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)

Interactions between the histone deacetylase inhibitors (HDACIs) suberoylanilide hydroxamic acid (SAHA) and sodium butyrate (SB) and the heat shock protein (Hsp) 90 antagonist 17-allylamino-17-demethoxygeldanamycin (17-AAG) have been examined in human leukemia cells (U937). Coadministration of marginally toxic concentrations of 17-AAG with sublethal concentrations of SB or SAHA resulted in highly synergistic induction of mitochondrial damage (i.e., cytochrome c release), caspase-3 and -8 activation, and apoptosis. Similar interactions were noted in human promyelocytic (HL-60) and lymphoblastic (Jurkat) leukemia cells. These events were accompanied by multiple perturbations in signal transduction, cell cycle, and survival-related pathways, including early down-regulation of Raf-1, inactivation of extracellular signal-regulated kinase (ERK) 1/2 and mitogen-activated protein/ERK kinase (MEK) 1/2, diminished expression of phospho-Akt, and late activation of c-Jun-NH(2)-terminal kinase, but no changes in expression of phospho-p38 mitogen-activated protein kinase. Coadministration of 17-AAG blocked SAHA-mediated induction of the cyclin-dependent kinase inhibitor p21(CIP1) and resulted in reduced expression of p27(KIP1) and p34(cdc2). 17-AAG/SAHA-treated cells also displayed down-regulation of the antiapoptotic protein Mcl-1 and evidence of Bcl-2 cleavage. Enforced expression of doxycycline-inducible p21(CIP1) or constitutively active MEK1 significantly diminished 17-AAG/SAHA-mediated lethality, indicating that interference with ERK activation and p21(CIP1) induction play important functional roles in the lethal effects of this regimen. In contrast, enforced expression of constitutively active Akt failed to exert cytoprotective actions. Together, these findings indicate that coadministration of SAHA or SB with the Hsp90 antagonist 17-AAG in human leukemia cells leads to multiple perturbations in signaling, cell cycle, and survival pathways that culminate in mitochondrial injury and apoptosis. They also raise the possibility that combining such agents with Hsp90 antagonists may represent a novel antileukemic strategy.
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PMID:Coadministration of the heat shock protein 90 antagonist 17-allylamino- 17-demethoxygeldanamycin with suberoylanilide hydroxamic acid or sodium butyrate synergistically induces apoptosis in human leukemia cells. 1467 5

The c-myc proto-oncogene encodes a ubiquitous transcription factor involved in the control of cell growth and differentiation and implicated in inducing tumorigenesis. Understanding the function of c-Myc and its role in cancer depends upon the identification of c-Myc target genes. Heat shock protein 90 (HSP90) is involved in the folding of proteins such as signal transduction molecules (Src, Raf1, cdk4) and steroid receptors and in enhancing the activity of telomerase and nitric-oxide synthase. Here we show that c-Myc directly activates HSP90A transcription. c-Myc-mediated induction of HSP90A transcription occurs in different tissues, is independent of cell proliferation, and is mediated by a c-Myc binding site in the proximal promoter region of HSP90A gene. Overexpression of HSP90A in Rat1a cells induces transformation. Short interference RNA of HSP90A/Hsp86alpha reduces transformation activity in HeLa and RatMyc cells. These results indicate that by induction of HSP90A c-Myc may control the activity of multiple signal pathways involved in cellular transformation.
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PMID:Direct activation of HSP90A transcription by c-Myc contributes to c-Myc-induced transformation. 1472 88

