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)

The role of proteasomes in T cell activation, proliferation, and apoptosis was investigated using a proteasome-specific inhibitor lactacystin (LAC). Inhibition of the proteasome activity by LAC repressed the mitogen-induced T cell proliferation. The proteasome activity was definitively required for the T cells to progress from the G0 to S phase. It was necessary to optimize the progress from the G1/S boundary to the G2/M phase, but not for the progress from the G2/M phase to the next G1 phase. Probably as a result of a blockage of cell cycle progress, the cycling, but not the resting, T cells underwent apoptosis when treated with LAC. Mechanistically, we have found that cyclin-dependent kinase-2 (CDK2) and the cyclin E-associated kinase (largely CDK2), but not CDK4, in the G1 phase were strongly inhibited by LAC. This could be an important mechanism for the proteasome to regulate the cell cycle. The degradation of cyclin E in the late G1 and early S phases was dependent on the proteasome, although it was unlikely that this accounted for the observed inhibition of T cell proliferation. There was a reduced decay of p27Kip1 in the late G1 phase when the proteasome activity was suppressed, and this might be a contributing mechanism for the observed inhibition of CDK2 activity. Interestingly, p21Cip1 was up-regulated during the G1 phase, and the up-regulation was inhibited by LAC. Our study shows that the proteasome plays pivotal roles in regulating T cell activation and proliferation, and its effect is probably exerted through multiple mechanisms.
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PMID:Role of proteasomes in T cell activation and proliferation. 955 14

During the three last years, the so-called p16 locus on human chromosome band 9p21 has been increasingly implicated in different cancers by a variety of alterations abolishing both copies of the p16INK4a/MTS1/CDKN2 gene and the adjacent p15INK4b gene, two members of a family of specific inhibitors of the cyclin D 1-3-CDK4/6 complexes that control cell cycle progression of the G1 to S phase. While these properties are characteristic of tumor suppressor genes, abundant experimental data have clearly identified a link between the loss of function of p16INK4a and tumorigenic processes. The role of p15INK4b alterations in the onset of natural and experimental tumors is less obvious. New light may be shed on the role of the p16 locus in tumor development by the recent finding that an alternative transcript from the p16INK4a gene encodes p19ARF, a negative regulator of cell cycle progression which is unrelated to p16 and p15 and does not act by binding any CDK. Hence, this protein appears to be an element of a novel negative cell cycle control mechanism, whose impairing might be involved in tumorigenesis.
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PMID:Contribution of the dual coding capacity of the p16INK4a/MTS1/CDKN2 locus to human malignancies. 955 10

Nonsteriodal anti-inflammatory drugs (NSAIDs) are among the most commonly used medications in the United States and elsewhere, mainly for the treatment of arthritis. The NSAID sulindac causes regression and prevents the recurrence of premalignant colonic polyps in patients with familial adenomatous polyposis and inhibits colon carcinogenesis in rodents. Sulindac and sulindac sulfone, a metabolite of sulindac that lacks cyclooxygenase (cox) inhibitory activity, also inhibit mammary carcinogenesis in rats. To obtain insights into the relevance of these findings to human breast cancer, we examined the mechanism of action of sulindac and its sulfide and sulfone metabolites on the normal human mammary epithelial cell line MCF-10F and the human breast cancer cell line MCF-7. Of the three compounds, the sulfide was the most potent inhibitor of cell growth, although the sulfone and sulfoxide were also active at higher concentrations. Treatment of MCF-10F and MCF-7 cells with 100 microM sulindac sulfide resulted in accumulation of cells in the G1 phase of the cell cycle and induction of apoptosis. Apoptosis occurred within 24 h as determined by the TUNEL assay and DNA laddering was observed at 72 h. The accumulation of cells in G1 was associated with decreased levels of expression of cyclin D1 but no effect was seen on the expression of CDK4 or the immediate early response gene c-jun. Treatment with sulindac sulfide caused a striking induction of the CDK inhibitor p21WAF1 in MCF-10F cells. The MCF-7 cell line expressed a high basal level of p21WAF1 which did not change significantly after drug treatment. The pro-apoptotic gene BAX was not induced in either MCF-10F or MCF-7 cells by sulindac sulfide. Stable overexpression of cyclin D1, which frequently occurs in breast cancers, did not protect mammary epithelial cells from inhibition by the sulfide. These studies suggest that this class of compounds warrants further study with respect to breast cancer prevention and treatment.
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PMID:Effects of sulindac and its metabolites on growth and apoptosis in human mammary epithelial and breast carcinoma cell lines. 959 66

