Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.4.2.30 (PARP)
 13,611 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Nanomolar concentrations of Taxol, and other antimitotic agents that interact with microtubules, mediate serine phosphorylation of the 66-kDa Shc isoform (p66shc) in A549 human lung carcinoma cells, 9-18 h after drug treatment. This event coincides with the release of PARP cleavage fragments that are early indicators of apoptosis. Taxol-induced serine phosphorylation of p66shc results from a MEK-independent signaling pathway that is activated in A549 cells that have a prolonged or abnormal mitotic phase of the cell cycle [Cancer Res. 60 (2000) 5171]. In contrast, in murine macrophage RAW 264.7 cells, micromolar concentrations of Taxol but not other microtubule-interacting agents induced serine phosphorylation of p66shc that correlated with the phosphorylation of Raf-1 and extracellular signal-regulated kinase (ERK1/2), within 15-30 min after Taxol treatment. This event also was induced by lipopolysaccharide (LPS). The MEK-inhibitor, U0126, that specifically inhibits the activation of ERK also blocked the phosphorylation of p66shc and Raf-1, suggesting that these processes were MEK-dependent, quite different from that which was observed in A549 cells. Taxol also induced phosphorylation of p38 and JNK MAP kinases within 8-15 min after drug treatment. It is known that Taxol, but not other microtubule-interacting agents, induces the production of cytokines, such as tumor necrosis factor alpha (TNF-alpha) in mouse macrophages. The time course of Taxol-induced TNF-alpha expression coincides with that of Taxol-induced p66shc phosphorylation, and U0126 inhibits significantly Taxol-induced TNF-alpha expression in RAW 264.7 cells. Our data indicate that the Taxol-induced serine phosphorylation of p66shc in RAW 264.7 cells is microtubule-independent and may be related to increased TNF-alpha expression after Taxol and LPS treatment. It is concluded that the mechanisms involved in Taxol-induced p66shc phosphorylation are distinct in A549 and RAW 264.7 cells.
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PMID:Distinct mechanisms of taxol-induced serine phosphorylation of the 66-kDa Shc isoform in A549 and RAW 264.7 cells. 1206 70

Despite the contrasts in chemistry and toxicity, for blister and nerve chemical warfare agents there may be some analogous proteolytic and inflammatory mediators and pathological pathways that can be pharmacological targets for a single-drug multi-threat medical countermeasure. The dermal-epidermal separation caused by proteases and bullous diseases compared with that observed following exposure to the blister agent sulfur mustard (2,2'-dichlorodiethyl sulfide) has fostered the hypothesis that sulfur mustard vesication involves proteolysis and inflammation. In conjunction with the paramount toxicological event of cholinergic crisis that causes acute toxicity and precipitates neuronal degeneration, both anaphylactoid reactions and pathological proteolytic activity have been reported in nerve-agent-intoxicated animals. Two classes of drugs already have demonstrated multi-threat activity for both nerve and blister agents. Serine protease inhibitors can prolong the survival of animals intoxicated with the nerve agent soman and can also protect against vesication caused by the blister agent sulfur mustard. Poly (ADP-ribose) polymerase (PARP) inhibitors can reduce both soman-induced neuronal degeneration and sulfur-mustard-induced epidermal necrosis. Protease and PARP inhibitors, like many of the other countermeasures for blister and nerve agents, have potent primary or secondary anti-inflammatory pharmacology. Accordingly, we hypothesize that drugs with anti-inflammatory actions against either nerve or blister agent might also display multi-threat efficacy for the inflammatory pathogenesis of both classes of chemical warfare agent.
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PMID:Putative role of proteolysis and inflammatory response in the toxicity of nerve and blister chemical warfare agents: implications for multi-threat medical countermeasures. 1279 39