The co-chaperone murine stress-inducible protein 1 (mSTI1), an Hsp70/Hsp90 organizing protein (Hop) homologue, mediates the assembly of the Hsp70/Hsp90 chaperone heterocomplex. The mSTI1 protein can be phosphorylated in vitro by cell cycle kinases proximal to a putative nuclear localization signal (NLS), which substantiated a predicted casein kinase II (CKII)-cdc2 kinase-NLS (CcN) motif at position 180-239 and suggested that mSTI1 might move between the cytoplasm and the nucleus under certain cell cycle conditions. The mechanism responsible for the cellular localization of mSTI1 was probed using NIH3T3 fibroblasts to investigate the localization of endogenous mSTI1 and enhanced green fluorescent protein (EGFP)-tagged mSTI1 mutants. Localization studies on cell lines stably expressing NLS(mSTI1)-EGFP and EGFP demonstrated that the NLS(mSTI1) was able to promote a nuclear localization of EGFP. The mSTI1 protein was exclusively cytoplasmic in most cells under normal conditions but was present in the nucleus of a subpopulation of cells and accumulated in the nucleus following inhibition of nuclear export (leptomycin B treatment). G1/S-phase arrest (using hydroxyurea) and inhibition of cdc2 kinase (using olomoucine) but not inhibition of casein kinase II (using 5,6-dichlorobenzimidazole riboside), increased the proportion of cells with endogenous mSTI1 nuclear staining. mSTI1-EGFP behaved identically to endogenous mSTI1. The functional importance of key residues was tested using modified mSTI1-EGFP proteins. Inactivation and phosphorylation mimicking of potential phosphorylation sites in mSTI1 altered the nuclear translocation. Mimicking of phosphorylation at the mSTI1 CKII phosphorylation site (S189E) promoted nuclear localization of mSTI1-EGFP. Mimicking phosphorylation at the cdc2 kinase phosphorylation site (T198E) promoted cytoplasmic localization of mSTI1-EGFP at the G1/S-phase transition,whereas removal of this site (T198A) promoted the nuclear localization of mSTI1-EGFP under the same conditions. These data provide the first evidence of nuclear import and export of a major Hsp70/Hsp90 co-chaperone and the regulation of this nuclear-cytoplasmic shuttling by cell cycle status and cell cycle kinases.
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PMID:Nuclear translocation of the Hsp70/Hsp90 organizing protein mSTI1 is regulated by cell cycle kinases. 1475 4

The effects of GA, an ansamycin antibiotic in development as a lead anticancer drug, were studied in mouse BP-A31 fibroblasts and in human cancer-derived cell lines. GA and related molecules act by inhibiting the chaperone function of the Hsp90 protein through competition for ATP binding. The antiproliferative effects of GA have been attributed to destabilization of the Raf-1 protein, one of the targets of Hsp90, and to the resulting inhibition of MAPK. Addition of GA to BP-A31 cells, synchronously progressing through the G(1) phase, inhibited Rb hyperphosphorylation and G(1)/S transition irrespective of the time of addition. The G(1) arrest was accompanied by a progressive decrease in Raf-1 content, especially of the phosphorylated form; however, GA caused only partial inhibition of MAPK phosphorylation. We show that GA triggers a rapid and marked decrease in the kinase activity of the cyclin E/cdk2 complex coupled with a decline in both total and cdk2-associated cyclin E. In transient transfection experiments, inhibition of cyclin E expression by GA was correlated with inhibition of the transcriptional activity of the cyclin E gene promoter. Inhibition of cdk4 activity by GA was observed 3 hr after addition of the drug to late G(1) cells but not after a short (1 hr) exposure, as revealed by the phosphorylation of Rb on the Ser(780) residue. In human cancer-derived cell lines expressing or not a functional Rb protein, GA blocked proliferation and inhibited the transcriptional activity of the cyclin E gene promoter. In these cell lines, the antiproliferative effect of GA was not limited to the G(1) phase, suggesting the existence of multiple cellular targets of the drug.
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PMID:Geldanamycin, an inhibitor of the chaperone activity of HSP90, induces MAPK-independent cell cycle arrest. 1499 69

The multichaperone heat shock protein (Hsp) 90 complex mediates the maturation and stability of a variety of proteins, many of which are crucial in oncogenesis, including epidermal growth factor receptor (EGF-R), Her-2, AKT, Raf, p53, and cdk4. These proteins are referred to as "clients" of Hsp90. Under unstressed conditions these proteins form complexes with Hsp90 and the cochaperones to attain their active conformations or enhance stability. Inhibition of Hsp90 function disrupts the complex and leads to degradation of client proteins in a proteasome-dependent manner. This results in simultaneous interruption of many signal transduction pathways pivotal to tumor progression and survival. Based on the unique role of the Hsp90 complex, extensive effort has been made in identifying Hsp90 inhibitors. Several compounds have been shown to inhibit Hsp90 in vitro and in vivo and the most advanced, 17-allylamino-17-demethoxygeldanamycin (AAG), is in phase I/II clinical trials. Recent findings with 17-AAG indicate that tumor cells utilize Hsp90 quite differently from normal cells, explaining the selectivity of the drug and suggesting a central role of Hsp90 in malignant progression. Thus these small molecule inhibitors have proved not only to be of great value in identifying new Hsp90 client proteins and in understanding the biology of Hsp90 but are also promising therapeutics in a variety of tumors.
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PMID:Targeting multiple signal transduction pathways through inhibition of Hsp90. 1516 26