The vast majority of glioblastomas have CDKN2A, CDK4, or RB gene alterations that perturb the p16-cdk4-pRb cell cycle regulatory cascade. To explore whether immunohistochemical methods provide an alternative means of assessing this pathway, we studied 25 glioblastomas using a combination of molecular genetic and immunohistochemical assays. Homozygous deletion of the CDKN2A gene was detected in 12 of 25 (48%) cases, CDK4 amplification in 4 of 25 (16%) tumors, and loss of heterozygosity at the RB gene in 8 of 22 (36%) informative cases. Five of 25 (20%) glioblastomas had diffuse p16 immunohistochemical positivity. Significantly, all of these had either CDK4 amplification or RB LOH, suggesting that p16 immunopositivity only occurs in those tumors with alterations of another component in the pathway. Nineteen (76%) cases were uniformly immunonegative for p16, and 12 (48%) had CDKN2A homozygous deletions, but the remaining 7 cases lacked CDKN2A deletions, mutations and promoter methylation. All glioblastomas stained diffusely for cdk4, irrespective of CDK4 gene amplification status. Extensive pRb staining was present in most cases that maintained both RB alleles, and absent in most cases with RB loss, but there were notable discrepancies. Thus, p16 and pRb immunohistochemistry cannot replace molecular genetic analysis of this critical regulatory cascade; instead, the combined results hint at complex regulation of this cell cycle checkpoint. From a practical point of view, although p16 immunonegativity does not necessarily indicate CDKN2A deletion, diffuse positive p16 immunostaining strongly suggests either CDK4 amplification or RB loss and excludes CDKN2A deletion.
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PMID:Molecular genetic correlates of p16, cdk4, and pRb immunohistochemistry in glioblastomas. 960 Feb 4

Recent methodological developments allow expression measurement of many genes simultaneously, thereby revealing patterns of gene expression that can be related to phenotype. We hypothesized that through the use of such methods we could identify patterns of gene expression associated with the malignant phenotype in human bronchial epithelial cells (BEC). To test this hypothesis, a recently developed quantitative reverse transcriptase polymerase chain reaction method was used to assess simultaneously expression of 15 genes mechanistically associated with cell-cycle control (c-myc, E2F-1, p21, rb, PCNA, cyclin D2, cyclin D3, cyclin E, cdc2, CDK2, CDK4, mad, max p21, max p22, and p53) in normal cell cultures from five individuals and in nine different malignant BEC lines. Relative to the mean expression levels in cultured normal cell populations, expression of c-myc, E2F-1, PCNA, cyclin E, and CDK4 messenger RNA (mRNA) were significantly increased and expression of p21 and p53 mRNA were significantly decreased in one or two, but not all three subtypes (squamous, adenocarcinoma and small cell) of carcinoma cell lines evaluated. No single cell-cycle control gene discriminated all three subtypes from normal cell populations. In contrast, the gene expression index c-myc x E2F-1/p21 separated all carcinoma cell lines from all normal cell populations initially evaluated. This malignancy index was validated in an additional three cultured normal BEC and three carcinoma cell lines, as well as three pairs of matched primary normal bronchial epithelial and primary bronchogenic carcinoma samples, and three pairs of matched primary normal lung parenchyma and primary bronchogenic carcinoma tissue. Again, the c-myc x E2F-1/ p21 index successfully discriminated all cultured and primary normal from malignant samples and thereby had a predictive value of 1 (no false positives and no false negatives). We hypothesize that because of functional mutations in cell-cycle regulatory genes (e.g., p53 and/or rb), cells lose the ability to maintain a pattern of gene expression mechanistically associated with normal, division-limited homeostatic equilibrium. Because the c-myc x E2F-1/p21 gene expression index has high specificity for malignant tissue, it will allow confirmation that there is a significant amount of tumor tissue present in small (e.g., fine-needle) biopsy specimens prior to evaluating them for expression of other genes, such as those involved in chemoresistance or radioresistance. In addition, the goal of most gene therapy efforts is to alter levels of gene expression quantitatively. This index and others derived in a similar manner may better define potential gene therapy targets as well as response of targeted genes to therapy.
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PMID:The gene expression index c-myc x E2F-1/p21 is highly predictive of malignant phenotype in human bronchial epithelial cells. 965 Nov 76