We recently characterized the interaction between poly(ADP-ribose) polymerase-1 (PARP-1) and the product of the tumor suppressor gene p53. We investigated which domains of human PARP-1 and of human wild-type (wt) p53 were involved in this protein-protein interaction. We generated baculoviral constructs encoding full length or distinct functional domains of both proteins. Full length PARP-1 was simultaneously coexpressed in insect cells with full length wt p53 protein or its distinct truncated fragments and vice versa. Reciprocal immunoprecipitation of Sf9 cell lysates revealed that the central and carboxy-terminal fragments of p53 were sufficient to confer binding to PARP-1, whereas the amino-terminal part harboring the transactivation functional domain was dispensable. On the other hand, the amino-terminal and central fragments of PARP-1 were necessary for complex formation with p53 protein. As the most important features of p53 protein are regulated by phosphorylation, we addressed the question of whether its phosphorylation is essential for binding between the two proteins. Baculovirally expressed wt p53 was post-translationally modified. At least six distinct p53 isomeres were resolved by immunoblotting following two-dimensional separation of baculovirally expressed wt p53 protein. Using specific phospho-serine antibodies, we identified phosphorylation of baculovirally expressed p53 protein at five distinct sites. To define the role of p53 phosphorylation, pull-down assays using untreated and dephosphorylated p53 protein were performed. Dephosphorylated p53 failed to bind PARP-1 indicating that complex formation between both proteins is regulated by phosphorylation of p53. The marked phosphorylation of p53 at Ser392 observed in unstressed cells suggests that the phosphorylated carboxy-terminal part of p53 undergoes complex formation with PARP-1 resulting in masking of the NES and thereby preventing its export. The functional significance of the interaction between both proteins was investigated at two different conditions: inactivation of PARP-1 and overexpression of PARP-1. Our results unequivocally show that the presence of PARP-1 regulates the basal expression of wt p53 in unstressed cells.
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PMID:Phosphorylation regulates the interaction and complex formation between wt p53 protein and PARP-1. 1289 23

The replication and transcription activator (RTA) of gamma-2 herpesvirus is sufficient to drive the entire virus lytic cycle. Hence, the control of RTA activity should play an important role in the maintenance of viral latency. Here, we demonstrate that cellular poly(ADP-ribose) polymerase 1 (PARP-1) and Ste20-like kinase hKFC interact with the serine/threonine-rich region of gamma-2 herpesvirus RTA and that these interactions efficiently transfer poly(ADP-ribose) and phosphate units to RTA. Consequently, these modifications strongly repressed RTA-mediated transcriptional activation by inhibiting its recruitment onto the promoters of virus lytic genes. Conversely, the genetic ablation of PARP-1 and hKFC interaction or the knockout of the PARP-1 gene and activity considerably enhanced gamma-2 herpesvirus lytic replication. Thus, this is the first demonstration that cellular PARP-1 and hKFC act as molecular sensors to regulate RTA activity and thereby, herpesvirus latency.
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PMID:Poly(ADP-ribose) polymerase 1 and Ste20-like kinase hKFC act as transcriptional repressors for gamma-2 herpesvirus lytic replication. 1458 85

Latent membrane protein 2A (LMP2A) blocks B-cell receptor signal transduction in vitro by binding the Syk and Lyn protein tyrosine kinases. As well as blocking B-cell signal transduction, LMP2A has been shown to activate the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway, which acts as a survival signal in both B cells and epithelial cells. Transforming growth factor beta 1 (TGF-beta 1) is a multifunctional cytokine that plays important roles in regulating cell growth and differentiation in many biological systems. The loss of the growth-inhibitory response to the TGF-beta 1 signal is found in many cancers and is widely thought to promote tumor development. In this study, we found that LMP2A induced the phosphorylation of Akt (serine 473) in Burkitt's lymphoma cell line Ramos and in gastric carcinoma cell line HSC-39 and partially enhanced cell viability following TGF-beta 1 treatment. In addition, LMP2A partially inhibited TGF-beta 1-induced DNA fragmentation and cleavage of poly(ADP-ribose) polymerase (PARP). In the presence of LY294002, an inhibitor of PI3-K, the LMP2A-mediated inhibitory effects on TGF-beta 1-induced DNA fragmentation and cleavage of PARP were alleviated. Furthermore, LMP2A did not alter the levels of expression of type I and type II TGF-beta 1 receptors. Taken together, these results suggest that LMP2A may inhibit TGF-beta 1-mediated apoptosis through activation of the PI3-K/Akt pathway.
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PMID:Latent membrane protein 2A inhibits transforming growth factor-beta 1-induced apoptosis through the phosphatidylinositol 3-kinase/Akt pathway. 1474 35

