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Sample records for kinase-1 ask-1 rescues

  1. Production of recombinant human apoptosis signal-regulating kinase 1 (ASK1) in Escherichia coli.

    PubMed

    Volynets, Galyna P; Gorbatiuk, Oksana B; Kukharenko, Oleksandr P; Usenko, Mariya O; Yarmoluk, Sergiy M

    2016-10-01

    Apoptosis signal-regulating kinase 1 (ASK1) is a mediator of the MAPK signaling cascade, which regulates different cellular processes including apoptosis, cell survival, and differentiation. The increased activity of ASK1 is associated with a number of human diseases and this protein kinase is considered as promising therapeutic target. In the present study, the kinase domain of human ASK1 was expressed in Escherichia coli (E. coli) in soluble form. The expression level of ASK1 was around 0.3-0.47 g per 1 L after using auto-induction protocol or IPTG induction. A one-step on column method for the efficient purification of recombinant ASK1 was performed. Our approach yields sufficient amount of recombinant ASK1, which can be used for inhibitor screening assays and different crystallographic studies. PMID:27245507

  2. Targeting Apoptosis Signalling Kinase-1 (ASK-1) Does Not Prevent the Development of Neuropathy in Streptozotocin-Induced Diabetic Mice

    PubMed Central

    Newton, Victoria L.; Ali, Sumia; Duddy, Graham; Whitmarsh, Alan J.; Gardiner, Natalie J.

    2014-01-01

    Apoptosis signal-regulating kinase-1 (ASK1) is a mitogen-activated protein 3 kinase (MAPKKK/MAP3K) which lies upstream of the stress-activated MAPKs, JNK and p38. ASK1 may be activated by a variety of extracellular and intracellular stimuli. MAP kinase activation in the sensory nervous system as a result of diabetes has been shown in numerous preclinical and clinical studies. As a common upstream activator of both p38 and JNK, we hypothesised that activation of ASK1 contributes to nerve dysfunction in diabetic neuropathy. We therefore wanted to characterize the expression of ASK1 in sensory neurons, and determine whether the absence of functional ASK1 would protect against the development of neuropathy in a mouse model of experimental diabetes. ASK1 mRNA and protein is constitutively expressed by multiple populations of sensory neurons of the adult mouse lumbar DRG. Diabetes was induced in male C57BL/6 and transgenic ASK1 kinase-inactive (ASK1n) mice using streptozotocin. Levels of ASK1 do not change in the DRG, spinal cord, or sciatic nerve following induction of diabetes. However, levels of ASK2 mRNA increase in the spinal cord at 4 weeks of diabetes, which could represent a future target for this field. Neither motor nerve conduction velocity deficits, nor thermal or mechanical hypoalgesia were prevented or ameliorated in diabetic ASK1n mice. These results suggest that activation of ASK1 is not responsible for the nerve deficits observed in this mouse model of diabetic neuropathy. PMID:25329046

  3. ASK1: a new therapeutic target for kidney disease.

    PubMed

    Tesch, Greg H; Ma, Frank Y; Nikolic-Paterson, David J

    2016-08-01

    Stress-induced activation of p38 MAPK and JNK signaling is a feature of both acute and chronic kidney disease and is associated with disease progression. Inhibitors of p38 MAPK or JNK activation provide protection against inflammation and fibrosis in animal models of kidney disease; however, clinical trials of p38 MAPK and JNK inhibitors in other diseases (rheumatoid arthritis and pulmonary fibrosis) have been disappointing. Apoptosis signal-regulating kinase 1 (ASK1) acts as an upstream regulator for the activation of p38 MAPK and JNK in kidney disease. Mice lacking the Ask1 gene are healthy with normal homeostatic functions and are protected from acute kidney injury induced by ischemia-reperfusion and from renal interstitial fibrosis induced by ureteric obstruction. Recent studies have shown that a selective ASK1 inhibitor substantially reduced renal p38 MAPK activation and halted the progression of nephropathy in diabetic mice, and this has led to a current clinical trial of an ASK1 inhibitor in patients with stage 3 or 4 diabetic kidney disease. This review explores the rationale for targeting ASK1 in kidney disease and the therapeutic potential of ASK1 inhibitors based on current experimental evidence. PMID:27226108

  4. NQDI-1, an inhibitor of ASK1 attenuates acute perinatal hypoxic-ischemic cerebral injury by modulating cell death

    PubMed Central

    HAO, HU; LI, SITAO; TANG, HUI; LIU, BINGQING; CAI, YAO; SHI, CONGCONG; XIAO, XIN

    2016-01-01

    Apoptosis signal-regulating kinase 1 (ASK1) is a ubiquitously expressed protein kinase, which regulates cell fate in numerous injury conditions. Therefore, ASK1 may be a promising novel therapeutic target for injury. However, the expression and distribution of ASK1 in the perinatal brain following hypoxia-ischemia (HI) remains to be elucidated. In the present study, western blotting and immunofluorescence were used to determine the expression and distribution of ASK1 and any associated downstream targets in the perinatal rat brain following HI. NQDI-1, a specific inhibitor of ASK1 was intracerebroventricularly injected following neonatal rats brain insult for neuroprotection. The results revealed an increased expression of ASK1 and this expression was localized to the neurons and astrocytes, compared with the sham controls. Additionally, it was demonstrated that the ASK1/c-Jun N-terminal kinases (JNK) pathway was involved in the brain damage following HI in neonatal rats. Notably, NQDI-1 significantly inhibited the in vivo expression levels of ASK1, phosphorylated (p-)JNK, p-c-Jun, p53 and caspase 3. Reduced acute hypoxic-ischemic cerebral injury and cell apoptosis was observed following the injection of NQDI-1. Collectively, NQDI-1 attenuated acute perinatal hypoxic-ischemic cerebral injury by inhibiting the expression of ASK1 and cell apoptosis. This may be a promising novel neuroprotective inhibitor for perinatal cerebra injury. PMID:27081917

  5. Diarachidonoylphosphoethanolamine induces apoptosis of malignant pleural mesothelioma cells through a Trx/ASK1/p38 MAPK pathway.

    PubMed

    Tsuchiya, Ayako; Kaku, Yoshiko; Nakano, Takashi; Nishizaki, Tomoyuki

    2015-11-01

    1,2-Diarachidonoyl-sn-glycero-3-phosphoethanolamine (DAPE) induces both necrosis/necroptosis and apoptosis of NCI-H28 malignant pleural mesothelioma (MPM) cells. The present study was conducted to understand the mechanism for DAPE-induced apoptosis of NCI-H28 cells. DAPE induced caspase-independent apoptosis of NCI-H28 malignant pleural mesothelioma (MPM) cells, and the effect of DAPE was prevented by antioxidants or an inhibitor of NADPH oxidase (NOX). DAPE generated reactive oxygen species (ROS) and inhibited activity of thioredoxin (Trx) reductase (TrxR). DAPE decreased an association of apoptosis signal-regulating kinase 1 (ASK1) with thioredoxin (Trx), thereby releasing ASK1. DAPE activated p38 mitogen-activated protein kinase (MAPK), which was inhibited by an antioxidant or knocking-down ASK1. In addition, DAPE-induced NCI-H28 cell death was also prevented by knocking-down ASK1. Taken together, the results of the present study indicate that DAPE stimulates NOX-mediated ROS production and suppresses TrxR activity, resulting in the decrease of reduced Trx and the dissociation of ASK1 from a complex with Trx, allowing sequential activation of ASK1 and p38 MAPK, to induce apoptosis of NCI-H28 MPM cells. PMID:26588871

  6. Ask1 Gene Deletion Blocks Maternal Diabetes–Induced Endoplasmic Reticulum Stress in the Developing Embryo by Disrupting the Unfolded Protein Response Signalosome

    PubMed Central

    Wang, Fang; Wu, Yanqing; Gu, Hui; Reece, E. Albert; Fang, Shengyun; Gabbay-Benziv, Rinat; Aberdeen, Graham

    2015-01-01

    Apoptosis signal–regulating kinase 1 (ASK1) is activated by various stresses. The link between ASK1 activation and endoplasmic reticulum (ER) stress, two causal events in diabetic embryopathy, has not been determined. We sought to investigate whether ASK1 is involved in the unfolded protein response (UPR) that leads to ER stress. Deleting Ask1 abrogated diabetes-induced UPR by suppressing phosphorylation of inositol-requiring enzyme 1α (IRE1α), and double-stranded RNA-activated protein kinase (PKR)-like ER kinase (PERK) blocked the mitochondrial translocation of proapoptotic Bcl-2 members and ER stress. ASK1 participated in the IRE1α signalosome, and removing ASK1 abrogated the proapoptotic kinase activity of IRE1α. Ask1 deletion suppressed diabetes-induced IRE1α endoriboneclease activities, which led to X-box binding protein 1 mRNA cleavage, an ER stress marker, decreased expression of microRNAs, and increased expression of a miR-17 target, thioredoxin-interacting protein (Txnip), a thioredoxin binding protein, which enhanced ASK1 activation by disrupting the thioredoxin-ASK1 complexes. ASK1 is essential for the assembly and function of the IRE1α signalosome, which forms a positive feedback loop with ASK1 through Txnip. ASK1 knockdown in C17.2 neural stem cells diminished high glucose– or tunicamycin-induced IRE1α activation, which further supports our hypothesis that ASK1 plays a causal role in diabetes-induced ER stress and apoptosis. PMID:25249581

  7. MiR-17 Downregulation by High Glucose Stabilizes Thioredoxin-Interacting Protein and Removes Thioredoxin Inhibition on ASK1 Leading to Apoptosis.

    PubMed

    Dong, Daoyin; Fu, Noah; Yang, Peixin

    2016-03-01

    Pregestational diabetes significantly increases the risk of neural tube defects (NTDs). Maternal diabetes activates an Apoptosis Signal-regulating Kinase 1 (ASK1)-initiated pathway, which triggers neural stem cell apoptosis of the developing neuroepithelium leading to NTD formation. How high glucose of diabetes activates ASK1 is still unclear. In this study, we investigated the mechanism underlying high glucose-induced ASK1 activation. High glucose suppressed miR-17 expression, which led to an increase in its target gene Txnip (Thioredoxin-interacting protein). High glucose-increased Txnip enhanced its binding to the ASK1 inhibitor, thioredoxin (Trx), and thereby sequestered Trx from the Trx-ASK1 complex. High glucose-induced ASK1 activation and consequent apoptosis were abrogated by either the miR-17 mimic or Txnip siRNA knockdown. In contrast, the miR-17 inhibitor or Txnip ectopic overexpression mimicked the stimulative effect of high glucose on ASK1 and apoptosis. Thus, our study demonstrated that miR-17 repression mediates the pro-apoptotic effect of high glucose, and revealed a new mechanism underlying ASK1 activation, in which decreased miR-17 removes Trx inhibition on ASK1 through Txnip. PMID:26660634

  8. Klotho Regulates 14-3-3ζ Monomerization and Binding to the ASK1 Signaling Complex in Response to Oxidative Stress

    PubMed Central

    Brobey, Reynolds K.; Dheghani, Mehdi; Foster, Philip P.; Kuro-o, Makoto; Rosenblatt, Kevin P

    2015-01-01

    The reactive oxygen species (ROS)-sensitive apoptosis signal-regulating kinase 1 (ASK1) signaling complex is a key regulator of p38 MAPK activity, a major modulator of stress-associated with aging disorders. We recently reported that the ratio of free ASK1 to the complex-bound ASK1 is significantly decreased in Klotho-responsive manner and that Klotho-deficient tissues have elevated levels of free ASK1 which coincides with increased oxidative stress. Here, we tested the hypothesis that: 1) covalent interactions exist among three identified proteins constituting the ASK1 signaling complex; 2) in normal unstressed cells the ASK1, 14-3-3ζ and thioredoxin (Trx) proteins simultaneously engage in a tripartite complex formation; 3) Klotho’s stabilizing effect on the complex relied solely on 14-3-3ζ expression and its apparent phosphorylation and dimerization changes. To verify the hypothesis, we performed 14-3-3ζ siRNA knock-down experiments in conjunction with cell-based assays to measure ASK1-client protein interactions in the presence and absence of Klotho, and with or without an oxidant such as rotenone. Our results show that Klotho activity induces posttranslational modifications in the complex targeting 14-3-3ζ monomer/dimer changes to effectively protect against ASK1 oxidation and dissociation. This is the first observation implicating all three proteins constituting the ASK1 signaling complex in close proximity. PMID:26517365

  9. 4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to inhibit hepatocellular carcinoma cells.

    PubMed

    Fu, Meili; Wan, Fuqiang; Li, Zhengling; Zhang, Fenghua

    2016-03-01

    The aim of the present study is to investigate the potential anti-hepatocellular carcinoma (HCC) cell activity by 4SC-202, a novel class I HDAC inhibitor (HDACi). The associated signaling mechanisms were also analyzed. We showed that 4SC-202 treatment induced potent cytotoxic and proliferation-inhibitory activities against established HCC cell lines (HepG2, HepB3, SMMC-7721) and patient-derived primary HCC cells. Further, adding 4SC-202 in HCC cells activated mitochondrial apoptosis pathway, which was evidenced by mitochondrial permeability transition pore (mPTP) opening, cytochrome C cytosol release and caspase-3/-9 activation. Inhibition of this apoptosis pathway, by caspase-3/-9 inhibitors, mPTP blockers, or by shRNA-mediated knockdown of cyclophilin-D (Cyp-D, a key component of mPTP), significantly attenuated 4SC-202-induced HCC cell death and apoptosis. Reversely, over-expression of Cyp-D enhanced 4SC-202's sensitivity in HCC cells. Further studies showed that 4SC-202 induced apoptosis signal-regulating kinase 1 (ASK1) activation, causing it translocation to mitochondria and physical association with Cyp-D. This mitochondrial ASK1-Cyp-D complexation appeared required for mediating 4SC-202-induced apoptosis activation. ASK1 stable knockdown by targeted-shRNAs largely inhibited 4SC-202-induced mPTP opening, cytochrome C release, and following HCC cell apoptotic death. Together, we suggest that 4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to potently inhibit human HCC cells. PMID:26773495

  10. ASK1 signalling regulates brown and beige adipocyte function

    PubMed Central

    Hattori, Kazuki; Naguro, Isao; Okabe, Kohki; Funatsu, Takashi; Furutani, Shotaro; Takeda, Kohsuke; Ichijo, Hidenori

    2016-01-01

    Recent studies suggest that adult humans have active brown or beige adipocytes, the activation of which might be a therapeutic strategy for the treatment of diverse metabolic diseases. Here we show that the protein kinase ASK1 regulates brown and beige adipocytes function. In brown or white adipocytes, the PKA-ASK1-p38 axis is activated in response to cAMP signalling and contributes to the cell-autonomous induction of genes, including Ucp1. Global and fat-specific ASK1 deficiency leads to impaired metabolic responses, including thermogenesis and oxygen consumption, at the cell and whole-body levels, respectively. Our data thus indicate that the ASK1 signalling axis is a regulator of brown and beige adipocyte gene expression and function. PMID:27045525

  11. ASK1 signalling regulates brown and beige adipocyte function.

    PubMed

    Hattori, Kazuki; Naguro, Isao; Okabe, Kohki; Funatsu, Takashi; Furutani, Shotaro; Takeda, Kohsuke; Ichijo, Hidenori

    2016-01-01

    Recent studies suggest that adult humans have active brown or beige adipocytes, the activation of which might be a therapeutic strategy for the treatment of diverse metabolic diseases. Here we show that the protein kinase ASK1 regulates brown and beige adipocytes function. In brown or white adipocytes, the PKA-ASK1-p38 axis is activated in response to cAMP signalling and contributes to the cell-autonomous induction of genes, including Ucp1. Global and fat-specific ASK1 deficiency leads to impaired metabolic responses, including thermogenesis and oxygen consumption, at the cell and whole-body levels, respectively. Our data thus indicate that the ASK1 signalling axis is a regulator of brown and beige adipocyte gene expression and function. PMID:27045525

  12. ASK1 restores the antiviral activity of APOBEC3G by disrupting HIV-1 Vif-mediated counteraction

    PubMed Central

    Miyakawa, Kei; Matsunaga, Satoko; Kanou, Kazuhiko; Matsuzawa, Atsushi; Morishita, Ryo; Kudoh, Ayumi; Shindo, Keisuke; Yokoyama, Masaru; Sato, Hironori; Kimura, Hirokazu; Tamura, Tomohiko; Yamamoto, Naoki; Ichijo, Hidenori; Takaori-Kondo, Akifumi; Ryo, Akihide

    2015-01-01

    APOBEC3G (A3G) is an innate antiviral restriction factor that strongly inhibits the replication of human immunodeficiency virus type 1 (HIV-1). An HIV-1 accessory protein, Vif, hijacks the host ubiquitin–proteasome system to execute A3G degradation. Identification of the host pathways that obstruct the action of Vif could provide a new strategy for blocking viral replication. We demonstrate here that the host protein ASK1 (apoptosis signal-regulating kinase 1) interferes with the counteraction by Vif and revitalizes A3G-mediated viral restriction. ASK1 binds the BC-box of Vif, thereby disrupting the assembly of the Vif–ubiquitin ligase complex. Consequently, ASK1 stabilizes A3G and promotes its incorporation into viral particles, ultimately reducing viral infectivity. Furthermore, treatment with the antiretroviral drug AZT (zidovudine) induces ASK1 expression and restores the antiviral activity of A3G in HIV-1-infected cells. This study thus demonstrates a distinct function of ASK1 in restoring the host antiviral system that can be enhanced by AZT treatment. PMID:25901786

  13. Klotho Protects Dopaminergic Neuron Oxidant-Induced Degeneration by Modulating ASK1 and p38 MAPK Signaling Pathways

    PubMed Central

    Brobey, Reynolds K.; German, Dwight; Sonsalla, Patricia K.; Gurnani, Prem; Pastor, Johanne; Hsieh, C-C; Papaconstantinou, John; Foster, Philip P.; Kuro-o, Makoto; Rosenblatt, Kevin P.