CDK11p110 (cyclin-dependent kinase 11p110, formerly known as PITSLRE) is a member of the CDK superfamily. It associates with cyclin L and is involved in the regulation of transcription and in premRNA splicing. During staurosporine-, Fas- and tumour necrosis factor a-induced apoptosis, CDK11p110, is cleaved by caspases to generate smaller 46-50 kDa proteins containing the catalytic kinase domain. Ectopic expression of the caspase-processed form CDK11p46 induces apoptosis. The mechanisms that regulate activation and stability of CDK11 isoforms are still unclear. In the present study, we demonstrate that in human melanoma cells CDK11p110 and CDK11p46 interact with Hsp90 (heat-shock protein 90) and its co-chaperone cdc37. Furthermore, we show that the treatment of cells with the Hsp90-specific inhibitor geldanamycin leads to ubiquitination and enhanced degradation of both CDK11p110 and CDK11p46 through a proteasome-dependent pathway. We also determined that geldanamycin-triggered degradation of CDK11p46 slows down the progression of apoptosis. These results indicate that Hsp90 and cdc37 stabilize CDK11 kinase, and suggest that this stabilization is crucial for its pro-apoptotic function.
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PMID:Regulation of stability of cyclin-dependent kinase CDK11p110 and a caspase-processed form, CDK11p46, by Hsp90. 1534 6

The heat shock protein Hsp90 is a potential target for drug discovery of novel anticancer agents. By affecting this protein, several cell signaling pathways may be simultaneously modulated. The geldanamycin analog 17AAG has been shown to inhibit Hsp90 and associated proteins. Its clinical use, however, is hampered by poor solubility and thus, difficulties in formulation. Therefore, a water-soluble derivative was desirable and 17-dimethylaminoethylamino-17-demethoxy-geldanamycin (17DMAG) is such a derivative. Studies were carried out in order to evaluate the activity and molecular mechanism(s) of 17DMAG in comparison with those of 17-allylamino-demethoxygeldanamycin (17AAG). 17DMAG was found to be more potent than 17AAG in a panel of 64 different patient-derived tumor explants studied in vitro in the clonogenic assay. The tumor types that responded best included mammary cancers (six of eight), head and neck cancers (two of two), sarcomas (four of four), pancreas carcinoma (two of three), colon tumors (four of eight for 17AAG and six of eight for 17DMAG), and melanoma (two of seven). Bioinformatic comparisons suggested that, while 17AAG and 17DMAG are likely to share the same mode(s) of action, there was very little similarity with standard anticancer agents. Using three permanent human melanoma cell lines with differing sensitivities to 17AAG and 17DMAG (MEXF 276L, MEXF 462NL and MEXF 514L), we found that Hsp90 protein was reduced following treatment at a concentration associated with total growth inhibition. The latter occurred in MEXF 276L cells only, which are most sensitive to both compounds. The depletion of Hsp90 was more pronounced in cells exposed to 17DMAG than in those treated with 17AAG. The reduction in Hsp90 was associated with the expression of erbB2 and erbB3 in MEXF 276L, while erbB2 and erbB3 were absent in the more resistant MEXF 462NL and MEXF 514L cells. Levels of known Hsp90 client proteins such as phosphorylated AKT followed by AKT, cyclin D1 preceding cdk4, and craf-1 declined as a result of drug treatment in all three melanoma cell lines. However, the duration of drug exposure needed to achieve these effects was variable. All cell lines showed increased expression of Hsp70 and activated cleavage of PARP. No change in PI3K expression was observed and all melanoma cells were found to harbor the activating V599E BRAF kinase mutation. The results of our in vitro studies are consistent with both 17AAG and 17DMAG acting via the same molecular mechanism, i.e. by modulating Hsp90 function. Since 17DMAG can be formulated in physiological aqueous solutions, the data reported here strongly support the development of 17DMAG as a more pharmaceutically practicable congener of 17AAG.
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PMID:Comparison of 17-dimethylaminoethylamino-17-demethoxy-geldanamycin (17DMAG) and 17-allylamino-17-demethoxygeldanamycin (17AAG) in vitro: effects on Hsp90 and client proteins in melanoma models. 1584 78