Overexpression of the anti-apoptotic protein Bcl-2 has been associated with several malignancies, including small cell lung cancer (SCLC). In the present study, we have investigated if Bcl-2 contributes to the emergence of cisplatin resistance in SCLC H69 cells. The ability of cisplatin to induce apoptosis was decreased in H69 cells that acquired resistance to cisplatin (H69/CP). The level of Bcl-2 was, however, substantially reduced in H69/CP cells compared to parental H69 cells. There was little change in Bcl-2 content in H69 cells that were resistant to etoposide (VP-16) or Taxol. Bcl-2 was constitutively phosphorylated at serine 70 in H69 cells but not in H69/CP cells and cisplatin had little effect on Bcl-2 phosphorylation. The level of procaspase-3 was elevated in H69/CP cells but the ability of cisplatin to induce mitochondrial depolarization, caspase-9 activation, and poly(ADP-ribose) polymerase (PARP) cleavage was compromised in H69/CP cells. The level of the anti-apoptotic protein Bcl-x(L) and the pro-apoptotic protein Bax was slightly reduced in H69/CP cells but the ratio of pro-apoptotic and anti-apoptotic Bcl-2 family proteins was not sufficient to explain cellular resistance to cisplatin. These results suggest that the acquisition of cisplatin resistance by H69 cells was not due to an increase in the level/phosphorylation status of the anti-apoptotic protein Bcl-2.
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PMID:Down-regulation of Bcl-2 is associated with cisplatin resistance in human small cell lung cancer H69 cells. 1502 53

6-O-Palmitoyl ascorbic acid (PAA) has recently been used as a substitute for ascorbic acid because of its greater potency as an antioxidant. In detailed concentration response studies distinct cytotoxic effects of PAA at concentrations exceeding 100 microM were reported. Here we examined and further characterized this cytotoxicity. While ascorbic acid was tolerated well up to millimolar concentrations, PAA revealed an LC50 between 125 and 150 microM in rat GH3 tumor cells. Morphological and biochemical observations suggested the induction of apoptosis at concentrations exceeding 125 microM with a prominent activation of caspase 3 at 250 microM after 4 hr. A subsequent pronounced fragmentation of DNA (DNA-ladder) was detected after 6 hr and was further enhanced after 12 hr. The activation of caspases and the cleavage of its substrate PARP was preceded by a distinct increase in the phosphorylation of stress activated JNK-kinases. This observation suggested that the agent affected signal transduction mechanisms regulating protein phosphorylation at serine/threonine residues in the cell. No effect of PAA on protein phosphatase 2A (PP2A)-like activity was observed while magnesium-dependent protein phosphatase activity, presumably PP2C, was inhibited concentration-dependently up to 75% at the respective concentrations. Thus, the cytotoxic, pro-apoptotic effect of PAA might be related to the inhibition of PP2C and the activation of JNK.
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PMID:Apoptosis by 6-O-palmitoyl-L-ascorbic acid coincides with JNK-phosphorylation and inhibition of Mg2+-dependent phosphatase activity. 1510 45