    2015-01-01

    Klotho transgenic mice exhibit resistance to oxidative stress as measured by their urinal levels of 8-hydroxy-2-deoxyguanosine, albeit this anti-oxidant defense mechanism has not been locally investigated in the brain. Here, we tested the hypothesis that the reactive oxygen species (ROS)-sensitive apoptosis signal-regulating kinase 1 (ASK1)/p38 MAPK pathway regulates stress levels in the brain of these mice and showed that: 1) the ratio of free ASK1 to thioredoxin (Trx)-bound ASK1 is relatively lower in the transgenic brain whereas the reverse is true for the Klotho knockout mice; 2) the reduced p38 activation level in the transgene corresponds to higher level of ASK1-bound Trx, while the KO mice showed elevated p38 activation and lower level of–bound Trx; and 3) that 14-3-3ζ is hyper phosphorylated (Ser-58) in the transgene which correlated with increased monomer forms. In addition, we evaluated the in vivo robustness of the protection by challenging the brains of Klotho transgenic mice with a neurotoxin, MPTP and analyzed for residual neuron numbers and integrity in the substantia nigra pars compacta. Our results show that Klotho overexpression significantly protects dopaminergic neurons against oxidative damage, partly by modulating p38 MAPK activation level. Our data highlight the importance of ASK1/p38 MAPK pathway in the brain and identify Klotho as a possible anti-oxidant effector. PMID:26452228

  14. Klotho Protects Dopaminergic Neuron Oxidant-Induced Degeneration by Modulating ASK1 and p38 MAPK Signaling Pathways.

    PubMed

    Brobey, Reynolds K; German, Dwight; Sonsalla, Patricia K; Gurnani, Prem; Pastor, Johanne; Hsieh, C-C; Papaconstantinou, John; Foster, Philip P; Kuro-o, Makoto; Rosenblatt, Kevin P

    2015-01-01

    Klotho transgenic mice exhibit resistance to oxidative stress as measured by their urinal levels of 8-hydroxy-2-deoxyguanosine, albeit this anti-oxidant defense mechanism has not been locally investigated in the brain. Here, we tested the hypothesis that the reactive oxygen species (ROS)-sensitive apoptosis signal-regulating kinase 1 (ASK1)/p38 MAPK pathway regulates stress levels in the brain of these mice and showed that: 1) the ratio of free ASK1 to thioredoxin (Trx)-bound ASK1 is relatively lower in the transgenic brain whereas the reverse is true for the Klotho knockout mice; 2) the reduced p38 activation level in the transgene corresponds to higher level of ASK1-bound Trx, while the KO mice showed elevated p38 activation and lower level of-bound Trx; and 3) that 14-3-3ζ is hyper phosphorylated (Ser-58) in the transgene which correlated with increased monomer forms. In addition, we evaluated the in vivo robustness of the protection by challenging the brains of Klotho transgenic mice with a neurotoxin, MPTP and analyzed for residual neuron numbers and integrity in the substantia nigra pars compacta. Our results show that Klotho overexpression significantly protects dopaminergic neurons against oxidative damage, partly by modulating p38 MAPK activation level. Our data highlight the importance of ASK1/p38 MAPK pathway in the brain and identify Klotho as a possible anti-oxidant effector. PMID:26452228

  15. ASK1 modulates the expression of microRNA Let7A in microglia under high glucose in vitro condition

    PubMed Central

    Song, Juhyun; Lee, Jong Eun

    2015-01-01

    Hyperglycemia results in oxidative stress and leads to neuronal apoptosis in the brain. Diabetes studies show that microglia participate in the progression of neuropathogenesis through their involvement in inflammation in vivo and in vitro. In high-glucose-induced inflammation, apoptosis signal regulating kinase 1 (ASK1) triggers the release of apoptosis cytokines and apoptotic gene expression. MicroRNA-Let7A (miR-Let7A) is reported to be a regulator of inflammation. In the present study, we investigated whether miR-Let7A regulates the function of microglia by controlling ASK1 in response to high-glucose-induced oxidative stress. We performed reverse transcription (RT) polymerase chain reaction, Taqman assay, real-time polymerase chain reaction, and immunocytochemistry to confirm the alteration of microglia function. Our results show that miR-Let7A is associated with the activation of ASK1 and the expression of anti-inflammatory cytokine (interleukin (IL)-10) and Mycs (c-Myc and N-Myc). Thus, the relationship between Let-7A and ASK1 could be a novel target for enhancing the beneficial function of microglia in central nervous system (CNS) disorders. PMID:26041997

  16. Apoptosis signal-regulating kinase 1 mediates denbinobin-induced apoptosis in human lung adenocarcinoma cells

    PubMed Central

    Kuo, Chen-Tzu; Chen, Bing-Chang; Yu, Chung-Chi; Weng, Chih-Ming; Hsu, Ming-Jen; Chen, Chien-Chih; Chen, Mei-Chieh; Teng, Che-Ming; Pan, Shiow-Lin; Bien, Mauo-Ying; Shih, Chung-Hung; Lin, Chien-Huang

    2009-01-01

    In the present study, we explore the role of apoptosis signal-regulating kinase 1 (ASK1) in denbinobin-induced apoptosis in human lung adenocarcinoma (A549) cells. Denbinobin-induced cell apoptosis was attenuated by an ASK1 dominant-negative mutant (ASK1DN), two antioxidants (N-acetyl-L-cysteine (NAC) and glutathione (GSH)), a c-Jun N-terminal kinase (JNK) inhibitor (SP600125), and an activator protein-1 (AP-1) inhibitor (curcumin). Treatment of A549 cells with denbinobin caused increases in ASK1 activity and reactive oxygen species (ROS) production, and these effects were inhibited by NAC and GSH. Stimulation of A549 cells with denbinobin caused JNK activation; this effect was markedly inhibited by NAC, GSH, and ASK1DN. Denbinobin induced c-Jun phosphorylation, the formation of an AP-1-specific DNA-protein complex, and Bim expression. Bim knockdown using a bim short interfering RNA strategy also reduced denbinobin-induced A549 cell apoptosis. The denbinobin-mediated increases in c-Jun phosphorylation and Bim expression were inhibited by NAC, GSH, SP600125, ASK1DN, JNK1DN, and JNK2DN. These results suggest that denbinobin might activate ASK1 through ROS production to cause JNK/AP-1 activation, which in turn induces Bim expression, and ultimately results in A549 cell apoptosis. PMID:19405983

  17. Inhibition of apoptosis signal-regulating kinase 1 enhances endochondral bone formation by increasing chondrocyte survival.

    PubMed

    Eaton, G J; Zhang, Q-S; Diallo, C; Matsuzawa, A; Ichijo, H; Steinbeck, M J; Freeman, T A

    2014-01-01

    Endochondral ossification is the result of chondrocyte differentiation, hypertrophy, death and replacement by bone. The careful timing and progression of this process is important for normal skeletal bone growth and development, as well as fracture repair. Apoptosis Signal-Regulating Kinase 1 (ASK1) is a mitogen-activated protein kinase (MAPK), which is activated by reactive oxygen species and other cellular stress events. Activation of ASK1 initiates a signaling cascade known to regulate diverse cellular events including cytokine and growth factor signaling, cell cycle regulation, cellular differentiation, hypertrophy, survival and apoptosis. ASK1 is highly expressed in hypertrophic chondrocytes, but the role of ASK1 in skeletal tissues has not been investigated. Herein, we report that ASK1 knockout (KO) mice display alterations in normal growth plate morphology, which include a shorter proliferative zone and a lengthened hypertrophic zone. These changes in growth plate dynamics result in accelerated long bone mineralization and an increased formation of trabecular bone, which can be attributed to an increased resistance of terminally differentiated chondrocytes to undergo cell death. Interestingly, under normal cell culture conditions, mouse embryonic fibroblasts (MEFs) derived from ASK1 KO mice show no differences in either MAPK signaling or osteogenic or chondrogenic differentiation when compared with wild-type (WT) MEFs. However, when cultured with stress activators, H2O2 or staurosporine, the KO cells show enhanced survival, an associated decrease in the activation of proteins involved in death signaling pathways and a reduction in markers of terminal differentiation. Furthermore, in both WT mice treated with the ASK1 inhibitor, NQDI-1, and ASK1 KO mice endochondral bone formation was increased in an ectopic ossification model. These findings highlight a previously unrealized role for ASK1 in regulating endochondral bone formation. Inhibition of ASK1 has

  18. Thymoquinone inhibits TNF-α-induced inflammation and cell adhesion in rheumatoid arthritis synovial fibroblasts by ASK1 regulation

    SciTech Connect

    Umar, Sadiq; Hedaya, Omar; Singh, Anil K.; Ahmed, Salahuddin

    2015-09-15

    Tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine produced by monocytes/macrophage that plays a pathological role in rheumatoid arthritis (RA). In this study, we investigate the effect of thymoquinone (TQ), a phytochemical found in Nigella sativa, in regulating TNF-α-induced RA synovial fibroblast (RA-FLS) activation. Treatment with TQ (1–5 μM) had no marked effect on the viability of human RA-FLS. Pre-treatment of TQ inhibited TNF-α-induced interleukin-6 (IL-6) and IL-8 production and ICAM-1, VCAM-1, and cadherin-11 (Cad-11) expression in RA-FLS (p < 0.01). Evaluation of the signaling events showed that TQ inhibited TNF-α-induced phospho-p38 and phospho-JNK expression, but had no inhibitory effect on NF-κB pathway, in RA-FLS (p < 0.05; n = 4). Interestingly, we observed that selective down-regulation of TNF-α-induced phospho-p38 and phospho-JNK activation by TQ is elicited through inhibition of apoptosis-regulated signaling kinase 1 (ASK1). Furthermore, TNF-α selectively induced phosphorylation of ASK1 at Thr845 residue in RA-FLS, which was inhibited by TQ pretreatment in a dose dependent manner (p < 0.01). Pre-treatment of RA-FLS with ASK1 inhibitor (TC ASK10), blocked TNF-α induced expression of ICAM-1, VCAM-1, and Cad-11. Our results suggest that TNF-α-induced ASK1-p38/JNK pathway is an important mediator of cytokine synthesis and enhanced expression of adhesion molecule in RA-FLS and TQ, by selectively inhibiting this pathway, may have a potential therapeutic value in regulating tissue destruction observed in RA. - Highlights: • Evolving evidence suggests that ASK1 plays a central role in rheumatic arthritis (RA). • TNF-α activates ASK1, which regulate downstream signaling through JNK/p38 activation in RA-FLS. • ASK1 may be used as a potential therapeutic target in RA. • Thymoquinone was able to selectively inhibit TNF-α-induced phosphorylation of ASK1 in RA-FLS. • Thymoquinone might serve as a potential small

  19. C-terminus of heat shock protein 70– interacting protein facilitates degradation of apoptosis signal-regulating kinase 1 and inhibits apoptosis signal-regulating kinase 1– dependent apoptosis

    PubMed Central

    Hwang, Jae Ryoung; Zhang, Chunlian; Patterson, Cam

    2005-01-01

    Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase kinase kinase (MAPKKK) that is regulated under conditions of cellular stress. ASK1 phosphorylates c-Jun N-terminal kinase (JNK) and elicits an apoptotic response. ASK1 activity is regulated at multiple levels, 1 of which is through inhibition by cytosolic chaperones of the heat shock protein (Hsp) 70 family. Among the proteins that determine Hsp70 function, CHIP (C-terminus of Hsp70-interacting protein) is a cochaperone and ubiquitin ligase that interacts with Hsp70 through an amino-terminal tetratricopeptide repeat (TPR) domain. Prominent among the cellular functions mediated by CHIP is protection against physiologic stress. Because ASK1 is known to contain a TPR-acceptor site, we examined the role of CHIP in regulating ASK1 function. CHIP interacted with ASK1 in a TPR-dependent fashion and induced ubiquitylation and proteasome-dependent degradation of ASK1. Targeting of ASK1 by CHIP inhibited JNK activation in response to oxidative challenge and reduced ASK1-dependent apoptosis, whereas short interfering RNA (siRNA)-dependent depletion of CHIP enhanced JNK activation. Consistent with its ability to reduce cytoplasmic ASK1 levels, CHIP triggered the translocation of ASK1 partner protein death-associated protein (Daxx) into the nucleus, where it is known to activate an antiapoptotic response. These results indicate that CHIP regulates ASK1 activity by inducing its ubiquitylation and degradation, which, together with its effects on Daxx localization, provides a mechanism for the antiapoptotic effects of CHIP observed in the face of cellular and physiologic stress. PMID:16038411

  20. Thymoquinone inhibits TNF-α-induced inflammation and cell adhesion in rheumatoid arthritis synovial fibroblasts by ASK1 regulation.

    PubMed

    Umar, Sadiq; Hedaya, Omar; Singh, Anil K; Ahmed, Salahuddin

    2015-09-15

    Tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine produced by monocytes/macrophage that plays a pathological role in rheumatoid arthritis (RA). In this study, we investigate the effect of thymoquinone (TQ), a phytochemical found in Nigella sativa, in regulating TNF-α-induced RA synovial fibroblast (RA-FLS) activation. Treatment with TQ (1-5μM) had no marked effect on the viability of human RA-FLS. Pre-treatment of TQ inhibited TNF-α-induced interleukin-6 (IL-6) and IL-8 production and ICAM-1, VCAM-1, and cadherin-11 (Cad-11) expression in RA-FLS (p<0.01). Evaluation of the signaling events showed that TQ inhibited TNF-α-induced phospho-p38 and phospho-JNK expression, but had no inhibitory effect on NF-κB pathway, in RA-FLS (p<0.05; n=4). Interestingly, we observed that selective down-regulation of TNF-α-induced phospho-p38 and phospho-JNK activation by TQ is elicited through inhibition of apoptosis-regulated signaling kinase 1 (ASK1). Furthermore, TNF-α selectively induced phosphorylation of ASK1 at Thr845 residue in RA-FLS, which was inhibited by TQ pretreatment in a dose dependent manner (p<0.01). Pre-treatment of RA-FLS with ASK1 inhibitor (TC ASK10), blocked TNF-α induced expression of ICAM-1, VCAM-1, and Cad-11. Our results suggest that TNF-α-induced ASK1-p38/JNK pathway is an important mediator of cytokine synthesis and enhanced expression of adhesion molecule in RA-FLS and TQ, by selectively inhibiting this pathway, may have a potential therapeutic value in regulating tissue destruction observed in RA. PMID:26134265

  1. Biophysical and Structural Characterization of the Thioredoxin-binding Domain of Protein Kinase ASK1 and Its Interaction with Reduced Thioredoxin*

    PubMed Central

    Kosek, Dalibor; Kylarova, Salome; Psenakova, Katarina; Rezabkova, Lenka; Herman, Petr; Vecer, Jaroslav; Obsilova, Veronika; Obsil, Tomas

    2014-01-01

    Apoptosis signal-regulating kinase 1 (ASK1), a mitogen-activated protein kinase kinase kinase, plays a key role in the pathogenesis of multiple diseases. Its activity is regulated by thioredoxin (TRX1) but the precise mechanism of this regulation is unclear due to the lack of structural data. Here, we performed biophysical and structural characterization of the TRX1-binding domain of ASK1 (ASK1-TBD) and its complex with reduced TRX1. ASK1-TBD is a monomeric and rigid domain that forms a stable complex with reduced TRX1 with 1:1 molar stoichiometry. The binding interaction does not involve the formation of intermolecular disulfide bonds. Residues from the catalytic WCGPC motif of TRX1 are essential for complex stability with Trp31 being directly involved in the binding interaction as suggested by time-resolved fluorescence. Small-angle x-ray scattering data reveal a compact and slightly asymmetric shape of ASK1-TBD and suggest reduced TRX1 interacts with this domain through the large binding interface without inducing any dramatic conformational change. PMID:25037217

  2. Rescuing the duty to rescue.

    PubMed

    Rulli, Tina; Millum, Joseph

    2016-04-01

    Clinicians and health researchers frequently encounter opportunities to rescue people. Rescue cases can generate a moral duty to aid those in peril. As such, bioethicists have leveraged a duty to rescue for a variety of purposes. Yet, despite its broad application, the duty to rescue is underanalysed. In this paper, we assess the state of theorising about the duty to rescue. There are large gaps in bioethicists' understanding of the force, scope and justification of the two most cited duties to rescue-the individual duty of easy rescue and the institutional rule of rescue. We argue that the duty of easy rescue faces unresolved challenges regarding its force and scope, and the rule of rescue is indefensible. If the duty to rescue is to help solve ethical problems, these theoretical gaps must be addressed. We identify two further conceptions of the duty to rescue that have received less attention-an institutional duty of easy rescue and the professional duty to rescue. Both provide guidance in addressing force and scope concerns and, thereby, traction in answering the outstanding problems with the duty to rescue. We conclude by proposing research priorities for developing accounts of duties to rescue in bioethics. PMID:24790055

  3. Apoptosis signal-regulating kinase 1 mediates striatal degeneration via the regulation of C1q

    PubMed Central

    Cho, Kyoung Joo; Cheon, So Young; Kim, Gyung Whan

    2016-01-01

    Apoptosis signal-regulating kinase-1 (ASK1), an early signaling element in the cell death pathway, has been hypothesized to participate in the pathology of neurodegenerative diseases. The systemic administration of 3-nitropropionic acid (3-NP) facilitates the development of selective striatal lesions. However, it remains unclear whether specific neurons are selectively targeted in 3-NP-infused striatal degeneration. Recently, it has been proposed that complement-mediated synapse elimination may be reactivated aberrantly in the pathology of neurodegenerative diseases. We hypothesized that ASK1 is involved in striatal astrocyte reactivation; reactive astrocyte secretes molecules detrimental to neuron; and striatal neurons are more susceptible to these factors. Our results indicate that striatal astrocyte is reactivated and ASK1 level increases after 3-NP general and chronic infusion. Reactive striatal astrocyte increases TGF-beta differentially to cortex and striatum. ASK1 may be involved in regulation of astrocyte TGF-beta and it is linked to the C1q level in spatial and temporal, and moreover in the earlier stage of progressing striatal neuronal loss. Conclusively the present study suggests that ASK1 mediates 3-NP toxicity and regulates C1q level through the astrocyte TGF-beta. And also it may suggest that C1q level may be a surrogate of prediction marker representing neurodegenerative disease progress before developing behavioral impairment. PMID:26728245

  4. Effect of silibinin and vitamin E on the ASK1-p38 MAPK pathway in D-galactosamine/lipopolysaccharide induced hepatotoxicity.