Osteosarcomas often suffer mutations of the RB (retinoblastoma) gene, with resultant inactivation of the pRb protein. pRb is one component in a cell-cycle control pathway that includes the p16 (encoded by the CDKN2A gene) and cyclin-dependent kinase 4 (cdk4, encoded by the CDK4 gene) proteins. We therefore sought to determine whether the CDKN2A and CDK4 genes were altered in those osteosarcomas that lacked RB inactivation. Twenty-one osteosarcomas (2 low-grade and 19 high-grade) were evaluated for homozygous deletion of the CDKN2A gene, CDK4 amplification, and allelic loss of the RB gene, as well as for expression of p16 and pRb proteins. Five high-grade osteosarcomas showed loss of p16 expression; four of these had homozygous CDKN2A deletions, and the fifth had a probable deletion obscured by numerous nonneoplastic, p16-immunopositive multinucleated giant cells. Thus, p16 immunohistochemistry may provide a sensitive means for assessing CDKN2A status. Twelve tumors (including the two low-grade osteosarcomas) were immunopositive for pRb, and nine tumors were immunonegative for pRb. Of the five cases with CDKN2A/p16 alterations, none had allelic loss of the RB gene and all expressed pRb, suggesting that each of these tumors had an intact RB gene. None of the tumors showed CDK4 amplification. No alterations were detected in the two low-grade osteosarcomas. This study suggests that CDKN2A is a tumor suppressor inactivated in osteosarcomas that lack RB mutations and that the p16-pRb cell-cycle control pathway is deregulated in a large number of high-grade osteosarcomas.
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PMID:CDKN2A gene deletions and loss of p16 expression occur in osteosarcomas that lack RB alterations. 966 76

AIDS-associated Kaposi's sarcoma (KS) cell, a key element for development of KS lesions, proliferates in response to external cytokines, such as oncostatin M, the soluble IL-6R-IL-6 complex, TNF-alpha, and IL-1beta. In addition, the KS cell-produced basic fibroblast growth factor (bFGF) was reported to function as an autocrine growth factor. However, little is known of the exact roles of these external growth factors and endogenous bFGF on proliferation of KS cells, and underlying intracellular events have remained to be defined. We obtained evidence that anti-bFGF Ab abolished growth of KS cells by preventing S phase entry of the cell cycle, even in the presence of the external growth factors. Blockade of the FGF action profoundly inhibited cyclin E expression and cyclin-dependent kinase-2 (CDK2) activity, but not D-type cyclin expression and CDK4 activity. Exogenously added acidic FGF (aFGF), which generated a rapid tyrosine phosphorylation of FGFR1 and FGFR2 on KS cells, reversed the inhibitory effects of anti-bFGF Ab. Thus, FGF actions are essential for cyclin E-CDK2 activity and S phase entry. We also observed that the presence of external growth factors markedly induced cyclin E-CDK2 activity and S phase entrance, while the addition of aFGF or bFGF alone was insufficient to induce these responses. All this evidence shows that integration of the activities of external growth factors and endogenous bFGF is required for full activation of cyclin E-CDK2 activity and KS cell proliferation.
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PMID:Endogenous basic fibroblast growth factor is essential for cyclin E-CDK2 activity in multiple external cytokine-induced proliferation of AIDS-associated Kaposi's sarcoma cells: dual control of AIDS-associated Kaposi's sarcoma cell growth and cyclin E-CDK2 activity by endogenous and external signals. 971 33

Normal human diploid fibroblasts (HDF) have a finite proliferative life-span at the end of which they are arrested with a G1 phase DNA content regardless of the culture conditions. Serum stimulated senescent HDF fail to phosphorylate their retinoblastoma protein (pRb) and consequently do not express a large cohort of late G1 phase genes whose products are necessary for entry into S phase. Because pRb is believed to be phosphorylated sequentially in G1 phase by cyclin D-CDK4/6 and cyclin E-CDK2 complexes, we and others have investigated the status of these complexes in senescent HDF. There is little or no cyclin E-associated kinase activity in senescent IMR90 even though potentially active cyclin E-CDK2 complexes are present, suggesting the presence of an inhibitor. Likewise, cyclin D is complexed with its catalytic partners CDK4 and CDK6 in senescent HDF, but it is not known whether these complexes are active. p21Sdi1,Cip1,Waf1, a ubiquitous inhibitor of the activity of cyclin-CDK complexes, increases progressively throughout the life-span of HDF, but then declines again after the cells become senescent. In contrast, p16Ink4a, which binds monomeric CDK4 and CDK6 thereby preventing their binding to cyclin D, is increased dramatically at the time of senescence and remains high for at least 2 mo. Thus, it is possible that increased p21 initiates the senescent cell cycle arrest in normal cells, but p16 is important for the long-term maintenance of that arrest.
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PMID:Molecular mechanisms for the senescent cell cycle arrest. 973 51

