 Sep 59

Genetic and biochemical studies have shown that Ser(20) phosphorylation in the transactivation domain of p53 mediates p300-catalyzed DNA-dependent p53 acetylation and B-cell tumor suppression. However, the protein kinases that mediate this modification are not well defined. A cell-free Ser(20) phosphorylation site assay was used to identify a broad range of calcium calmodulin kinase superfamily members, including CHK2, CHK1, DAPK-1, DAPK-3, DRAK-1, and AMPK, as Ser(20) kinases. Phosphorylation of a p53 transactivation domain fragment at Ser(20) by these enzymes in vitro can be mediated in trans by a docking site peptide derived from the BOX-V domain of p53, which also harbors the ubiquitin signal for MDM2. Evaluation of these calcium calmodulin kinase superfamily members as candidate Ser(20) kinases in vivo has shown that only CHK1 or DAPK-1 can stimulate p53 transactivation and induce Ser(20) phosphorylation of p53. Using CHK1 as a prototypical in vivo Ser(20) kinase, we demonstrate that (i) CHK1 protein depletion using small interfering RNA can attenuate p53 phosphorylation at Ser(20), (ii) an enhanced green fluorescent protein (EGFP)-BOX-V fusion peptide can attenuate Ser(20) phosphorylation of p53 in vivo, (iii) the EGFP-BOX-V fusion peptide can selectively bind to CHK1 in vivo, and (iv) the Deltap53 spliced variant lacking the BOX-V motif is refractory to Ser(20) phosphorylation by CHK1. These data indicate that the BOX-V motif of p53 has evolved the capacity to bind to enzymes that mediate either p53 phosphorylation or ubiquitination, thus controlling the specific activity of p53 as a transcription factor.
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PMID:The MDM2 ubiquitination signal in the DNA-binding domain of p53 forms a docking site for calcium calmodulin kinase superfamily members. 1733 37

Deletion mutations of mitochondrial DNA (mtDNA) accumulate somatically on a cell-by-cell basis with age, resulting in decreased cell function in muscle and substantia nigra. In osteosarcoma cells deletions incapacitate mitochondria and induce the autophagic transcript ATG12, which is involved in an early step of the mammalian autophagy pathway. We discuss here which consequences of mtDNA deletions could induce ATG12, and provide two new pieces of data. Our previous studies demonstrated that mtDNA deletions decreased mitochondrial ATP production and proteasomal function, induced the AMPK transcript (likely as a consequence of bioenergetic depletion), and decreased the intracellular concentration of 20 amino acids (possibly as a consequence of decreased proteasomal activity). Deletions eliminate essential tRNAs for mitochondrial protein synthesis, as well as essential components of mitochondrial multisubunit enzymes; therefore, the increased level of ATG12 could result from decreased bioenergetic function, increased oxidative damage, or decreased mitochondrial protein synthesis. However, the bioenergetic inhibitor rotenone does not induce ATG12. We show here that chloramphenicol, which inhibits mitochondrial protein synthesis, induces ATG12, and that mtDNA deletions result in an increased burden of oxidatively damaged protein. Thus, mtDNA deletions could induce ATG12 through a mechanism such as the following: deletions > mitochondrial protein synthesis inhibition or ROS > proteasome inhibition > amino acid depletion > ATG12.
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PMID:Mitochondrial DNA deletions and chloramphenicol treatment stimulate the autophagic transcript ATG12. 1715 91


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