    PubMed

    Hashem, Reem M; Hassanin, Kamel Ma; Rashed, Laila A; Mahmoud, Mohamed O; Hassan, Mohamed G

    2016-06-01

    Apoptosis signal-regulating kinase 1 (ASK1), a redox-sensor mitogen-activated protein kinase kinase kinase (MAPKKK) that activates p38 MAPK pathways in oxidative stress-induced hepatotoxicity in D-galactosamine/lipopolysaccharide (D-GalN/LPS) model, is a key central pathway in which specific targeting of ASK1 deactivation is of a great therapeutic potential. We tested the effect of silibinin and vitamin E in curative and prophylactic manner of treatment on the negative modulators of ASK1, thioredoxin1 (Trx1), thioredoxin reductase1 (TrxR1), and the protein phosphatase (PP5), whereas they have previously proven to have hepatoprotective effect. Either curative or prophylactic silibinin and vitamin E groups significantly decreased ASK1 and p38 MAPK levels through increasing the gene expression of Trx1, TrxR1, and PP5 to reduce the oxidative stress as demonstrated by decreasing the levels of NADPH oxidase 4 (NOX4), TBARS and conjugated diene with a concomitant increase in the levels of GSH, CAT, and SOD. These results were confirmed by histopathology examination which illustrated progressive degenerative changes of hepatocytes such as hydropic degeneration, vacuolation, pyknosis, karyolysis, and loss of architecture of some cells in D-GalN/LPS treatment, and these features were alleviated with silibinin and vitamin E administration. In conclusion, silibinin and vitamin E decreased ASK1-p38 MAPK pathway through deactivating the upstream signalling ASK1 molecule via increasing the levels of Trx1 and TrxR1 as well as the PP5 to alleviate in D-GalN/LPS induced hepatotoxicity. PMID:26941058

  5. Lys29-linkage of ASK1 by Skp1−Cullin 1−Fbxo21 ubiquitin ligase complex is required for antiviral innate response

    PubMed Central

    Yu, Zhou; Chen, Taoyong; Li, Xuelian; Yang, Mingjin; Tang, Songqing; Zhu, Xuhui; Gu, Yan; Su, Xiaoping; Xia, Meng; Li, Weihua; Zhang, Xuemin; Wang, Qingqing; Cao, Xuetao; Wang, Jianli

    2016-01-01

    Protein ubiquitination regulated by ubiquitin ligases plays important roles in innate immunity. However, key regulators of ubiquitination during innate response and roles of new types of ubiquitination (apart from Lys48- and Lys63-linkage) in control of innate signaling have not been clearly understood. Here we report that F-box only protein Fbxo21, a functionally unknown component of SCF (Skp1–Cul1–F-box protein) complex, facilitates Lys29-linkage and activation of ASK1 (apoptosis signal-regulating kinase 1), and promotes type I interferon production upon viral infection. Fbxo21 deficiency in mice cells impairs virus-induced Lys29-linkage and activation of ASK1, attenuates c-Jun N-terminal kinase (JNK) and p38 signaling pathway, and decreases the production of proinflammatory cytokines and type I interferon, resulting in reduced antiviral innate response and enhanced virus replication. Therefore Fbxo21 is required for ASK1 activation via Lys29-linkage of ASK1 during antiviral innate response, providing mechanistic insights into non-proteolytic roles of SCF complex in innate immune response. DOI: http://dx.doi.org/10.7554/eLife.14087.001 PMID:27063938

  6. Inhibitor of apoptosis signal-regulating kinase 1 protects against acetaminophen-induced liver injury

    SciTech Connect

    Xie, Yuchao; Ramachandran, Anup; Breckenridge, David G.; Liles, John T.; Lebofsky, Margitta; Farhood, Anwar; Jaeschke, Hartmut

    2015-07-01

    Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and liver injury as indicated by plasma ALT activities and area of necrosis over a 24 h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affecting the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5 h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. - Highlights: • Two ASK1 inhibitors protected against acetaminophen-induced liver injury. • The ASK1 inhibitors protect when used as pre- or post-treatment. • Protection by ASK1 inhibitor is

  7. Integration of Apoptosis Signal-Regulating Kinase 1-Mediated Stress Signaling with the Akt/Protein Kinase B-IκB Kinase Cascade

    PubMed Central

    Puckett, Mary C.; Goldman, Erinn H.; Cockrell, Lisa M.; Huang, Bei; Kasinski, Andrea L.; Du, Yuhong; Wang, Cun-Yu; Lin, Anning; Ichijo, Hidenori; Khuri, Fadlo

    2013-01-01

    Cellular processes are tightly controlled through well-coordinated signaling networks that respond to conflicting cues, such as reactive oxygen species (ROS), endoplasmic reticulum (ER) stress signals, and survival factors to ensure proper cell function. We report here a direct interaction between inhibitor of κB kinase (IKK) and apoptosis signal-regulating kinase 1 (ASK1), unveiling a critical node at the junction of survival, inflammation, and stress signaling networks. IKK can be activated by growth factor stimulation or tumor necrosis factor alpha engagement. IKK forms a complex with and phosphorylates ASK1 at a sensor site, Ser967, leading to the recruitment of 14-3-3, counteracts stress signal-triggered ASK1 activation, and suppresses ASK1-mediated functions. An inhibitory role of IKK in JNK signaling has been previously reported to depend on NF-κB-mediated gene expression. Our data suggest that IKK has a dual role: a transcription-dependent and a transcription-independent action in controlling the ASK1-JNK axis, coupling IKK to ROS and ER stress response. Direct phosphorylation of ASK1 by IKK also defines a novel IKK phosphorylation motif. Because of the intimate involvement of ASK1 in diverse diseases, the IKK/ASK1 interface offers a promising target for therapeutic development. PMID:23530055

  8. Space Rescue

    NASA Technical Reports Server (NTRS)

    Muratore, John F.

    2007-01-01

    Space Rescue has been a topic of speculation for a wide community of people for decades. Astronauts, aerospace engineers, diplomats, medical and rescue professionals, inventors and science fiction writers have all speculated on this problem. Martin Caidin's 1964 novel Marooned dealt with the problems of rescuing a crew stranded in low earth orbit. Legend at the Johnson Space Center says that Caidin's portrayal of a Russian attempt to save the American crew played a pivotal role in convincing the Russians to join the real joint Apollo-Soyuz mission. Space Rescue has been a staple in science fiction television and movies portrayed in programs such as Star Trek, Stargate-SG1 and Space 1999 and movies such as Mission To Mars and Red Planet. As dramatic and as difficult as rescue appears in fictional accounts, in the real world it has even greater drama and greater difficulty. Space rescue is still in its infancy as a discipline and the purpose of this chapter is to describe the issues associated with space rescue and the work done so far in this field. For the purposes of this chapter, the term space rescue will refer to any system which allows for rescue or escape of personnel from situations which endanger human life in a spaceflight operation. This will span the period from crew ingress prior to flight through crew egress postlanding. For the purposes of this chapter, the term primary system will refer to the spacecraft system that a crew is either attempting to escape from or from which an attempt is being made to rescue the crew.

  9. Involvement of ASK1 activation in apoptosis induced by NPe6-PDT

    NASA Astrophysics Data System (ADS)

    Liu, Lei; Zhang, Zhen-zhen; Zhang, Zhigang

    2010-02-01

    Photodynamic therapy (PDT) employing photosensiter N-aspartyl chlorin e6 (NPe6) can induce lysosome disruption and initiate apoptotic pathway. Apoptosis signal-regulating kinase (ASK1) is an important regulator of apoptosis in response to various stresses, such as reactive oxygen species (ROS), endoplasmic reticulum (ER) stress, lipopolysaccharide (LPS) and calcium influx. In this study, we investigated the molecular mechanisms of apoptosis induced by NPe6-PDT in ASTC-a-1 cells. The results showed that the activities of ASK1 increased in response to NPe6-PDT. Over-expression of wild-type or activated mutant of ASK1 could obviously decrease cell viability and increase cell death; while inhibition of ASK1 significantly decreased cell apoptosis. These results suggested that ASK1 plays an important role in apoptosis induced by NPe6-PDT.

  10. ASK1 promotes the contact hypersensitivity response through IL-17 production.

    PubMed

    Mizukami, Junya; Sato, Takehiro; Camps, Montserrat; Ji, Hong; Rueckle, Thomas; Swinnen, Dominique; Tsuboi, Ryoji; Takeda, Kohsuke; Ichijo, Hidenori

    2014-01-01

    Contact hypersensitivity (CHS) is a form of delayed-type hypersensitivity triggered by the response to reactive haptens (sensitization) and subsequent challenge (elicitation). Here, we show that ASK1 promotes CHS and that suppression of ASK1 during the elicitation phase is sufficient to attenuate CHS. ASK1 knockout (KO) mice exhibited impaired 2,4-dinitrofluorobenzene (DNFB)-induced CHS. The suppression of ASK1 activity during the elicitation phase through a chemical genetic approach or a specific inhibitory compound significantly reduced the CHS response to a level similar to that observed in ASK1 KO mice. The reduced response was concomitant with the strong inhibition of production of IL-17, a cytokine that plays an important role in CHS and other inflammatory diseases, from sensitized lymph node cells. These results suggest that ASK1 is relevant to the overall CHS response during the elicitation phase and that ASK1 may be a promising therapeutic target for allergic contact dermatitis and other IL-17-related inflammatory diseases. PMID:24736726

  11. Glycogen synthase kinase-3β antagonizes ROS-induced hepatocellular carcinoma cell death through suppression of the apoptosis signal-regulating kinase 1.

    PubMed

    Zhang, Na; Liu, Lu; Dou, Yueying; Song, Danqing; Deng, Hongbin

    2016-07-01

    Glycogen synthase kinase-3β (GSK-3β), a multifunctional kinase, is an important regulator of cancer cell survival. Apoptosis signal-regulating kinase 1 (ASK1) is also a key factor for controlling several cellular events including the cell cycle, senescence, and apoptosis, in response to reactive oxygen species (ROS). The role of GSK-3β regulating the activity and protein level of ASK1 in the cancer cells remains largely unexplored. In this study, we showed that GSK-3β inhibits ROS-induced hepatocellular carcinoma cell death by suppressing ASK1. We first found that ectopic expression of GSK-3β suppressed hydrogen peroxide (H2O2)-induced cell death in HepG2 cells and knockdown of endogenous GSK-3β expression exhibited opposite effects. Moreover, GSK-3β expression clearly inhibited H2O2-induced phosphorylation of ASK1 in HepG2 cells, in association with a decrease in ASK1 protein level. Further exploration revealed that GSK-3β induced ubiquitination and proteasome-dependent degradation of ASK1 via inhibition of ubiquitin-specific protease USP9X. Our results thus suggest that GSK-3β is a key factor involved in ASK1 activation and ROS-induced cell death. PMID:27221474

  12. Coupling between endocytosis and sphingosine kinase 1 recruitment.

    PubMed

    Shen, Hongying; Giordano, Francesca; Wu, Yumei; Chan, Jason; Zhu, Chen; Milosevic, Ira; Wu, Xudong; Yao, Kai; Chen, Bo; Baumgart, Tobias; Sieburth, Derek; De Camilli, Pietro

    2014-07-01

    Genetic studies have suggested a functional link between cholesterol/sphingolipid metabolism and endocytic membrane traffic. Here we show that perturbing the cholesterol/sphingomyelin balance in the plasma membrane results in the massive formation of clusters of narrow endocytic tubular invaginations positive for N-BAR proteins. These tubules are intensely positive for sphingosine kinase 1 (SPHK1). SPHK1 is also targeted to physiologically occurring early endocytic intermediates, and is highly enriched in nerve terminals, which are cellular compartments specialized for exo/endocytosis. Membrane recruitment of SPHK1 involves a direct, curvature-sensitive interaction with the lipid bilayer mediated by a hydrophobic patch on the enzyme's surface. The knockdown of SPHKs results in endocytic recycling defects, and a mutation that disrupts the hydrophobic patch of Caenorhabditis elegans SPHK fails to rescue the neurotransmission defects in loss-of-function mutants of this enzyme. Our studies support a role for sphingosine phosphorylation in endocytic membrane trafficking beyond the established function of sphingosine-1-phosphate in intercellular signalling. PMID:24929359

  13. Regulation of the death-associated protein kinase 1 expression and autophagy via ATF6 requires apoptosis signal-regulating kinase 1.

    PubMed

    Gade, Padmaja; Manjegowda, Srikanta B; Nallar, Shreeram C; Maachani, Uday B; Cross, Alan S; Kalvakolanu, Dhananjaya V

    2014-11-01

    The death-associated protein kinase 1 (DAPK1) is an important regulator of cell death and autophagy. Recently, we have identified that ATF6, an endoplasmic reticulum-resident transcription factor, in association with the transcription factor CEBP-β, regulates the gamma interferon (IFN-γ)-induced expression of Dapk1 (P. Gade et al., Proc. Natl. Acad. Sci. U. S. A. 109:10316-10321, 2012, doi.org/10.1073/pnas.1119273109). IFN-γ-induced proteolytic processing of ATF6 and phosphorylation of C/EBP-β were essential for the formation of a novel transcriptional complex that regulates DAPK1. Here, we report that IFN-γ activates the ASK1-MKK3/MKK6-p38 mitogen-activated protein kinase (MAPK) pathway for controlling the activity of ATF6. The terminal enzyme in this pathway, p38 MAPK, phosphorylates a critical threonine residue in ATF6 upstream of its DNA binding domain. ATF6 mutants defective for p38 MAPK phosphorylation fail to undergo proteolytic processing in the Golgi apparatus and drive IFN-γ-induced gene expression and autophagy. We also show that mice lacking Ask1 are highly susceptible to lethal bacterial infection owing to defective autophagy. Together, these results identify a novel host defense pathway controlled by IFN-γ signaling. PMID:25135476

  14. Inhibitor of Apoptosis Signal-Regulating Kinase 1 Protects Against Acetaminophen-induced Liver Injury

    PubMed Central

    Xie, Yuchao; Ramachandran, Anup; Breckenridge, David G.; Liles, John T.; Lebofsky, Margitta; Farhood, Anwar; Jaeschke, Hartmut

    2015-01-01

    Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and liver injury as indicated by plasma ALT activities and area of necrosis over a 24h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affected the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. PMID:25818599

  15. Inhibitor of apoptosis signal-regulating kinase 1 protects against acetaminophen-induced liver injury.

    PubMed

    Xie, Yuchao; Ramachandran, Anup; Breckenridge, David G; Liles, John T; Lebofsky, Margitta; Farhood, Anwar; Jaeschke, Hartmut

    2015-07-01

    Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and liver injury as indicated by plasma ALT activities and area of necrosis over a 24h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affecting the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. PMID:25818599

  16. ASK1 Inhibitor Halts Progression of Diabetic Nephropathy in Nos3-Deficient Mice.

    PubMed

    Tesch, Greg H; Ma, Frank Y; Han, Yingjie; Liles, John T; Breckenridge, David G; Nikolic-Paterson, David J

    2015-11-01

    p38 mitogen-activated protein kinase (MAPK) signaling promotes diabetic kidney injury. Apoptosis signal-regulating kinase (ASK)1 is one of the upstream kinases in the p38 MAPK-signaling pathway, which is activated by inflammation and oxidative stress, suggesting a possible role for ASK1 in diabetic nephropathy. In this study, we examined whether a selective ASK1 inhibitor can prevent the induction and progression of diabetic nephropathy in mice. Diabetes was induced in hypertensive endothelial nitric oxide synthase (Nos3)-deficient mice by five low-dose streptozotocin (STZ) injections. Groups of diabetic Nos3(-/-) mice received ASK1 inhibitor (GS-444217 delivered in chow) as an early intervention (2-8 weeks after STZ) or late intervention (weeks 8-15 after STZ). Control diabetic and nondiabetic Nos3(-/-) mice received normal chow. Treatment with GS-444217 abrogated p38 MAPK activation in diabetic kidneys but had no effect upon hypertension in Nos3(-/-) mice. Early intervention with GS-444217 significantly inhibited diabetic glomerulosclerosis and reduced renal dysfunction but had no effect on the development of albuminuria. Late intervention with GS-444217 improved renal function and halted the progression of glomerulosclerosis, renal inflammation, and tubular injury despite having no effect on established albuminuria. In conclusion, this study identifies ASK1 as a new therapeutic target in diabetic nephropathy to reduce renal inflammation and fibrosis independent of blood pressure control. PMID:26180085

  17. Rescue Equipment

    NASA Technical Reports Server (NTRS)

    1995-01-01

    The Lifeshear cutter, a rescue tool for freeing accident victims from wreckage, was developed under the Clinton Administration's Technology Reinvestment Program. Prior cutting equipment was cumbersome and expensive; the new cutter is 50 percent lighter and 70 percent cheaper. The cutter is pyrotechnically-actuated, using a miniature version of the power cartridges used for separation devices on the Space Shuttle and other NASA spacecraft. Hi-Shear Technology Corporation developed the cutter with the Jet Propulsion Laboratory and input from the City of Torrance (California) Fire Department.

  18. Glutathione S-transferase class mu regulation of apoptosis signal-related kinase 1 protein during VCD-induced ovotoxicity in neonatal rat ovaries

    PubMed Central

    Bhattacharya, Poulomi; Madden, Jill A.; Sen, Nivedita; Hoyer, Patricia B.; Keating, Aileen F.