The molecular chaperone Hsp90 is distinct from Hsp70 and chaperonin in that client proteins are apparently restricted to a subset of proteins categorized as cellular signaling molecules. Among these, many specific protein kinases require the assistance of Hsp90 and its co-chaperone Cdc37/p50 for their biogenesis. A series of Cdc37 deletion mutants revealed that all mutants capable of binding Raf-1 possess amino acid residues between 181 and 200. The 20-residue region is sufficient and, in particular, a five-residue segment (residue 191-195) is essential for binding to Raf-1. These five residues are present in one alpha helix (residues 184-199) in the middle of Cdc37, which is unexpectedly nested within the Hsp90-interacting domain of Cdc37, which was recently determined by crystallography, but does not seem to contribute to direct contact with Hsp90. Furthermore, an N-terminally truncated mutant of Cdc37 composed of residues 181-378 was shown to bind the N-terminal portion of Raf-1 (subdomains I-IV). This mutant can bind not only other Hsp90 client protein kinases, Akt1, Aurora B and Cdk4, but also Cdc2 and Cdk2, which to date have not been shown to physically interact with Cdc37. These results suggest that a region of Cdc37 other than the client-binding site may be responsible for discriminating client protein kinases from others.
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PMID:A client-binding site of Cdc37. 1615 89

Hsp90 and its cochaperone Cdc37 cooperate to provide requisite support to numerous protein kinases involved in cellular signal transduction. In this report, we studied the interactions of Hsp90 and Cdc37 with the cyclin-dependent kinase, Cdk2. Treatment of K562 cells with the Hsp90 inhibitor, geldanamycin, caused a 75% reduction in Cdk2 levels and reduced the levels of its activating kinase, Cdk7, by more than 60%, suggesting that both of these kinases may be Hsp90 clients. Using classical pull-down assays and the Hsp90 inhibitory agents geldanamycin and molybdate, Cdk2 is shown to be a genuine client of the Hsp90 chaperone complex. Subsequently, pull-down assays directed at helix alphaC of Cdk2 are shown to disrupt Hsp90 and Cdc37 binding and explain the initial difficulties in demonstrating these interactions. Mutant constructs containing deletions of secondary structural elements from the N- and C-termini of Cdk2 were prepared and assayed for their ability to coadsorb Hsp90 and Cdc37 in a salt-stable high-affinity manner with and without the addition of molybdate. Consistent with similar work done with the cyclin-dependent kinase relative Cdk4, the presence of the G-box motif of Cdk2 was shown to be critical for Cdc37 binding, whereas consistent with work done with the Src-family tyrosine kinase Lck, the presence of helix alphaC and the stabilization of helix alphaE were shown to be needed for Hsp90 binding.
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PMID:Cdk2: a genuine protein kinase client of Hsp90 and Cdc37. 1628 32

DAF-21, a Caenorhabditis elegans homologue of Hsp90, is expressed primarily in germline cells. Although mutations in the daf-21 gene affect animal fertility, its cellular roles have remained elusive. To phenocopy daf-21 mutations, we impaired the daf-21 function by RNA interference (RNAi), and found that oocytes skipped the diakinesis arrest and displayed a defective diakinesis arrest, which led to the production of endomitotic oocytes with polyploid chromosomes (Emo phenotype). The same Emo phenotype was also observed with RNAi against wee-1.3. To identify a cause for Emo, we examined the CDK-1 (Cdc2) phosphorylation status in Emo animals, since CDK-1 is a key regulator of the prophase/metaphase transition and is kept inactivated by WEE-1.3 kinase during prophase. We immunostained both daf-21(RNAi) and wee-1.3(RNAi) animals with anti-phosphorylated-CDK-1 antibody and observed no detectable phosphates on CDK-1 in either of the animals. We also examined WEE-1.3 expression in daf-21(RNAi) and found a significant reduction of WEE-1.3. These results indicate that CDK-1 was not phosphorylated in either daf-21(RNAi) or wee-1.3(RNAi) animals, and suggest that daf-21 was necessary for producing functional WEE-1.3. Thus, all together, we propose that DAF-21 indirectly regulates the meiotic prophase/metaphase transition during oocyte development by ensuring the normal function of WEE-1.3.
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PMID:Cell cycle control by daf-21/Hsp90 at the first meiotic prophase/metaphase boundary during oogenesis in Caenorhabditis elegans. 1646 90


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