We show here that the adenovirus E1A oncoprotein prevents growth arrest by the CDK2 inhibitor p27(Kip1) (p27) in rodent fibroblasts. However, E1A neither binds p27 nor prevents inhibition of CDK2 complexes in vivo. In contrast, the amount of free p27 available to inhibit cyclin E/CDK2 is increased in E1A-expressing cells, owing to reduced expression of cyclins D1 and D3. Moreover, E1A allows cell proliferation in the presence of supraphysiological p27 levels, while c-Myc, known to induce a cellular p27-inhibitory activity, is only effective against physiological p27 concentrations. E1A also bypasses G1 arrest by roscovitine, a chemical inhibitor of CDK2. Altogether, these findings imply that E1A can act downstream of p27 and CDK2. Retinoblastoma (pRb)-family proteins are known CDK substrates; as expected, association of E1A with these proteins (but not with p300/CBP) is required for E1A to prevent growth arrest by either p27 or the CDK4/6 inhibitor p16(INK4a). Bypassing CDK2 inhibition requires an additional function of E1A: the mutant E1A Delta26-35 does not overcome p27-induced arrest, while it binds pRb-family proteins, prevents p16-induced arrest, and alleviates pRb-mediated repression of E2F-1 transcriptional activity (although E1A Delta26-35 fails to restore expression of E2F-regulated genes in p27-arrested cells). We propose that besides the pRb family, E1A targets specific effector(s) of CDK2 in G1-S control.
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PMID:A novel function of adenovirus E1A is required to overcome growth arrest by the CDK2 inhibitor p27(Kip1). 977 42

EBV-immortalized lymphoblastoid B cell lines (LCLs) are a suitable in vitro model for the study of EBV-related lymphoproliferative disorders of immunosuppressed patients. We have previously shown that 9-cis-, 13-cis- and all-trans-retinoic acid (RA) powerfully inhibit LCL proliferation at concentrations corresponding to therapeutically achievable plasma levels (10(-6) M). Herein we show that RA-induced LCL accumulation in the G0/G1 phases correlated with the loss of the catalytic activity of all three G1-associated CDKs (CDK2, CDK4 and CDK6) and with increased levels of underphosphorylated pRb and, in some LCLs, p130. LCLs arrested in G0/G1 by RA also showed a significant decrease in the protein levels of cyclins D2, D3 and A, together with a reduction in the amount of cyclin D associated with CDK4 and CDK6, probably accounting for the inhibition of the relative kinase activity. In addition, RA-treated LCLs showed a marked up-regulation of the CDK inhibitor (CKI) p27Kip-1 at the protein but not mRNA level, which correlated with a progressive increase of p27Kip-1 in CDK2 complexes (more than 2.5-fold) and with a reduction in the active phosphorylated form of CDK2. p27Kip-1 may also contribute to the inhibition of CDK4 kinase activity, as the amount of CDK4-associated p27Kip-1 was increased by 50% after RA exposure. p27Kip-1 up-regulation stably persisted for more than one week after RA withdrawal concomitantly with the maintenance of the proliferative block. Moreover, neutralization of TGFbeta did not affect the growth inhibitory activity of RA, suggesting that LCL growth arrest induced by these retinoids is probably not mediated by a pathway directly involving TGFbeta. Overall, these results demonstrate that RA treatment of EBV-immortalized B lymphocytes is associated with multiple effects on G1 regulatory proteins, including p27Kip1 up-regulation, decreased levels of cyclins D2, D3 and A, and inhibition of CDK2, CDK4 and CDK6 activity, which ultimately result in reduced pRb phosphorylation and G0/G1 growth arrest.
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PMID:Retinoic acid-mediated growth arrest of EBV-immortalized B lymphocytes is associated with multiple changes in G1 regulatory proteins: p27Kip1 up-regulation is a relevant early event. 977 49


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