    2013-01-01

    4-vinylcyclohexene diepoxide (VCD) destroys ovarian primordial and small primary follicles via apoptosis. In mice, VCD exposure induces ovarian mRNA expression of glutathione S-transferase (GST) family members, including isoform mu (Gstm). Extra-ovarian GSTM negatively regulates pro-apoptotic apoptosis signal-related kinase 1 (ASK1) through protein complex formation, which dissociates during stress, thereby initiating ASK1-induced apoptosis. The present study investigated the ovarian response of Gstm mRNA and protein to VCD. Induction of Ask1 mRNA at VCD-induced follicle loss onset was determined. Ovarian GSTM:ASK1 protein complex formation was investigated and VCD exposure effects thereon evaluated. Phosphatidylinositol-3 kinase (PI3K) regulation of GSTM protein was also studied. Postnatal day (PND) 4 rat ovaries were cultured in control media ±: 1) VCD (30 μM) for 2–8d; 2) VCD (30 μM) for 2d, followed by incubation in control media for 4d (acute VCD exposure); or 3) LY294002 (20 μM) for 6d. VCD exposure did not alter Gstm mRNA expression, however, GSTM protein increased (P < 0.05) after 6d of both the acute and chronic treatments. Ask1 mRNA increased (0.33-fold; P < 0.05) relative to control after 6d of VCD exposure. Ovarian GSTM:ASK1 protein complex formation was confirmed and, relative to control, the amount of GSTM bound to ASK1 increased 33% (P < 0.05) by chronic but with no effect of acute VCD exposure. PI3K inhibition increased (P < 0.05) GSTM protein by 40% and 71% on d4 and d6, respectively. These findings support involvement of GSTM in the ovarian response to VCD exposure, through regulation of pro-apoptotic ASK1. PMID:23274565

  19. Glutathione S-transferase class μ regulation of apoptosis signal-regulating kinase 1 protein during VCD-induced ovotoxicity in neonatal rat ovaries.

    PubMed

    Bhattacharya, Poulomi; Madden, Jill A; Sen, Nivedita; Hoyer, Patricia B; Keating, Aileen F

    2013-02-15

    4-Vinylcyclohexene diepoxide (VCD) destroys ovarian primordial and small primary follicles via apoptosis. In mice, VCD exposure induces ovarian mRNA expression of glutathione S-transferase (GST) family members, including isoform mu (Gstm). Extra-ovarian GSTM negatively regulates pro-apoptotic apoptosis signal-regulating kinase 1 (ASK1) through protein complex formation, which dissociates during stress, thereby initiating ASK1-induced apoptosis. The present study investigated the ovarian response of Gstm mRNA and protein to VCD. Induction of Ask1 mRNA at VCD-induced follicle loss onset was determined. Ovarian GSTM:ASK1 protein complex formation was investigated and VCD exposure effects thereon evaluated. Phosphatidylinositol-3 kinase (PI3K) regulation of GSTM protein was also studied. Postnatal day (PND) 4 rat ovaries were cultured in control media ± 1) VCD (30 μM) for 2-8 days; 2) VCD (30 μM) for 2 days, followed by incubation in control media for 4 days (acute VCD exposure); or 3) LY294002 (20 μM) for 6 days. VCD exposure did not alter Gstm mRNA expression, however, GSTM protein increased (P<0.05) after 6 days of both the acute and chronic treatments. Ask1 mRNA increased (0.33-fold; P<0.05) relative to control after 6 days of VCD exposure. Ovarian GSTM:ASK1 protein complex formation was confirmed and, relative to control, the amount of GSTM bound to ASK1 increased 33% (P<0.05) by chronic but with no effect of acute VCD exposure. PI3K inhibition increased (P<0.05) GSTM protein by 40% and 71% on d4 and d6, respectively. These findings support involvement of GSTM in the ovarian response to VCD exposure, through regulation of pro-apoptotic ASK1. PMID:23274565

  20. Checkpoint kinase 1 (Chk1) is required for mitotic progression through negative regulation of polo-like kinase 1 (Plk1)

    PubMed Central

    Tang, Jiabin; Erikson, Raymond L.; Liu, Xiaoqi

    2006-01-01

    Although the essential function of checkpoint kinase 1 (Chk1) in DNA damage response has been well established, the role of Chk1 in normal cell cycle progression is unclear. By using RNAi to specifically deplete Chk1, we determined loss-of-function phenotypes in HeLa cells. A vector-based RNAi approach showed that Chk1 is required for normal cell proliferation and survival, inasmuch as a dramatic cell-cycle arrest at G2/M phase and massive apoptosis were observed in Chk1-deficient cells. Coupling of siRNA with cell synchronization further revealed that Chk1 depletion leads to metaphase block, as indicated by various mitotic markers. Neither bipolar spindle formation nor centrosome functions were affected by Chk1 depletion; however, the depleted cells exhibited chromosome misalignment during metaphase, chromosome lagging during anaphase, and kinetochore defects within the regions of misaligned/lagging chromosomes. Moreover, we showed that Chk1 is a negative regulator of polo-like kinase 1 (Plk1), in either the absence or presence of DNA damage. Finally, Chk1 depletion leads to the activation of the spindle checkpoint because codepletion of spindle checkpoint proteins rescues the Chk1 depletion-induced mitotic arrest. PMID:16873548

  1. Rescue Manual. Module 8.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. Instructional Materials Lab.

    This learner manual for rescuers covers the current techniques or practices required in the rescue service. The eighth of 10 modules contains 6 chapters: (1) trench rescue; (2) shoring and tunneling techniques; (3) farm accident rescue; (4) wilderness search and rescue; (5) aircraft rescue; and (6) helicopter information. Key points, an…

  2. Rescue Manual. Module 9.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. Instructional Materials Lab.

    This learner manual for rescuers covers the current techniques or practices required in the rescue service. The ninth of 10 modules contains 7 chapters: (1) ice characteristics; (2) river characteristics and tactics for rescue; (3) water rescue techniques; (4) water rescue/recovery operations; (5) dive operations; (6) water rescue equipment; and…

  3. Platycodin D induces reactive oxygen species-mediated apoptosis signal-regulating kinase 1 activation and endoplasmic reticulum stress response in human breast cancer cells.

    PubMed

    Yu, Ji Sun; Kim, An Keun

    2012-08-01

    Platycodin D (PD), a natural compound found in Platycodon grandiflorum, induces apoptotic cell death in various carcinoma cells. One mechanism of PD-mediated cell death is by activation of mitogen-activated protein kinases, as suggested in a recent report. In this study, we further examined upstream signal pathways and the relationship between these signals and reactive oxygen species (ROS). Using immunoblotting assays, we found that PD activated apoptosis signal-regulating kinase 1 (ASK1) through phosphorylation of ASK1 at threonine and dephosphorylation of ASK1 at serine. We also showed that PD caused activation of the endoplasmic reticulum (ER) stress response. This was supported by observations showing that treatment with PD induces phosphorylation of PKR-like ER kinase (PERK) and eukaryotic initiation factor 2 α (eIF 2α), up-regulating expression of glucose-regulated protein 78/immunoglobulin heavy chain binding protein (GRP78/Bip) and CCAAT/enhancer-binding protein homologous protein/growth arrest and DNA damage-inducible gene 153 (CHOP/GADD153) and activation of caspase-4. Furthermore, PD-induced ASK1 and ER stress responses were inhibited by the antioxidant N-acetyl-l-cysteine. These results suggest that ROS play a critical role for activation of ASK1 and ER stress in PD-treated cancer cells. PMID:22784044

  4. Rescue Manual. Module 6.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. Instructional Materials Lab.

    This learner manual for rescuers covers the current techniques or practices required in the rescue service. The sixth of 10 modules contains 4 chapters: (1) industrial rescue; (2) rescue from a confined space; (3) extrication from heavy equipment; and (4) rescue operations involving elevators. Key points, an introduction, and conclusion accompany…

  5. Glutathione S-transferase class mu regulation of apoptosis signal-regulating kinase 1 protein during VCD-induced ovotoxicity in neonatal rat ovaries

    SciTech Connect

    Bhattacharya, Poulomi; Madden, Jill A.; Sen, Nivedita; Hoyer, Patricia B.; Keating, Aileen F.

    2013-02-15

    4-Vinylcyclohexene diepoxide (VCD) destroys ovarian primordial and small primary follicles via apoptosis. In mice, VCD exposure induces ovarian mRNA expression of glutathione S-transferase (GST) family members, including isoform mu (Gstm). Extra-ovarian GSTM negatively regulates pro-apoptotic apoptosis signal-regulating kinase 1 (ASK1) through protein complex formation, which dissociates during stress, thereby initiating ASK1-induced apoptosis. The present study investigated the ovarian response of Gstm mRNA and protein to VCD. Induction of Ask1 mRNA at VCD-induced follicle loss onset was determined. Ovarian GSTM:ASK1 protein complex formation was investigated and VCD exposure effects thereon evaluated. Phosphatidylinositol-3 kinase (PI3K) regulation of GSTM protein was also studied. Postnatal day (PND) 4 rat ovaries were cultured in control media ± 1) VCD (30 μM) for 2–8 days; 2) VCD (30 μM) for 2 days, followed by incubation in control media for 4 days (acute VCD exposure); or 3) LY294002 (20 μM) for 6 days. VCD exposure did not alter Gstm mRNA expression, however, GSTM protein increased (P < 0.05) after 6 days of both the acute and chronic treatments. Ask1 mRNA increased (0.33-fold; P < 0.05) relative to control after 6 days of VCD exposure. Ovarian GSTM:ASK1 protein complex formation was confirmed and, relative to control, the amount of GSTM bound to ASK1 increased 33% (P < 0.05) by chronic but with no effect of acute VCD exposure. PI3K inhibition increased (P < 0.05) GSTM protein by 40% and 71% on d4 and d6, respectively. These findings support involvement of GSTM in the ovarian response to VCD exposure, through regulation of pro-apoptotic ASK1. - Highlights: ► GSTM protein increases in response to ovarian VCD exposure. ► VCD increases Ask1 mRNA at the onset of follicle loss. ► Ovarian GSTM binds more ASK1 protein during VCD-induced ovotoxicity. ► PI3K regulates ovarian GSTM protein.

  6. The ASK1 gene regulates B function gene expression in cooperation with UFO and LEAFY in Arabidopsis.

    PubMed

    Zhao, D; Yu, Q; Chen, M; Ma, H

    2001-07-01

    The Arabidopsis floral regulatory genes APETALA3 (AP3) and PISTILLATA (PI) are required for the B function according to the ABC model for floral organ identity. AP3 and PI expression are positively regulated by the LEAFY (LFY) and UNUSUAL FLORAL ORGANS (UFO) genes. UFO encodes an F-box protein, and we have shown previously that UFO genetically interacts with the ASK1 gene encoding a SKP1 homologue; both the F-box containing protein and SKP1 are subunits of ubiquitin ligases. We show here that the ask1-1 mutation can enhance the floral phenotypes of weak lfy and ap3 mutants; therefore, like UFO, ASK1 also interacts with LFY and AP3 genetically. Furthermore, our results from RNA in situ hybridizations indicate that ASK1 regulates early AP3 and PI expression. These results support the idea that UFO and ASK1 together positively regulate AP3 and PI expression. We propose that the UFO and ASK1 proteins are components of a ubiquitin ligase that mediates the proteolysis of a repressor of AP3 and PI expression. Our genetic studies also indicate that ASK1 and UFO play a role in regulating the number of floral organ primordia, and we discuss possible mechanisms for such a regulation. PMID:11526079

  7. MYOSIN PHOSPHATASE TARGETING SUBUNIT1 REGULATES MITOSIS BY ANTAGONIZING POLO-LIKE KINASE1

    PubMed Central

    Yamashiro, S.; Yamakita, Y.; Totsukawa, G.; Goto, H.; Kaibuchi, K.; Ito, M.; Hartshorne, D.; Matsumura, F.

    2009-01-01

    SUMMARY Myosin phosphatase targeting subunit1 (MYPT1) binds to the catalytic subunit of protein phosphatase1 (PP1C). This binding is believed to target PP1C to specific substrates including myosin II, thus controlling cellular contractility. Surprisingly, we found that during mitosis mammalian MYPT1 binds to polo-like kinase1 (PLK1). MYPT1 is phosphorylated during mitosis by proline-directed kinases including cdc2, which generates the binding motif for the polo box domain of PLK1. Depletion of PLK1 by small interfering RNAs is known to results in loss of γ-tubulin recruitment to the centrosomes, blocking centrosome maturation, leading to mitotic arrest. We found that co-depletion of MYPT1 and PLK1 reinstates γ-tubulin at the centrosomes, rescuing the mitotic arrest. MYPT1 depletion increases phosphorylation of PLK1 at its activating site (Thr210) in vivo, explaining, at least in part, the rescue phenotype by co-depletion. Taken together, our results identify a previously unrecognized role for MYPT1 in regulating mitosis by antagonizing PLK1. PMID:18477460

  8. Rescue Skills and Techniques.

    ERIC Educational Resources Information Center

    Defense Civil Preparedness Agency (DOD), Washington, DC.

    The guide has been prepared for use as a textbook in rescue training courses at DCPA (Defense Civil Preparedness Agency) approved training schools and is to be used in rescue training programs of State and local governments. The document explains the various types of rescue missions, command structure, the personnel of the operating unit,…

  9. Rescue Manual. Module 4.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. Instructional Materials Lab.

    This learner manual for rescuers covers the current techniques or practices required in the rescue service. The fourth of 10 modules contains 8 chapters: (1) construction and characteristics of rescue rope; (2) knots, bends, and hitches; (3) critical angles; (4) raising systems; (5) rigging; (6) using the brake-bar rack for rope rescue; (7) rope…

  10. Zebrafish WNK Lysine Deficient Protein Kinase 1 (wnk1) Affects Angiogenesis Associated with VEGF Signaling

    PubMed Central

    Chen, Wen-Chuan; Kou, Fong-Ji; Lu, Jeng-Wei; Wang, Horng-Dar; Huang, Chou-Long; Yuh, Chiou-Hwa

    2014-01-01

    The WNK1 (WNK lysine deficient protein kinase 1) protein is a serine/threonine protein kinase with emerging roles in cancer. WNK1 causes hypertension and hyperkalemia when overexpressed and cardiovascular defects when ablated in mice. In this study, the role of Wnk1 in angiogenesis was explored using the zebrafish model. There are two zebrafish wnk1 isoforms, wnk1a and wnk1b, and both contain all the functional domains found in the human WNK1 protein. Both isoforms are expressed in the embryo at the initiation of angiogenesis and in the posterior cardinal vein (PCV), similar to fms-related tyrosine kinase 4 (flt4). Using morpholino antisense oligonucleotides against wnk1a and wnk1b, we observed that wnk1 morphants have defects in angiogenesis in the head and trunk, similar to flk1/vegfr2 morphants. Furthermore, both wnk1a and wnk1b mRNA can partially rescue the defects in vascular formation caused by flk1/vegfr2 knockdown. Mutation of the kinase domain or the Akt/PI3K phosphorylation site within wnk1 destroys this rescue capability. The rescue experiments provide evidence that wnk1 is a downstream target for Vegfr2 (vascular endothelial growth factor receptor-2) and Akt/PI3K signaling and thereby affects angiogenesis in zebrafish embryos. Furthermore, we found that knockdown of vascular endothelial growth factor receptor-2 (flk1/vegfr2) or vascular endothelial growth factor receptor-3 (flt4/vegfr3) results in a decrease in wnk1a expression, as assessed by in situ hybridization and q-RT-PCR analysis. Thus, the Vegf/Vegfr signaling pathway controls angiogenesis in zebrafish via Akt kinase-mediated phosphorylation and activation of Wnk1 as well as transcriptional regulation of wnk1 expression. PMID:25171174

  11. Receptor tyrosine kinase-like orphan receptor 1, a target of NKX2-1/TTF-1 lineage-survival oncogene, inhibits apoptosis signal-regulating kinase 1-mediated pro-apoptotic signaling in lung adenocarcinoma.