Benzyl isothiocyanate (BITC), a cruciferous vegetable-derived compound, has been shown to inhibit chemically induced cancer in animal models. Moreover, epidemiological studies have provided compelling evidence to suggest that cruciferous vegetables may be protective against cancer risk. Here, we report that BITC significantly inhibits growth of human pancreatic cancer BxPC-3 cells in a concentration-dependent manner with an IC(50) of approximately 8 micro M, a concentration that can be generated through dietary intake of cruciferous vegetables. Treatment of BxPC-3 cells with growth suppressive concentrations of BITC resulted in G(2)/M phase cell cycle arrest that was associated with a marked decline in protein levels of G(2)/M regulatory proteins including cyclin-dependent kinase 1 (Cdk1), cyclin B1 and cell division cycle 25B (Cdc25B). Further, BITC-mediated growth inhibition of BxPC-3 cells correlated with apoptosis induction that was characterized by an increase in Bax/Bcl-2 ratio, cleavage of procaspase-3 and poly(ADP-ribose)polymerase (PARP), and an increase in cytoplasmic histone-associated DNA fragmentation. Interestingly, BITC treatment caused inhibition of nuclear factor kappaB (NF-kappaB) activation, which is constitutively activated in human pancreatic cancer. Western blotting revealed concentration-dependent decrease in NF-kappaB/Rel-p65 protein level in BxPC-3 cells upon exposure to BITC. An increase in protein level of inhibitory subunit kappaB (IkappaBa) in association with reduced serine-32 phosphorylation was also observed in BITC-treated BxPC-3 cells. Consistent with these findings, BITC treatment caused a decrease in nuclear translocation of NF-kappaB as reflected by reduced DNA-binding capacity of NF-kappaB. Furthermore, the protein level of cyclin D1, a transcriptional target of NF-kappaB, was reduced significantly in BITC-treated BxPC-3 cells. To the best of our knowledge, this study is the first published report to implicate suppression of NF-kappaB activation as a potential mechanism for anti-proliferative activity of BITC against human pancreatic cancer cells.
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PMID:Cell cycle arrest, apoptosis induction and inhibition of nuclear factor kappa B activation in anti-proliferative activity of benzyl isothiocyanate against human pancreatic cancer cells. 1511 14

DNA double-strand breaks (DSB) mobilize DNA-repair machinery and cell cycle checkpoint by activating the ataxia-telangiectasia (A-T) mutated (ATM). Here we show that ATM kinase activity is inhibited by poly(ADP-ribose) polymerase-1 (PARP-1) in vitro. It was shown by biochemical fractionation procedure that PARP-1 as well as ATM increases at chromatin level after induction of DSB with neocarzinostatin (NCS). Phosphorylation of histone H2AX on serine 139 and p53 on serine 15 in Parp-1 knockout (Parp-1(-/-)) mouse embryonic fibroblasts (MEF) was significantly induced by NCS treatment compared with MEF derived from wild-type (Parp-1(+/+)) mouse. NCS-induced phosphorylation of histone H2AX on serine 139 in Parp-1(-/-) embryonic stem cell (ES) clones was also higher than that in Parp-1(+/+) ES clone. Furthermore, in vitro, PARP-1 inhibited phosphorylation of p53 on serine 15 and (32)P-incorporation into p53 by ATM in a DNA-dependent manner. These results suggest that PARP-1 negatively regulates ATM kinase activity in response to DSB.
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PMID:Poly(ADP-ribose) polymerase-1 inhibits ATM kinase activity in DNA damage response. 1517 48

Although several lines of evidence support a role for serine proteases in apoptosis, little is known about the mechanisms involved. In the present study, we have examined the apoptosis-inducing potential and dissected the death-signalling pathways of N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) and N-tosyl-L-lysine chloromethyl ketone (TLCK), inhibitors of chymotrypsin- and trypsin-like proteases, respectively. Our results designate two distinct roles for serine proteases. Firstly, we show that both inhibitors induce biochemical and morphological characteristics of apoptosis, including proteolysis of poly(ADP-ribose) polymerase 1 (PARP-1) and inhibitor of caspase-activated DNase (ICAD), as well as mitochondrial dysfunction, and that their action is abrogated by the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp.fluoromethylketone (z-VAD.fmk). These results suggest that inhibition of anti-apoptotic serine proteases governs the onset of the caspase-dependant apoptotic cascade. Secondly, we also demonstrate the involvement of a serine protease in the terminal stage of apoptosis. We showed that chymotrypsin-like protease activity is required for internucleosomal DNA fragmentation in apoptotic cells. Hence, DNA fragmentation is abrogated in TPCK-pre-treated WEHI 231 cells undergoing apoptosis triggered either by anti-IgM or TLCK. These results indicate that internucleosomal DNA cleavage in apoptotic cells is mediated by a chymotrypsin-like protease.
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PMID:Internucleosomal DNA cleavage in apoptotic WEHI 231 cells is mediated by a chymotrypsin-like protease. 1550 21


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