    PubMed

    Ida, Lisa; Yamaguchi, Tomoya; Yanagisawa, Kiyoshi; Kajino, Taisuke; Shimada, Yukako; Suzuki, Motoshi; Takahashi, Takashi

    2016-02-01

    We previously identified receptor tyrosine kinase-like orphan receptor 1 (ROR1) as a transcriptional target of the NKX2-1/TTF-1 lineage-survival oncogene in lung adenocarcinoma. ROR1 consequently sustains a favorable balance between pro-survival phosphatidylinositol 3-kinase-protein kinase B and pro-apoptotic apoptosis signal-regulating kinase 1 (ASK1)-p38MAPK signaling. In contrast to recent advances in understanding how ROR1 sustains pro-survival signaling, the mechanism of ROR1 repression of pro-apoptotic signaling remains rather elusive. In the present study, we investigated the underlying mechanism of ROR1-mediated inhibition of the ASK1-p38MAPK signaling pathway. Growth inhibition mediated by siROR1 was partially but significantly alleviated by ASK1 co-knockdown in lung adenocarcinoma cell lines. Also, ASK1 phosphorylation at Thr845, which reflects its activated state, was clearly inhibited by ROR1 overexpression in both steady state and oxidative stress-elicited conditions in MSTO-211H cells. In addition, we found that ROR1 was physically associated with ASK1 at the C-terminal serine threonine-rich domain of ROR1. Furthermore, ROR1 kinase activity was shown to be required to repress the ASK1-p38 axis and oxidative stress-induced cell death. The present findings thus support our notion that ROR1 sustains lung adenocarcinoma survival, at least in part, through direct physical interaction with ASK1 and consequential repression of the pro-apoptotic ASK1-p38 axis in a ROR1 kinase activity-dependent manner. PMID:26661061

  12. Apoptosis signal-regulating kinase 1 is mediated in TNF-α-induced CCL2 expression in human synovial fibroblasts.

    PubMed

    Tsou, Hsi-Kai; Chen, Hsien-Te; Chang, Chia-Hao; Yang, Wan-Yu; Tang, Chih-Hsin

    2012-11-01

    Tumor necrosis factor-α (TNF-α), a pro-inflammatory cytokine with a critical role in osteoarthritis (OA), was primarily produced by monocytes/macrophages and plays a crucial role in the inflammatory response. Here, we investigated the intracellular signaling pathways involved in TNF-α-induced monocyte chemoattractant protein 1 (MCP-1)/CCL2 expression in human synovial fibroblast cells. Stimulation of synovial fibroblasts (OASF) with TNF-α induced concentration- and time-dependent increases in CCL2 expression. TNF-α-mediated CCL2 production was attenuated by TNFR1 monoclonal antibody (Ab). Pretreatment with an apoptosis signal-regulating kinase 1 (ASK1) inhibitor (thioredoxin), JNK inhibitor (SP600125), p38 inhibitor (SB203580), or AP-1 inhibitor (curcumin or tanshinone IIA) also blocked the potentiating action of TNF-α. Stimulation of cells with TNF-α enhanced ASK1, JNK, and p38 activation. Treatment of OASF with TNF-α also increased the accumulation of phosphorylated c-Jun in the nucleus, AP-1-luciferase activity, and c-Jun binding to the AP-1 element on the CCL2 promoter. TNF-α-mediated AP-1-luciferase activity and c-Jun binding to the AP-1 element were inhibited by TNFR1 Ab, thioredoxin, SP600125, and SB203580. Our results suggest that the interaction between TNF-α and TNFR1 increases CCL2 expression in human synovial fibroblasts via the ASK1, JNK/p38, c-Jun, and AP-1 signaling pathway. PMID:22711527

  13. CLDN6-induced apoptosis via regulating ASK1-p38/JNK signaling in breast cancer MCF-7 cells.

    PubMed

    Guo, Yaxiong; Lin, Dongjing; Zhang, Mingzi; Zhang, Xiaowei; Li, Yanru; Yang, Ruan; Lu, Yan; Jin, Xiangshu; Yang, Minlan; Wang, Miaomiao; Zhao, Shuai; Quan, Chengshi

    2016-06-01

    Claudin 6 (CLDN6), a member of tight junction protein claudin (CLDN) family, inhibits proliferation and induces apoptosis of MCF-7 breast cancer cells. However, these molecular mechanisms of CLDN6-induced apoptosis remain largely elusive. We previously found that restoration of human CLDN6 gene expression was correlated with the expression level of apoptosis signal-regulating kinase 1 (ASK1) using cDNA array and bioinformatics analysis. ASK1, a mitogen-activated protein kinase kinase kinase, is involved in environmental stress-activation of the c-jun N-terminal kinase (JNK) and p38 pathways, which contribute to apoptosis-associated tumor cell death. In the present study, we show that the restoration of CLDN6 gene expression in MCF-7 cells marhedly decreased ASK1 phosphorylation at Ser967. Activated ASK1ser967 further induced the activation of downstream targets, JNK and p38 kinase. MCF-7/CLDN6 stable transfection cell clone treated with TRX1, an ASK1 inhibitor, showed suppressed JNK and p38 activation, and showed substantially increased survival and colony formation and reduced percent of apoptotic cells using TUNEL staining and DNA ladder. Furthermore, TRX1 treatment increased Bcl-2/Bax ratio and reduced caspase-3 cleavage in MCF-7/CLDN6 stable transfection cell clone. Therefore, these data show that CLDN6 mediates ASK1-p38/JNK apoptotic signaling in MCF-7 cells, and it is correlated with constitutive deregulation of the balance of Bcl-2 family proteins and activation of caspase-3. PMID:27035750

  14. Enhanced Rescue Lift Capability

    NASA Technical Reports Server (NTRS)

    Young, Larry A.

    2007-01-01

    The evolving and ever-increasing demands of emergency response and disaster relief support provided by rotorcraft dictate, among other things, the development of enhanced rescue lift capability for these platforms. This preliminary analysis is first-order in nature but provides considerable insight into some of the challenges inherent in trying to effect rescue using a unique form of robotic rescue device deployed and operated from rotary-wing aerial platforms.

  15. Sphingosine kinase-1--a potential therapeutic target in cancer.

    PubMed

    Cuvillier, Olivier

    2007-02-01

    Sphingolipid metabolites play critical functions in the regulation of a number of fundamental biological processes including cancer. Whereas ceramide and sphingosine mediate and trigger apoptosis or cell growth arrest, sphingosine 1-phosphate promotes proliferation and cell survival. The delicate equilibrium between the intracellular levels of each of these sphingolipids is controlled by the enzymes that either produce or degrade these metabolites. Sphingosine kinase-1 is a crucial regulator of this two-pan balance, because it produces the prosurvival sphingosine 1-phosphate, and reduces the content of both ceramide and sphingosine, the proapoptotic sphingolipids. Sphingosine kinase-1 controls the levels of sphingolipids having opposite effects on cell survival/death, its gene was found to be of oncogenic nature, its mRNA is overexpressed in many solid tumors, its overexpression protects cells from apoptosis and its activity is decreased during anticancer treatments. Therefore, sphingosine kinase-1 appears to be a target of interest for therapeutic manipulation via its pharmacological inhibition. Strategies to kill tumor cells by increasing their ceramide and/or sphingosine content while blocking sphingosine 1-phosphate generation should have a favorable therapeutic index. PMID:17159597

  16. The ASK1-specific inhibitors K811 and K812 prolong survival in a mouse model of amyotrophic lateral sclerosis.

    PubMed

    Fujisawa, Takao; Takahashi, Motoo; Tsukamoto, Yuka; Yamaguchi, Namiko; Nakoji, Masayoshi; Endo, Megumi; Kodaira, Hiroshi; Hayashi, Yuki; Nishitoh, Hideki; Naguro, Isao; Homma, Kengo; Ichijo, Hidenori

    2016-01-15

    Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with no cure. To develop effective treatments for this devastating disease, an appropriate strategy for targeting the molecule responsible for the pathogenesis of ALS is needed. We previously reported that mutant SOD1 protein causes motor neuron death through activation of ASK1, a mitogen-activated protein kinase kinase kinase. Additionally, we recently developed K811 and K812, which are selective inhibitors for ASK1. Here, we report the effect of K811 and K812 in a mouse model of ALS (SOD1(G93A) transgenic mice). Oral administration of K811 or K812 significantly extended the life span of SOD1(G93A) transgenic mice (1.06 and 1.08% improvement in survival). Moreover, ASK1 activation observed in the lumbar spinal cord of mice at the disease progression stage was markedly decreased in the K811- and K812-treated groups. In parallel, immunohistochemical analysis revealed that K811 and K812 treatment inhibited glial activation in the lumbar spinal cord of SOD1(G93A) transgenic mice. These results reinforce the importance of ASK1 as a therapeutic target for ALS treatment. PMID:26604152

  17. Rescue Manual. Module 2.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. Instructional Materials Lab.

    This learner manual for rescuers covers the current techniques or practices required in the rescue service. The second of 10 modules contains 5 chapters: (1) patient care and handling techniques; (2) rescue carries and drags; (3) emergency vehicle operations; (4) self-contained breathing apparatus; and (5) protective clothing. Key points, an…

  18. Palmitoylation of LIM Kinase-1 ensures spine-specific actin polymerization and morphological plasticity

    PubMed Central

    George, Joju; Soares, Cary; Montersino, Audrey; Beique, Jean-Claude; Thomas, Gareth M

    2015-01-01

    Precise regulation of the dendritic spine actin cytoskeleton is critical for neurodevelopment and neuronal plasticity, but how neurons spatially control actin dynamics is not well defined. Here, we identify direct palmitoylation of the actin regulator LIM kinase-1 (LIMK1) as a novel mechanism to control spine-specific actin dynamics. A conserved palmitoyl-motif is necessary and sufficient to target LIMK1 to spines and to anchor LIMK1 in spines. ShRNA knockdown/rescue experiments reveal that LIMK1 palmitoylation is essential for normal spine actin polymerization, for spine-specific structural plasticity and for long-term spine stability. Palmitoylation is critical for LIMK1 function because this modification not only controls LIMK1 targeting, but is also essential for LIMK1 activation by its membrane-localized upstream activator PAK. These novel roles for palmitoylation in the spatial control of actin dynamics and kinase signaling provide new insights into structural plasticity mechanisms and strengthen links between dendritic spine impairments and neuropathological conditions. DOI: http://dx.doi.org/10.7554/eLife.06327.001 PMID:25884247

  19. PAR-1 antagonist SCH79797 ameliorates apoptosis following surgical brain injury through inhibition of ASK1-JNK in rats

    PubMed Central

    Manaenko, Anatol; Sun, Xuejun; Kim, Cherine; Yan, Junhao; Ma, Qingyi

    2012-01-01

    Neurosurgical procedures inevitably produce intraoperative hemorrhage. The subsequent entry of blood into the brain parenchyma results in the release of large amounts of thrombin, a known contributor to perihematomal edema formation and apoptosis after brain injury. The present study seeks to test 1) the effect of surgically induced brain injury (SBI) on thrombin activity, expression of thrombin’s receptor PAR-1, and PAR-1 mediated apoptosis; 2) the effect of thrombin inhibition by argatroban and PAR-1 inhibition by SCH79797 on the development of secondary brain injury in the SBI model on rats. A total of 88 Sprague-Dawley male rats were randomly divided into sham, vehicle-, argatroban-, or SCH79797-treated groups. SBI involved partial resection of the right frontal lobe under inhalation isoflurane anesthesia. Sham-operated animals received only craniotomy. Thrombin activity, brain water content, and neurological deficits were measured at 24 hours following SBI. Involvement of the Ask1/JNK pathway in PAR-1-induced post-SBI apoptosis was characterized by using Ask1 or JNK inhibitors. We observed that SBI increased thrombin activity, yet failed to demonstrate any effect on PAR-1 expression. Argatroban and SCH79797 reduced SBI-induced brain edema and neurological deficits in a dose-dependent manner. SBI-induced apoptosis seemed mediated by the PAR-1/Ask1/JNK pathways. Administration of SCH79797 ameliorated the apoptosis following SBI. Our finding indicate that PAR-1 antagonist protects against secondary brain injury after SBI by decreasing both brain edema and apoptosis by inactivating PAR-1/Ask1/JNK pathway. The anti-apoptotic effect of PAR-1 antagonists may provide a promising path for therapy following SBI. PMID:23000356

  20. Countries renew rescue agreement

    NASA Astrophysics Data System (ADS)

    Bush, Susan M.

    To insure long-term continuity for the international satellite search and rescue system, COSPAS/SARSAT, an intergovernmental agreement binding the four sponsoring nations to cooperate was signed July 1 in Paris. According to Russell Vollmers of the National Oceanic and Atmospheric Administration, the agreement is binding for 15 years, with an automatic extension.The system marked the fifth anniversary of its first rescue last year, when on September 10, 1982, three persons were rescued. Begun in the 1970s by NASA as an experiment, COSPAS/SARSAT (a Russian-English acronym) is now a cooperative project among the United States, Canada, France, and the Soviet Union. Its goal is to reduce the time required to rescue air and maritime distress victims and also to locate victims who otherwise may not be found, thus using the satellite system as a life-saving device.

  1. Thioredoxin inhibits MPK38-induced ASK1, TGF-β, and p53 function in a phosphorylation-dependent manner.

    PubMed

    Manoharan, Ravi; Seong, Hyun-A; Ha, Hyunjung

    2013-10-01

    Murine protein serine-threonine kinase 38 (MPK38) is a member of the AMP-activated protein kinase-related serine/threonine kinase family. The factors that regulate MPK38 activity and function are not yet elucidated. Here, thioredoxin (Trx) was shown to be a negative regulator of MPK38. The redox-dependent association of MPK38 and Trx was mediated through the C-terminal domain of MPK38. Single and double amino acid substitution mutagenesis of MPK38 (C286S, C339S, C377S, and C339S/C377S) and Trx (C32S, C35S, and C32S/C35S) demonstrated that Cys(339) and Cys(377) of MPK38 and Cys(32) and Cys(35) of Trx are required for MPK38-Trx complex formation. MPK38 directly interacted with and phosphorylated Trx at Thr(76). Expression of wild-type Trx, but not the Trx mutants C32S/C35S and T76A, inhibited MPK38-induced ASK1, TGF-β, and p53 function by destabilizing MPK38. The E3 ubiquitin-protein ligase Mdm2 played a critical role in the regulation of MPK38 stability by Trx. Treatment of cells with 1-chloro-2,4-dinitrobenzene, a specific inhibitor of Trx reductase, decreased MPK38-Trx complex formation and subsequently increased MPK38 stability and activity, indicating that Trx negatively regulates MPK38 activity in vivo. Finally, we used ASK1-, Smad3-, and p53-null mouse embryonic fibroblasts to demonstrate that ASK1, Smad3, and p53 play important roles in the activity and function of MPK38, suggesting a functional link between MPK38 and ASK1, TGF-β, and p53 signaling pathways. These results indicate that Trx functions as a physiological inhibitor of MPK38, which plays an important role in inducing ASK1-, TGF-β-, and p53-mediated activity. PMID:23747528

  2. Ribosomal protein S6 kinase 1 signaling regulates mammalian lifespan

    PubMed Central

    Selman, Colin; Tullet, Jennifer M.A.; Wieser, Daniela; Irvine, Elaine; Lingard, Steven J.; Choudhury, Agharul I.; Claret, Marc; Al-Qassab, Hind; Carmignac, Danielle; Ramadani, Faruk; Woods, Angela; Robinson, Iain C.A.; Schuster, Eugene; Batterham, Rachel L.; Kozma, Sara C.; Thomas, George; Carling, David; Okkenhaug, Klaus; Thornton, Janet M.; Partridge, Linda; Gems, David; Withers, Dominic J.

    2016-01-01

    Caloric restriction (CR) protects against aging and disease but the mechanisms by which this affects mammalian lifespan are unclear. We show in mice that deletion of the nutrient-responsive mTOR (mammalian target of rapamycin) signaling pathway component ribosomal S6 protein kinase 1 (S6K1) led to increased lifespan and resistance to age-related pathologies such as bone, immune and motor dysfunction and loss of insulin sensitivity. Deletion of S6K1 induced gene expression patterns similar to those seen in CR or with pharmacological activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK), a conserved regulator of the metabolic response to CR. Our results demonstrate that S6K1 influences healthy mammalian lifespan, and suggest therapeutic manipulation of S6K1 and AMPK might mimic CR and provide broad protection against diseases of aging. PMID:19797661

  3. Sphingosine kinase-1 mediates androgen-induced osteoblast cell growth.

    PubMed

    Martin, Claire; Lafosse, Jean-Michel; Malavaud, Bernard; Cuvillier, Olivier

    2010-01-01

    Herein we report that the lipid kinase sphingosine kinase-1 (SphK1) is instrumental in mediating androgen-induced cell proliferation in osteoblasts. Dihydrotestosterone (DHT) triggered cell growth in steroid-deprived MC3T3 cells, which was associated with a rapid stimulation of SphK1 and activation of both Akt and ERK signaling pathways. This mechanism relied on functional androgen receptor/PI3K/Akt nongenotropic signaling as pharmacological antagonists could block SphK1 stimulation by DHT and its consequences. Finally, SphK1 inhibition not only abrogated DHT-induced ERK activation but also blocked cell proliferation, while ERK inhibition had no impact, suggesting that SphK1 was critical for DHT signaling yet independently of the ERK. PMID:19932089

  4. Sphingosine kinase-1 mediates androgen-induced osteoblast cell growth

    SciTech Connect

    Martin, Claire; Lafosse, Jean-Michel; Malavaud, Bernard; Cuvillier, Olivier

    2010-01-01

    Herein we report that the lipid kinase sphingosine kinase-1 (SphK1) is instrumental in mediating androgen-induced cell proliferation in osteoblasts. Dihydrotestosterone (DHT) triggered cell growth in steroid-deprived MC3T3 cells, which was associated with a rapid stimulation of SphK1 and activation of both Akt and ERK signaling pathways. This mechanism relied on functional androgen receptor/PI3K/Akt nongenotropic signaling as pharmacological antagonists could block SphK1 stimulation by DHT and its consequences. Finally, SphK1 inhibition not only abrogated DHT-induced ERK activation but also blocked cell proliferation, while ERK inhibition had no impact, suggesting that SphK1 was critical for DHT signaling yet independently of the ERK.

  5. Sphingosine kinase 1 as an anticancer therapeutic target

    PubMed Central

    Gao, Ying; Gao, Fei; Chen, Kan; Tian, Mei-li; Zhao, Dong-li

    2015-01-01

    The development of chemotherapeutic resistance is a major challenge in oncology. Elevated sphingosine kinase 1 (SK1) levels is predictive of a poor prognosis, and SK1 overexpression may confer resistance to chemotherapeutics. The SK/sphingosine-1-phosphate (S1P)/sphingosine-1-phosphate receptor (S1PR) signaling pathway has been implicated in the progression of various cancers and in chemotherapeutic drug resistance. Therefore, SK1 may represent an important target for cancer therapy. Targeting the SK/S1P/S1PR signaling pathway may be an effective anticancer therapeutic strategy, particularly in the context of overcoming drug resistance. This review summarizes our current understanding of the role of SK/S1P/S1PR signaling in cancer and development of SK1 inhibitors. PMID:26150697

  6. Hesperetin Induces Apoptosis in Breast Carcinoma by Triggering Accumulation of ROS and Activation of ASK1/JNK Pathway.

    PubMed

    Palit, Shreyasi; Kar, Susanta; Sharma, Gunjan; Das, Pijush K

    2015-08-01

    Hesperetin, a flavanone glycoside predominantly found in citrus fruits, exhibits a wide array of biological properties. In the present study hesperetin exhibited a significant cytotoxic effect in human breast carcinoma MCF-7 cells in a concentration- and time-dependent manner without affecting normal (HMEC) as well as immortalized normal mammary epithelial cells (MCF-10A). The cytotoxic effect of hesperetin was due to the induction of apoptosis as evident from the phosphatidyl-serine externalization, DNA fragmentation, caspase-7 activation, and PARP cleavage. Apoptosis was associated with caspase-9 activation, mitochondrial membrane potential loss, release of cytochrome c, and increase in Bax:Bcl-2 ratio. Pre-treatment with caspase-9 specific inhibitor (Z-LEHD-fmk) markedly attenuated apoptosis suggesting an involvement of intrinsic mitochondrial apoptotic cascade. Further, DCFDA flow-cytometric analysis revealed triggering of ROS in a time-dependent manner. Pre-treatment with ROS scavenger N-acetylcysteine (NAC) and glutathione markedly abrogated hesperetin-mediated apoptosis whereas carbonyl cyanide m-chlorophenylhydrazone (CCCP) pretreatment along with DHR123-based flow-cytometry indicated the generation of cytosolic ROS. Profiling of MAPKs revealed activation of JNK upon hesperetin treatment which was abrogated upon NAC pre-treatment. Additionally, inhibition of JNK by SP600125 significantly reversed hesperetin-mediated apoptosis. The activation of JNK was associated with the activation of ASK1. Silencing of ASK1 resulted in significant attenuation of JNK activation as well as reversed the hesperetin-mediated apoptosis suggesting that hesperetin-mediated apoptosis of MCF-7 cells involves accumulation of ROS and activation of ASK1/JNK pathway. In addition, hesperetin also induced apoptosis in triple negative breast cancer MDA-MB-231 cells via intrinsic pathway via activation of caspase -9 and -3 and increase in Bax:Bcl-2 ratio. PMID:25204891

  7. Airborne rescue system

    NASA Technical Reports Server (NTRS)

    Haslim, Leonard A. (Inventor)

    1991-01-01

    The airborne rescue system includes a boom with telescoping members for extending a line and collar to a rescue victim. The boom extends beyond the tip of the helicopter rotor so that the victim may avoid the rotor downwash. The rescue line is played out and reeled in by winch. The line is temporarily retained under the boom. When the boom is extended, the rescue line passes through clips. When the victim dons the collar and the tension in the line reaches a predetermined level, the clips open and release the line from the boom. Then the rescue line can form a straight line between the victim and the winch, and the victim can be lifted to the helicopter. A translator is utilized to push out or pull in the telescoping members. The translator comprises a tape and a rope. Inside the telescoping members the tape is curled around the rope and the tape has a tube-like configuration. The tape and rope are provided from supply spools.

  8. Activation-Induced Cell Death of Dendritic Cells Is Dependent on Sphingosine Kinase 1

    PubMed Central

    Schwiebs, Anja; Friesen, Olga; Katzy, Elisabeth; Ferreirós, Nerea; Pfeilschifter, Josef M.; Radeke, Heinfried H.

    2016-01-01

    Sphingosine 1-phosphate (S1P) is an immune modulatory lipid mediator and has been implicated in numerous pathophysiological processes. S1P is produced by sphingosine kinase 1 (Sphk1) and Sphk2. Dendritic cells (DCs) are central for the direction of immune responses and crucially involved in autoimmunity and cancerogenesis. In this study we examined the function and survival of bone marrow-derived DCs under long-term inflammatory stimulation. We observed that differentiated cells undergo activation-induced cell death (AICD) upon LPS stimulation with an increased metabolic activity shortly after stimulation, followed by a rapid activation of caspase 3 and subsequent augmented apoptosis. Importantly, we highlight a profound role of Sphk1 in secretion of inflammatory cytokines and survival of dendritic cells that might be mediated by a change in sphingolipid levels as well as by a change in STAT3 expression. Cell growth during differentiation of Sphk1-deficient cells treated with the functional S1P receptor antagonist FTYP was reduced. Importantly, in dendritic cells we did not observe a compensatory regulation of Sphk2 mRNA in Sphk1-deficient cells. Instead, we discovered a massive increase in Sphk1 mRNA concentration upon long-term stimulation with LPS in wild type cells that might function as an attempt to rescue from inflammation-caused cell death. Taken together, in this investigation we describe details of a crucial involvement of sphingolipids and Sphk1 in AICD during long-term immunogenic activity of DCs that might play an important role in autoimmunity and might explain the differences in immune response observed in in vivo studies of Sphk1 modulation. PMID:27148053

  9. Arabidopsis Receptor of Activated C Kinase1 Phosphorylation by WITH NO LYSINE8 KINASE

    DOE PAGESBeta

    Urano, Daisuke; Czarnecki, Olaf; Wang, Xiaoping; Jones, Alan M.; Chen, Jin-Gui

    2014-12-08

    Receptor of activated C kinase1 (RACK1) is a versatile scaffold protein that binds to numerous proteins to regulate diverse cellular pathways in mammals. In Arabidopsis (Arabidopsis thaliana), RACK1 has been shown to regulate plant hormone signaling, stress responses, and multiple processes of growth and development. However, little is known about the molecular mechanism underlying these regulations. In this paper, we show that an atypical serine (Ser)/threonine (Thr) protein kinase, WITH NO LYSINE8 (WNK8), phosphorylates RACK1. WNK8 physically interacted with and phosphorylated RACK1 proteins at two residues: Ser-122 and Thr-162. Genetic epistasis analysis of rack1 wnk8 double mutants indicated that RACK1more » acts downstream of WNK8 in the glucose responsiveness and flowering pathways. The phosphorylation-dead form, RACK1AS122A/T162A, but not the phosphomimetic form, RACK1AS122D/T162E, rescued the rack1a null mutant, implying that phosphorylation at Ser-122 and Thr-162 negatively regulates RACK1A function. The transcript of RACK1AS122D/T162E accumulated at similar levels as those of RACK1S122A/T162A. However, although the steady-state level of the RACK1AS122A/T162A protein was similar to wild-type RACK1A protein, the RACK1AS122D/T162E protein was nearly undetectable, suggesting that phosphorylation affects the stability of RACK1A proteins. In conclusion, these results suggest that RACK1 is phosphorylated by WNK8 and that phosphorylation negatively regulates RACK1 function by influencing its protein stability.« less

  10. Arabidopsis Receptor of Activated C Kinase1 Phosphorylation by WITH NO LYSINE8 KINASE

    SciTech Connect

    Urano, Daisuke; Czarnecki, Olaf; Wang, Xiaoping; Jones, Alan M.; Chen, Jin-Gui

    2014-12-08

    Receptor of activated C kinase1 (RACK1) is a versatile scaffold protein that binds to numerous proteins to regulate diverse cellular pathways in mammals. In Arabidopsis (Arabidopsis thaliana), RACK1 has been shown to regulate plant hormone signaling, stress responses, and multiple processes of growth and development. However, little is known about the molecular mechanism underlying these regulations. In this paper, we show that an atypical serine (Ser)/threonine (Thr) protein kinase, WITH NO LYSINE8 (WNK8), phosphorylates RACK1. WNK8 physically interacted with and phosphorylated RACK1 proteins at two residues: Ser-122 and Thr-162. Genetic epistasis analysis of rack1 wnk8 double mutants indicated that RACK1 acts downstream of WNK8 in the glucose responsiveness and flowering pathways. The phosphorylation-dead form, RACK1AS122A/T162A, but not the phosphomimetic form, RACK1AS122D/T162E, rescued the rack1a null mutant, implying that phosphorylation at Ser-122 and Thr-162 negatively regulates RACK1A function. The transcript of RACK1AS122D/T162E accumulated at similar levels as those of RACK1S122A/T162A. However, although the steady-state level of the RACK1AS122A/T162A protein was similar to wild-type RACK1A protein, the RACK1AS122D/T162E protein was nearly undetectable, suggesting that phosphorylation affects the stability of RACK1A proteins. In conclusion, these results suggest that RACK1 is phosphorylated by WNK8 and that phosphorylation negatively regulates RACK1 function by influencing its protein stability.

  11. Escape and rescue model

    NASA Astrophysics Data System (ADS)

    Alvord, D.; Nelson, H. E.

    The Escape and Rescue model is a discrete-event simulation program written in Simscript. It was developed to simulate the emergency movement involved in escape and/or rescue of people from a Board and Care Home housing a group of persons with varying degrees of physical or mental disabilities along with a small live-in staff. It may, however, be used in a much more general setting. It can reasonably handle a building with up to 100 residents and 100 rooms.

  12. SAFER Rescue System Tested

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Astronauts Carl J. Meade and Mark C. Lee (red strip on suit) test the new Simplified Aid for EVA Rescue (SAFER) system some 130 nautical miles above Earth. The pair was actually performing an in-space rehearsal or demonstration of a contingency rescue using never-before flown hardware. Meade, who here wears the small back-pack unit with its complementary chest-mounted control unit, and Lee anchored to the Space Shuttle Discovery's Remote Manipulator System (RMS) robot arm, took turns using the SAFER hardware during their shared space walk.

  13. Identification of inhibitors of checkpoint kinase 1 through template screening.

    PubMed

    Matthews, Thomas P; Klair, Suki; Burns, Samantha; Boxall, Kathy; Cherry, Michael; Fisher, Martin; Westwood, Isaac M; Walton, Michael I; McHardy, Tatiana; Cheung, Kwai-Ming J; Van Montfort, Rob; Williams, David; Aherne, G Wynne; Garrett, Michelle D; Reader, John; Collins, Ian

    2009-08-13

    Checkpoint kinase 1 (CHK1) is an oncology target of significant current interest. Inhibition of CHK1 abrogates DNA damage-induced cell cycle checkpoints and sensitizes p53 deficient cancer cells to genotoxic therapies. Using template screening, a fragment-based approach to small molecule hit generation, we have identified multiple CHK1 inhibitor scaffolds suitable for further optimization. The sequential combination of in silico low molecular weight template selection, a high concentration biochemical assay and hit validation through protein-ligand X-ray crystallography provided 13 template hits from an initial in silico screening library of ca. 15000 compounds. The use of appropriate counter-screening to rule out nonspecific aggregation by test compounds was essential for optimum performance of the high concentration bioassay. One low molecular weight, weakly active purine template hit was progressed by iterative structure-based design to give submicromolar pyrazolopyridines with good ligand efficiency and appropriate CHK1-mediated cellular activity in HT29 colon cancer cells. PMID:19572549

  14. Serum and Glucocorticoid Regulated Kinase 1 in Sodium Homeostasis

    PubMed Central

    Lou, Yiyun; Zhang, Fan; Luo, Yuqin; Wang, Liya; Huang, Shisi; Jin, Fan

    2016-01-01

    The ubiquitously expressed serum and glucocorticoid regulated kinase 1 (SGK1) is tightly regulated by osmotic and hormonal signals, including glucocorticoids and mineralocorticoids. Recently, SGK1 has been implicated as a signal hub for the regulation of sodium transport. SGK1 modulates the activities of multiple ion channels and carriers, such as epithelial sodium channel (ENaC), voltage-gated sodium channel (Nav1.5), sodium hydrogen exchangers 1 and 3 (NHE1 and NHE3), sodium-chloride symporter (NCC), and sodium-potassium-chloride cotransporter 2 (NKCC2); as well as the sodium-potassium adenosine triphosphatase (Na+/K+-ATPase) and type A natriuretic peptide receptor (NPR-A). Accordingly, SGK1 is implicated in the physiology and pathophysiology of Na+ homeostasis. Here, we focus particularly on recent findings of SGK1’s involvement in Na+ transport in renal sodium reabsorption, hormone-stimulated salt appetite and fluid balance and discuss the abnormal SGK1-mediated Na+ reabsorption in hypertension, heart disease, edema with diabetes, and embryo implantation failure. PMID:27517916

  15. Casein Kinase 1 Promotes Synchrony of the Circadian Clock Network

    PubMed Central

    Zheng, Xiangzhong; Sowcik, Mallory; Chen, Dechun

    2014-01-01

    Casein kinase 1, known as DOUBLETIME (DBT) in Drosophila melanogaster, is a critical component of the circadian clock that phosphorylates and promotes degradation of the PERIOD (PER) protein. However, other functions of DBT in circadian regulation are not clear, in part because severe reduction of dbt causes preadult lethality. Here we report the molecular and behavioral phenotype of a viable dbtEY02910 loss-of-function mutant. We found that DBT protein levels are dramatically reduced in adult dbtEY02910 flies, and the majority of mutant flies display arrhythmic behavior, with a few showing weak, long-period (∼32 h) rhythms. Peak phosphorylation of PER is delayed, and both hyper- and hypophosphorylated forms of the PER and CLOCK proteins are present throughout the day. In addition, molecular oscillations of the circadian clock are dampened. In the central brain, PER and TIM expression is heterogeneous and decoupled in the canonical clock neurons of the dbtEY02910 mutants. We also report an interaction between dbt and the signaling pathway involving pigment dispersing factor (PDF), a synchronizing peptide in the clock network. These data thus demonstrate that overall reduction of DBT causes long and arrhythmic behavior, and they reveal an unexpected role of DBT in promoting synchrony of the circadian clock network. PMID:24820422

  16. The proliferation marker thymidine kinase 1 in clinical use

    PubMed Central

    ZHOU, JI; HE, ELLEN; SKOG, SVEN

    2013-01-01

    Tumor-related biomarkers are used for the diagnosis, prognosis and monitoring of treatments and follow-up of cancer patients, although only a few are fully accepted for the detection of invisible/visible tumors in health screening. Thymidine kinase 1 (TK1), a cell cycle-dependent and thus a proliferation-related marker, has been extensively studied during the last decades, using both biochemical and immunological techniques. Therefore, TK1 is an emerging potential proliferating biomarker in oncology that may be used for the prognosis and monitoring of tumor therapy, relapse and survival. In addition, TK1 concentration in serum (STK1p) is a useful biomarker in healthy screening for the detection of potential malignancy development as well as the identification of early-stage tumors, with a few false-positive cases (ROC value, 0.96; tumor proliferation sensitivity, 0.80; specificity, 0.99). In this review, we examine results regarding the expression of STK1p and TK1 in relation to cancer patients and STK1p in health screening published between 2000 and 2012. The use of tumor-related markers recommended by international cancer organizations is also discussed. This review provides valuable information for applications in tumor patients, in health screening and for cancer research. PMID:24649117

  17. Serum and Glucocorticoid Regulated Kinase 1 in Sodium Homeostasis.

    PubMed

    Lou, Yiyun; Zhang, Fan; Luo, Yuqin; Wang, Liya; Huang, Shisi; Jin, Fan

    2016-01-01

    The ubiquitously expressed serum and glucocorticoid regulated kinase 1 (SGK1) is tightly regulated by osmotic and hormonal signals, including glucocorticoids and mineralocorticoids. Recently, SGK1 has been implicated as a signal hub for the regulation of sodium transport. SGK1 modulates the activities of multiple ion channels and carriers, such as epithelial sodium channel (ENaC), voltage-gated sodium channel (Nav1.5), sodium hydrogen exchangers 1 and 3 (NHE1 and NHE3), sodium-chloride symporter (NCC), and sodium-potassium-chloride cotransporter 2 (NKCC2); as well as the sodium-potassium adenosine triphosphatase (Na⁺/K⁺-ATPase) and type A natriuretic peptide receptor (NPR-A). Accordingly, SGK1 is implicated in the physiology and pathophysiology of Na⁺ homeostasis. Here, we focus particularly on recent findings of SGK1's involvement in Na⁺ transport in renal sodium reabsorption, hormone-stimulated salt appetite and fluid balance and discuss the abnormal SGK1-mediated Na⁺ reabsorption in hypertension, heart disease, edema with diabetes, and embryo implantation failure. PMID:27517916

  18. Casein kinase 1 promotes synchrony of the circadian clock network.

    PubMed

    Zheng, Xiangzhong; Sowcik, Mallory; Chen, Dechun; Sehgal, Amita

    2014-07-01

    Casein kinase 1, known as DOUBLETIME (DBT) in Drosophila melanogaster, is a critical component of the circadian clock that phosphorylates and promotes degradation of the PERIOD (PER) protein. However, other functions of DBT in circadian regulation are not clear, in part because severe reduction of dbt causes preadult lethality. Here we report the molecular and behavioral phenotype of a viable dbt(EY02910) loss-of-function mutant. We found that DBT protein levels are dramatically reduced in adult dbt(EY02910) flies, and the majority of mutant flies display arrhythmic behavior, with a few showing weak, long-period (∼32 h) rhythms. Peak phosphorylation of PER is delayed, and both hyper- and hypophosphorylated forms of the PER and CLOCK proteins are present throughout the day. In addition, molecular oscillations of the circadian clock are dampened. In the central brain, PER and TIM expression is heterogeneous and decoupled in the canonical clock neurons of the dbt(EY02910) mutants. We also report an interaction between dbt and the signaling pathway involving pigment dispersing factor (PDF), a synchronizing peptide in the clock network. These data thus demonstrate that overall reduction of DBT causes long and arrhythmic behavior, and they reveal an unexpected role of DBT in promoting synchrony of the circadian clock network. PMID:24820422

  19. Casein Kinase 1 Promotes Initiation of Clathrin-Mediated Endocytosis

    PubMed Central

    Peng, Yutian; Grassart, Alexandre; Lu, Rebecca; Wong, Catherine C. L.; Yates, John; Barnes, Georjana; Drubin, David G.

    2014-01-01

    Summary In budding yeast, over 60 proteins functioning in at least 5 modules are recruited to endocytic sites with predictable order and timing. However, how sites of clathrin-mediated endocytosis are initiated and stabilized is not well understood. Here, the casein kinase 1 (CK1) Hrr25 is shown to be an endocytic protein and to be among the earliest proteins to appear at endocytic sites. Hrr25 absence or overexpression decreases or increases the rate of endocytic site initiation, respectively. Ede1, an early endocytic Eps15-like protein important for endocytic initiation, is an Hrr25 target and is required for Hrr25 recruitment to endocytic sites. Hrr25 phosphorylation of Ede1 is required for Hrr25-Ede1 interaction and promotes efficient initiation of endocytic sites. These observations indicate that Hrr25 kinase and Ede1 cooperate to initiate and stabilize endocytic sites. Analysis of the mammalian homologs CK1δ/ε suggests a conserved role for these protein kinases in endocytic site initiation and stabilization. PMID:25625208

  20. Casein kinase 1 promotes initiation of clathrin-mediated endocytosis.

    PubMed

    Peng, Yutian; Grassart, Alexandre; Lu, Rebecca; Wong, Catherine C L; Yates, John; Barnes, Georjana; Drubin, David G

    2015-01-26

    In budding yeast, over 60 proteins functioning in at least five modules are recruited to endocytic sites with predictable order and timing. However, how sites of clathrin-mediated endocytosis are initiated and stabilized is not well understood. Here, the casein kinase 1 (CK1) Hrr25 is shown to be an endocytic protein and to be among the earliest proteins to appear at endocytic sites. Hrr25 absence or overexpression decreases or increases the rate of endocytic site initiation, respectively. Ede1, an early endocytic Eps15-like protein important for endocytic initiation, is an Hrr25 target and is required for Hrr25 recruitment to endocytic sites. Hrr25 phosphorylation of Ede1 is required for Hrr25-Ede1 interaction and promotes efficient initiation of endocytic sites. These observations indicate that Hrr25 kinase and Ede1 cooperate to initiate and stabilize endocytic sites. Analysis of the mammalian homologs CK1δ/ε suggests a conserved role for these protein kinases in endocytic site initiation and stabilization. PMID:25625208

  1. Operation Rescue. Final Report.

    ERIC Educational Resources Information Center

    McKee, Neena; Crawford, Georgette

    The Operation Rescue project was designed to develop a classroom setting for the delivery of comprehensive educational services to "at risk" young adolescents. The classroom was established as part of the pre-existing Jonesboro Alternative School, and it utilized the basic academic and social program developed by this entity over 12 years of work…

  2. Close Call: Unwanted Rescue.

    ERIC Educational Resources Information Center

    Journal of Adventure Education and Outdoor Leadership, 1991

    1991-01-01

    Describes incident where group engaged in training exercise was almost "rescued" by Coast Guard, although Coast Guard had been alerted that training exercise would be taking place. On another occasion Coast Guard did not react to actual report, thinking it was training group. Group was studying grey seal breeding colonies in Pembrokeshire. (KS)

  3. Rescue Manual. Module 5.

    ERIC Educational Resources Information Center

    Ohio State Univ., Columbus. Instructional Materials Lab.

    This learner manual for rescuers covers the current techniques or practices required in the rescue service. The fifth of 10 modules contains information on hazardous materials. Key points, an introduction, and conclusion accompany substantive material in this module. In addition, the module contains a Department of Transportation guide chart on…

  4. Macrophage Migration Inhibitory Factor: A Novel Inhibitor of Apoptosis Signal-Regulating Kinase 1-p38-Xanthine Oxidoreductase-Dependent Cigarette Smoke-Induced Apoptosis.

    PubMed

    Fallica, Jonathan; Varela, Lidenys; Johnston, Laura; Kim, Bo; Serebreni, Leonid; Wang, Lan; Damarla, Mahendra; Kolb, Todd M; Hassoun, Paul M; Damico, Rachel

    2016-04-01

    Cigarette smoke (CS) exposure is the leading cause of emphysema. CS mediates pathologic emphysematous remodeling of the lung via apoptosis of lung parenchymal cells resulting in enlargement of the airspaces, loss of the capillary bed, and diminished surface area for gas exchange. Macrophage migration inhibitory factor (MIF), a pleiotropic cytokine, is reduced both in a preclinical model of CS-induced emphysema and in patients with chronic obstructive pulmonary disease, particularly those with the most severe disease and emphysematous phenotype. MIF functions to antagonize CS-induced DNA damage, p53-dependent apoptosis of pulmonary endothelial cells (EndoCs) and resultant emphysematous tissue remodeling. Using primary alveolar EndoCs and a mouse model of CS-induced lung damage, we investigated the capacity and molecular mechanism(s) by which MIF modifies oxidant injury. Here, we demonstrate that both the activity of xanthine oxidoreductase (XOR), a superoxide-generating enzyme obligatory for CS-induced DNA damage and EndoC apoptosis, and superoxide concentrations are increased after CS exposure in the absence of MIF. Both XOR hyperactivation and apoptosis in the absence of MIF occurred via a p38 mitogen-activated protein kinase-dependent mechanism. Furthermore, a mitogen-activated protein kinase kinase kinase family member, apoptosis signal-regulating kinase 1 (ASK1), was necessary for CS-induced p38 activation and EndoC apoptosis. MIF was sufficient to directly suppress ASK1 enzymatic activity. Taken together, MIF suppresses CS-mediated cytotoxicity in the lung, in part by antagonizing ASK1-p38-XOR-dependent apoptosis. PMID:26390063

  5. Collectivizing rescue obligations in bioethics.

    PubMed

    Garrett, Jeremy R

    2015-01-01

    Bioethicists invoke a duty to rescue in a wide range of cases. Indeed, arguably, there exists an entire medical paradigm whereby vast numbers of medical encounters are treated as rescue cases. The intuitive power of the rescue paradigm is considerable, but much of this power stems from the problematic way that rescue cases are conceptualized-namely, as random, unanticipated, unavoidable, interpersonal events for which context is irrelevant and beneficence is the paramount value. In this article, I critique the basic assumptions of the rescue paradigm, reframe the ethical landscape in which rescue obligations are understood, and defend the necessity and value of a wider social and institutional view. Along the way, I move back and forth between ethical theory and a concrete case where the duty to rescue has been problematically applied: the purported duty to regularly return incidental findings and individual research results in genomic and genetic research. PMID:25674948

  6. Cross-talk between Arg methylation and Ser phosphorylation modulates apoptosis signal–regulating kinase 1 activation in endothelial cells

    PubMed Central

    Chen, Ming; Qu, Xiaosheng; Zhang, Zhiqing; Wu, Huayu; Qin, Xia; Li, Fuji; Liu, Zhenfang; Tian, Liyuan; Miao, Jianhua; Shu, Wei

    2016-01-01

    We describe a novel functional interaction between ASK1 and PRMT5. We show that PRMT5 interacts with and methylates ASK1 at arginine residue 89 and thereby negatively regulates its activity by promoting the interaction between ASK1 and Akt and thus phosphorylating ASK1 at serine residue 83. Furthermore, the association between ASK1 and Akt is enhanced by VEGF stimulation, and PRMT5 is required for this association. Moreover, PRMT5-mediated ASK1 methylation impaired the H2O2-induced activity of ASK1, and this inhibitory effect of PRMT5 was abolished by replacement of arginine 89 with Trp or depletion of PRMT5 expression by RNA interference. Together the results demonstrate cross-talk between arginine methylation and serine phosphorylation in ASK1. PMID:26912789

  7. Peroxiredoxin 1 has an anti-apoptotic role via apoptosis signal-regulating kinase 1 and p38 activation in mouse models with oral precancerous lesions

    PubMed Central

    ZHANG, JIANFEI; JING, XINYING; NIU, WENWEN; ZHANG, MIN; GE, LIHUA; MIAO, CONGCONG; TANG, XIAOFEI

    2016-01-01

    Peroxiredoxin 1 (Prx1) is important in the protection of cells from oxidative damage and the regulation of cell proliferation and apoptosis. Prx1 is overexpressed in oral precancerous lesions of oral leukoplakia (OLK) and oral cancer; however, the association between Prx1 expression and OLK pathogenesis remains unknown. The present study investigated the role of Prx1 and its molecular mechanisms in oxidative stress-induced apoptosis during the pathogenesis of OLK. Wild-type and Prx1 knockout mice were treated with 50 µg/ml 4-nitroquinoline-1-oxide (4NQO) or 4NQO + H2O2 for 16 weeks to establish mouse models with tongue precancerous lesions. Apoptotic cells were detected using terminal deoxynucleotidyl transferase dUTP nick-end labeling assay. The expression of Prx1, apoptosis signal-regulating kinase 1 (ASK1), phosphor-ASK1, p38 and phosphor-p38 was analyzed using immunohistochemical staining, and their mRNA expression levels were evaluated by reverse transcription quantitative polymerase chain reaction. The present results demonstrated that 4NQO or 4NQO + H2O2 induced the development of tongue precancerous lesions in Prx1 knockout and wild-type mice. Prx1 was overexpressed in tongue precancerous lesions compared with normal tongue mucosa. There was a significant decrease in the degree of moderate or severe epithelial dysplasia, and mild epithelial dysplasia was clearly elevated, in Prx1 knockout mice treated with 4NQO + H2O2 compared with wild-type mice treated with 4NQO + H2O2. Prx1 suppressed apoptosis and upregulated phosphor-ASK1 and phosphor-p38 expression in tongue precancerous lesions. The present results suggest that Prx1 suppresses oxidative stress-induced apoptosis via the ASK1/p38 signalling pathway in mouse tongue precancerous lesions. In conclusion, Prx1 and H2O2 have a coordination role in promoting the progression of tongue precancerous mucosa lesions. The present findings provide novel insight into Prx1 function and the mechanisms of Prx1 in OLK

  8. Carbamoylating Activity Associated with the Activation of the Antitumor Agent Laromustine Inhibits Angiogenesis by Inducing ASK1-Dependent Endothelial Cell Death

    PubMed Central

    Praggastis, Alexandra; Li, Yonghao; Zhou, Huanjiao Jenny; He, Yun; Ghazvinian, Roxanne; Cincotta, Dylan J.; Rice, Kevin P.; Min, Wang

    2014-01-01

    The anticancer agent 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[(methylamino)carbonyl]hydrazine (laromustine), upon decomposition in situ, yields methyl isocyanate and the chloroethylating species 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE). 90CE has been shown to kill tumor cells via a proposed mechanism that involves interstrand DNA cross-linking. However, the role of methyl isocyanate in the antineoplastic function of laromustine has not been delineated. Herein, we show that 1,2-bis(methylsulfonyl)-1-[(methylamino)carbonyl]hydrazine (101MDCE), an analog of laromustine that generates only methyl isocyanate, activates ASK1-JNK/p38 signaling in endothelial cells (EC). We have previously shown that ASK1 forms a complex with reduced thioredoxin (Trx1) in resting EC, and that the Cys residues in ASK1 and Trx1 are critical for their interaction. 101MDCE dissociated ASK1 from Trx1, but not from the phosphoserine-binding inhibitor 14-3-3, in whole cells and in cell lysates, consistent with the known ability of methyl isocyanate to carbamoylate free thiol groups of proteins. 101MDCE had no effect on the kinase activity of purified ASK1, JNK, or the catalytic activity of Trx1. However, 101MDCE, but not 90CE, significantly decreased the activity of Trx reductase-1 (TrxR1). We conclude that methyl isocyanate induces dissociation of ASK1 from Trx1 either directly by carbamoylating the critical Cys groups in the ASK1-Trx1 complex or indirectly by inhibiting TrxR1. Furthermore, 101MDCE (but not 90CE) induced EC death through a non-apoptotic (necroptotic) pathway leading to inhibition of angiogenesis in vitro. Our study has identified methyl isocyanates may contribute to the anticancer activity in part by interfering with tumor angiogenesis. PMID:25068797

  9. Hybridization facilitates evolutionary rescue

    PubMed Central

    Stelkens, Rike B; Brockhurst, Michael A; Hurst, Gregory D D; Greig, Duncan

    2014-01-01

    The resilience of populations to rapid environmental degradation is a major concern for biodiversity conservation. When environments deteriorate to lethal levels, species must evolve to adapt to the new conditions to avoid extinction. Here, we test the hypothesis that evolutionary rescue may be enabled by hybridization, because hybridization increases genetic variability. Using experimental evolution, we show that interspecific hybrid populations of Saccharomyces yeast adapt to grow in more highly degraded environments than intraspecific and parental crosses, resulting in survival rates far exceeding those of their ancestors. We conclude that hybridization can increase evolutionary responsiveness and that taxa able to exchange genes with distant relatives may better survive rapid environmental change. PMID:25558281

  10. Inflatable rescue device

    NASA Technical Reports Server (NTRS)

    Swan, Scott A. (Inventor)

    1995-01-01

    This invention discloses, in one aspect, a personal rescue device for use in outer space which has an inflatable flexible tube with a shaper apparatus herein. Gas under pressure flows through the shaper apparatus and into the flexible tube. The flexible tube is mounted to the shaper so that as it inflates it expands and deploys lengthwise away from the shaper. In one embodiment a housing contains the shaper and the flexible tube and the housing is designed to facilitate movement of the expanding tube from the housing so the expanding tube does not bunch up in the housing.

  11. [Importance of helicopter rescue].

    PubMed

    Hofer, G; Voelckel, W G

    2014-03-01

    Helicopter emergency medical service (HEMS) have become a main part of prehospital emergency medical services over the last 40 years. Recently, an ongoing discussion about financial shortage and personal shortcomings question the role of cost-intensive air rescue. Thus, the value of HEMS must be examined and discussed appropriately. Since the number of physician-staffed ground ambulances may decrease due to the limited availability of qualified physicians, HEMS may fill the gap. In addition patient transfer to specialized hospitals will require an increasing number of air transports in order to minimize prehospital time. The higher risk ratio for HEMS missions when compared with ground rescue requires a rigorous quality management system. When it comes to missions in remote and exposed areas, the scope of medical treatment must be adjusted to the individual situation. Medical competence is key in order to balance guideline compliant or maximal care versus optimal care characterized as a mission-specific, individualized emergency care concept. Although, medical decision making and treatment is typically based on the best scientific evidence, personal skills, competence, and the mission scenario will determine the scope of interventions suitable to improve outcome. Thus, the profile of requirements for the HEMS medical crew is high. PMID:24618925

  12. Curcumin induced human gastric cancer BGC-823 cells apoptosis by ROS-mediated ASK1-MKK4-JNK stress signaling pathway.

    PubMed

    Liang, Tao; Zhang, Xiaojian; Xue, Wenhua; Zhao, Songfeng; Zhang, Xiang; Pei, Jianying

    2014-01-01

    The signaling mediated by stress-activated MAP kinases (MAPK), c-Jun N-terminal kinase (JNK) has well-established importance in cancer. In the present report, we investigated the effects of curcumin on the signaling pathway in human gastric cancer BGC-823 cells. Curcumin induced reactive oxygen species (ROS) production and BGC-823 cells apoptosis. Inhibition of ROS generation by antioxidant (NAC or Trion) significantly prevented curcumin-mediated apoptosis. Notably, we observed that curcumin activated ASK1, a MAPKKK that is oxidative stress sensitive and responsible to phosphorylation of JNK via triggering cascades, up-regulated an upstream effector of the JNK, MKK4, and phosphorylated JNK protein expression in BGC-823 cells. However, curcumin induced ASK1-MKK4-JNK signaling was attenuated by NAC. All the findings confirm the possibility that oxidative stress-activated ASK1-MKK4-JNK signaling cascade promotes the apoptotic response in curcumin-treated BGC-823 cells. PMID:25198898

  13. Curcumin Induced Human Gastric Cancer BGC-823 Cells Apoptosis by ROS-Mediated ASK1-MKK4-JNK Stress Signaling Pathway

    PubMed Central

    Liang, Tao; Zhang, Xiaojian; Xue, Wenhua; Zhao, Songfeng; Zhang, Xiang; Pei, Jianying

    2014-01-01

    The signaling mediated by stress-activated MAP kinases (MAPK), c-Jun N-terminal kinase (JNK) has well-established importance in cancer. In the present report, we investigated the effects of curcumin on the signaling pathway in human gastric cancer BGC-823 cells. Curcumin induced reactive oxygen species (ROS) production and BGC-823 cells apoptosis. Inhibition of ROS generation by antioxidant (NAC or Trion) significantly prevented curcumin-mediated apoptosis. Notably, we observed that curcumin activated ASK1, a MAPKKK that is oxidative stress sensitive and responsible to phosphorylation of JNK via triggering cascades, up-regulated an upstream effector of the JNK, MKK4, and phosphorylated JNK protein expression in BGC-823 cells. However, curcumin induced ASK1-MKK4-JNK signaling was attenuated by NAC. All the findings confirm the possibility that oxidative stress-activated ASK1-MKK4-JNK signaling cascade promotes the apoptotic response in curcumin-treated BGC-823 cells. PMID:25198898

  14. Psychological Factors in Wilderness Rescue.

    ERIC Educational Resources Information Center

    Ogilvie, Bruce C.

    This presentation provides wilderness rescue workers with an overview of the psychological reactions of victims of accidents and natural disasters and suggested responses for rescuers and caregivers. A personal account of rescue and death in a drowning accident illustrates how the rescuer can also be traumatized by such an incident and may suffer…

  15. Locate and rescue system components

    NASA Technical Reports Server (NTRS)

    1976-01-01

    Two types of search and rescue systems not involving satellites are studied; one using a network radio communications link and the other a characteristic beacon signal. Line of slight limitation of VHF radio and beacon signals limit the range (approximately 25 miles) between the origin of the distress signal and the mobile rescue unit.

  16. How competition affects evolutionary rescue

    PubMed Central

    Osmond, Matthew Miles; de Mazancourt, Claire

    2013-01-01

    Populations facing novel environments can persist by adapting. In nature, the ability to adapt and persist will depend on interactions between coexisting individuals. Here we use an adaptive dynamic model to assess how the potential for evolutionary rescue is affected by intra- and interspecific competition. Intraspecific competition (negative density-dependence) lowers abundance, which decreases the supply rate of beneficial mutations, hindering evolutionary rescue. On the other hand, interspecific competition can aid evolutionary rescue when it speeds adaptation by increasing the strength of selection. Our results clarify this point and give an additional requirement: competition must increase selection pressure enough to overcome the negative effect of reduced abundance. We therefore expect evolutionary rescue to be most likely in communities which facilitate rapid niche displacement. Our model, which aligns to previous quantitative and population genetic models in the absence of competition, provides a first analysis of when competitors should help or hinder evolutionary rescue. PMID:23209167

  17. Electronic search and rescue aids

    NASA Technical Reports Server (NTRS)

    Trudell, B. J.

    1980-01-01

    There are two elements to the basic electronic search and rescue problem: a means for immediately alerting potential rescuers and an effective method to guide the rescue forces to the scene of the emergency. An Emergency Locator Transmitter (ELT) used by aircraft or an Emergency Position Indicating Radio Beacon (EPIRB) used by maritime vessels has the capability of providing for both an immediate alert and a homing signal to assist rescue forces in locating the site of the distress. This paper describes the development of ELT/EPIRB systems. Emphasis is placed on the SARSAT project, the COSPAS/SARSAT project, and an experimental 406 MHz ELT/EPIRB system.

  18. Community rescue in experimental metacommunities

    PubMed Central

    Low-Décarie, Etienne; Kolber, Marcus; Homme, Paige; Lofano, Andrea; Dumbrell, Alex; Gonzalez, Andrew; Bell, Graham

    2015-01-01

    The conditions that allow biodiversity to recover following severe environmental degradation are poorly understood. We studied community rescue, the recovery of a viable community through the evolutionary rescue of many populations within an evolving community, in metacommunities of soil microbes adapting to a herbicide. The metacommunities occupied a landscape of crossed spatial gradients of the herbicide (Dalapon) and a resource (glucose), whereas their constituent communities were either isolated or connected by dispersal. The spread of adapted communities across the landscape and the persistence of communities when that landscape was degraded were strongly promoted by dispersal, and the capacity to adapt to lethal stress was also related to community size and initial diversity. After abrupt and lethal stress, community rescue was most frequent in communities that had previously experienced sublethal levels of stress and had been connected by dispersal. Community rescue occurred through the evolutionary rescue of both initially common taxa, which remained common, and of initially rare taxa, which grew to dominate the evolved community. Community rescue may allow productivity and biodiversity to recover from severe environmental degradation. PMID:26578777

  19. Community rescue in experimental metacommunities.

    PubMed

    Low-Décarie, Etienne; Kolber, Marcus; Homme, Paige; Lofano, Andrea; Dumbrell, Alex; Gonzalez, Andrew; Bell, Graham

    2015-11-17

    The conditions that allow biodiversity to recover following severe environmental degradation are poorly understood. We studied community rescue, the recovery of a viable community through the evolutionary rescue of many populations within an evolving community, in metacommunities of soil microbes adapting to a herbicide. The metacommunities occupied a landscape of crossed spatial gradients of the herbicide (Dalapon) and a resource (glucose), whereas their constituent communities were either isolated or connected by dispersal. The spread of adapted communities across the landscape and the persistence of communities when that landscape was degraded were strongly promoted by dispersal, and the capacity to adapt to lethal stress was also related to community size and initial diversity. After abrupt and lethal stress, community rescue was most frequent in communities that had previously experienced sublethal levels of stress and had been connected by dispersal. Community rescue occurred through the evolutionary rescue of both initially common taxa, which remained common, and of initially rare taxa, which grew to dominate the evolved community. Community rescue may allow productivity and biodiversity to recover from severe environmental degradation. PMID:26578777

  20. 46 CFR 180.210 - Rescue boats.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 7 2013-10-01 2013-10-01 false Rescue boats. 180.210 Section 180.210 Shipping COAST...) LIFESAVING EQUIPMENT AND ARRANGEMENTS Number and Type of Survival Craft § 180.210 Rescue boats. (a) A vessel... that the vessel itself cannot serve as an adequate rescue craft. (c) In general, a rescue boat must...

  1. 46 CFR 180.210 - Rescue boats.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 7 2010-10-01 2010-10-01 false Rescue boats. 180.210 Section 180.210 Shipping COAST...) LIFESAVING EQUIPMENT AND ARRANGEMENTS Number and Type of Survival Craft § 180.210 Rescue boats. (a) A vessel... that the vessel itself cannot serve as an adequate rescue craft. (c) In general, a rescue boat must...

  2. 46 CFR 180.210 - Rescue boats.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 7 2014-10-01 2014-10-01 false Rescue boats. 180.210 Section 180.210 Shipping COAST...) LIFESAVING EQUIPMENT AND ARRANGEMENTS Number and Type of Survival Craft § 180.210 Rescue boats. (a) A vessel... that the vessel itself cannot serve as an adequate rescue craft. (c) In general, a rescue boat must...

  3. Targeting Translation Control with p70 S6 Kinase 1 Inhibitors to Reverse Phenotypes in Fragile X Syndrome Mice.

    PubMed

    Bhattacharya, Aditi; Mamcarz, Maggie; Mullins, Caitlin; Choudhury, Ayesha; Boyle, Robert G; Smith, Daniel G; Walker, David W; Klann, Eric

    2016-07-01

    Aberrant neuronal translation is implicated in the etiology of numerous brain disorders. Although mTORC1-p70 ribosomal S6 kinase 1 (S6K1) signaling is critical for translational control, pharmacological manipulation in vivo has targeted exclusively mTORC1 due to the paucity of specific inhibitors to S6K1. However, small molecule inhibitors of S6K1 could potentially ameliorate pathological phenotypes of diseases, which are based on aberrant translation and protein expression. One such condition is fragile X syndrome (FXS), which is considered to be caused by exaggerated neuronal translation and is the most frequent heritable cause of autism spectrum disorder (ASD). To date, potential therapeutic interventions in FXS have focused largely on targets upstream of translational control to normalize FXS-related phenotypes. Here we test the ability of two S6K1 inhibitors, PF-4708671 and FS-115, to normalize translational homeostasis and other phenotypes exhibited by FXS model mice. We found that although the pharmacokinetic profiles of the two S6K1 inhibitors differed, they overlapped in reversing multiple disease-associated phenotypes in FXS model mice including exaggerated protein synthesis, inappropriate social behavior, behavioral inflexibility, altered dendritic spine morphology, and macroorchidism. In contrast, the two inhibitors differed in their ability to rescue stereotypic marble-burying behavior and weight gain. These findings provide an initial pharmacological characterization of the impact of S6K1 inhibitors in vivo for FXS, and have therapeutic implications for other neuropsychiatric conditions involving aberrant mTORC1-S6K1 signaling. PMID:26708105

  4. Characterization of a conserved C-terminal motif (RSPRR) in ribosomal protein S6 kinase 1 required for its mammalian target of rapamycin-dependent regulation.

    PubMed

    Schalm, Stefanie S; Tee, Andrew R; Blenis, John

    2005-03-25

    The mammalian target of rapamycin, mTOR, is a Ser/Thr kinase that promotes cell growth and proliferation by activating ribosomal protein S6 kinase 1 (S6K1). We previously identified a conserved TOR signaling (TOS) motif in the N terminus of S6K1 that is required for its mTOR-dependent activation. Furthermore, our data suggested that the TOS motif suppresses an inhibitory function associated with the C terminus of S6K1. Here, we have characterized the mTOR-regulated inhibitory region within the C terminus. We have identified a conserved C-terminal "RSPRR" sequence that is responsible for an mTOR-dependent suppression of S6K1 activation. Deletion or mutations within this RSPRR motif partially rescue the kinase activity of the S6K1 TOS motif mutant (S6K1-F5A), and this rescued activity is rapamycin resistant. Furthermore, we have shown that the RSPRR motif significantly suppresses S6K1 phosphorylation at two phosphorylation sites (Thr-389 and Thr-229) that are crucial for S6K1 activation. Importantly, introducing both the Thr-389 phosphomimetic and RSPRR motif mutations into the catalytically inactive S6K1 mutant S6K1-F5A completely rescues its activity and renders it fully rapamycin resistant. These data show that the N-terminal TOS motif suppresses an inhibitory function mediated by the C-terminal RSPRR motif. We propose that the RSPRR motif interacts with a negative regulator of S6K1 that is normally suppressed by mTOR. PMID:15659381

  5. Inhibiting Polo-like kinase 1 causes growth reduction and apoptosis in pediatric acute lymphoblastic leukemia cells

    PubMed Central

    Hartsink-Segers, Stefanie A.; Exalto, Carla; Allen, Matthew; Williamson, Daniel; Clifford, Steven C.; Horstmann, Martin; Caron, Huib N.; Pieters, Rob; Den Boer, Monique L.

    2013-01-01

    This study investigated Polo-like kinase 1, a mitotic regulator often over-expressed in solid tumors and adult hematopoietic malignancies, as a potential new target in the treatment of pediatric acute lymphoblastic leukemia. Polo-like kinase 1 protein and Thr210 phosphorylation levels were higher in pediatric acute lymphoblastic leukemia (n=172) than in normal bone marrow mononuclear cells (n=10) (P<0.0001). High Polo-like kinase 1 protein phosphorylation, but not expression, was associated with a lower probability of event-free survival (P=0.042) and was a borderline significant prognostic factor (P=0.065) in a multivariate analysis including age and initial white blood cell count. Polo-like kinase 1 was necessary for leukemic cell survival, since short hairpin-mediated Polo-like kinase 1 knockdown in acute lymphoblastic leukemia cell lines inhibited cell proliferation by G2/M cell cycle arrest and induced apoptosis through caspase-3 and poly (ADP-ribose) polymerase cleavage. Primary patient cells with a high Polo-like kinase 1 protein expression were sensitive to the Polo-like kinase 1-specific inhibitor NMS-P937 in vitro, whereas cells with a low expression and normal bone marrow cells were resistant. This sensitivity was likely not caused by Polo-like kinase 1 mutations, since only one new mutation (Ser335Arg) was found by 454-sequencing of 38 pediatric acute lymphoblastic leukemia cases. This mutation did not affect Polo-like kinase 1 expression or NMS-P937 sensitivity. Together, these results indicate a pivotal role for Polo-like kinase 1 in pediatric acute lymphoblastic leukemia and show potential for Polo-like kinase 1-inhibiting drugs as an addition to current treatment strategies for cases expressing high Polo-like kinase 1 levels. PMID:23753023

  6. ASK1 (MAP3K5) as a potential therapeutic target in malignant fibrous histiocytomas with 12q14-q15 and 6q23 amplifications.

    PubMed

    Chibon, Frédéric; Mariani, Odette; Derré, Josette; Mairal, Aline; Coindre, Jean-Michel; Guillou, Louis; Sastre, Xavier; Pédeutour, Florence; Aurias, Alain

    2004-05-01

    Malignant fibrous histiocytomas (MFHs) are aggressive tumors without any definable line of differentiation. We recently demonstrated that about 20% of them are characterized by high-level amplifications of the 12q14-q15 chromosome region, associated with either 1p32 or 6q23 band amplification. This genetic finding, very similar to that in well-differentiated liposarcomas, strongly suggests that these tumors actually correspond to undifferentiated liposarcomas. It also suggests that the lack of differentiation could be the consequence of amplification of target genes localized in the 1p32 or 6q23 bands. We report here the characterization by array CGH of the 6q23 minimal region of amplification. Our findings demonstrate that amplification and overexpression of ASK1 (MAP3K5), a gene localized in the 6q23 band and encoding a mitogen-activated protein kinase kinase kinase of the JNK-MAPK signaling pathway, could inhibit the adipocytic differentiation process of the tumor cells. Treatment of a cell line with specific inhibitors of ASK1 protein resulted in the bypass of the differentiation block and induction of a strong adipocytic differentiation. These observations indicate that ASK1 is a target for new therapeutic management of these aggressive tumors. PMID:15034865

  7. Biochemical methods for quantifying sphingolipids: ceramide, sphingosine, sphingosine kinase-1 activity, and sphingosine-1-phosphate.

    PubMed

    Brizuela, Leyre; Cuvillier, Olivier

    2012-01-01

    Sphingolipids (ceramide, sphingosine, and sphingosine-1-phosphate) are bioactive lipids with important biological functions in proliferation, apoptosis, angiogenesis, and inflammation. Herein, we describe easy and rapid biochemical methods with the use of radiolabeled molecules ((3)H, (32)P) for their mass determination. Quantitation of sphingosine kinase-1 activity, the most studied isoform, is also included. PMID:22528435

  8. Cell division cycle 6, a mitotic substrate of polo-like kinase 1, regulates chromosomal segregation mediated by cyclin-dependent kinase 1 and separase

    PubMed Central

    Yim, Hyungshin; Erikson, Raymond L.

    2010-01-01

    Defining the links between cell division and DNA replication is essential for understanding normal cell cycle progression and tumorigenesis. In this report we explore the effect of phosphorylation of cell division cycle 6 (Cdc6), a DNA replication initiation factor, by polo-like kinase 1 (Plk1) on the regulation of chromosomal segregation. In mitosis, the phosphorylation of Cdc6 was highly increased, in correlation with the level of Plk1, and conversely, Cdc6 is hypophosphorylated in Plk1-depleted cells, although cyclin A- and cyclin B1-dependent kinases are active. Binding between Cdc6 and Plk1 occurs through the polo-box domain of Plk1, and Cdc6 is phosphorylated by Plk1 on T37. Immunohistochemistry studies reveal that Cdc6 and Plk1 colocalize to the central spindle in anaphase. Expression of T37V mutant of Cdc6 (Cdc6-TV) induces binucleated cells and incompletely separated nuclei. Wild-type Cdc6 but not Cdc6-TV binds cyclin-dependent kinase 1 (Cdk1). Expression of wild-type Plk1 but not kinase-defective mutant promotes the binding of Cdc6 to Cdk1. Cells expressing wild-type Cdc6 display lower Cdk1 activity and higher separase activity than cells expressing Cdc6-TV. These results suggest that Plk1-mediated phosphorylation of Cdc6 promotes the interaction of Cdc6 and Cdk1, leading to the attenuation of Cdk1 activity, release of separase, and subsequent anaphase progression. PMID:21041660

  9. Joseph Conrad's tormented Rescue (fantasy).

    PubMed

    Freedman, William

    2014-02-01

    Joseph Conrad was a notoriously tormented writer for whom the creative act was often a punishment severe enough to drive him into paralyzing depressions that delayed the completion of his novels, sometimes for years. By far the most agonizing of these projects was The Rescue, a novel he began in 1898, abandoned a year later, tried unsuccessfully to continue several times over the next two decades, but was only able to resume in 1918 and to complete, after another tortured two-year struggle, in 1920. An explanation for this incapacity, that is powerfully suggested by the novel's evocative title and perhaps unintentionally ironic subtitle (A Romance of the Shallows) has not yet been explored. Using Freud's 1910 essay on the rescue fantasy, "Contributions to the Psychology of Love: A Special Type of Choice of Object Made by Men," and Emanuel Berman's instructive revision and expansion of the concept in his 2003 American Imago essay, "Ferenczi, Rescue, and Utopia," I argue that a substantial explanation for Conrad's tormented history with The Rescue is ascribable to its quite remarkably faithful treatment of a rescue fantasy with deep and disabling resonance for its author. More specifically, the difficulty was compounded by the novel's dramatization of the soul-crushing conflict between two such fantasies: one in the service of the masculine ideal of unflinching dedication to a heroic purpose, the other promising satisfaction to the equally potent demands of emotional and sexual desire. Features of Conrad's narrative fit so tightly and consistently with the theory as Freud (and Abraham) proposed and as Berman elaborated it that The Rescue offers itself as one of those rare and reinforcing instances wherein the literary text seems to validate the psychoanalytic theory at least as persuasively as the theory "understands" the text. PMID:24555549

  10. A high-throughput kinome screen reveals serum/glucocorticoid-regulated kinase 1 as a therapeutic target for NF2-deficient meningiomas

    PubMed Central

    Beauchamp, Roberta L.; James, Marianne F.; DeSouza, Patrick A.; Wagh, Vilas; Zhao, Wen-Ning; Jordan, Justin T.; Stemmer-Rachamimov, Anat; Plotkin, Scott R.; Gusella, James F.; Haggarty, Stephen J.; Ramesh, Vijaya

    2015-01-01

    Meningiomas are the most common primary intracranial adult tumor. All Neurofibromatosis 2 (NF2)-associated meningiomas and ~60% of sporadic meningiomas show loss of NF2 tumor suppressor protein. There are no effective medical therapies for progressive and recurrent meningiomas. Our previous work demonstrated aberrant activation of mTORC1 signaling that led to ongoing clinical trials with rapamycin analogs for NF2 and sporadic meningioma patients. Here we performed a high-throughput kinome screen to identify kinases responsible for mTORC1 pathway activation in NF2-deficient meningioma cells. Among the emerging top candidates were the mTORC2-specific target serum/glucocorticoid-regulated kinase 1 (SGK1) and p21-activated kinase 1 (PAK1). In NF2-deficient meningioma cells, inhibition of SGK1 rescues mTORC1 activation, and SGK1 activation is sensitive to dual mTORC1/2 inhibitor AZD2014, but not to rapamycin. PAK1 inhibition also leads to attenuated mTORC1 but not mTORC2 signaling, suggesting that mTORC2/SGK1 and Rac1/PAK1 pathways are independently responsible for mTORC1 activation in NF2-deficient meningiomas. Using CRISPR-Cas9 genome editing, we generated isogenic human arachnoidal cell lines (ACs), the origin cell type for meningiomas, expressing or lacking NF2. NF2-null CRISPR ACs recapitulate the signaling of NF2-deficient meningioma cells. Interestingly, we observe increased SGK1 transcription and protein expression in NF2-CRISPR ACs and in primary NF2-negative meningioma lines. Moreover, we demonstrate that the dual mTORC1/mTORC2 inhibitor, AZD2014 is superior to rapamycin and PAK inhibitor FRAX597 in blocking proliferation of meningioma cells. Importantly, AZD2014 is currently in use in several clinical trials of cancer. Therefore, we believe that AZD2014 may provide therapeutic advantage over rapalogs for recurrent and progressive meningiomas. PMID:26219339

  11. A high-throughput kinome screen reveals serum/glucocorticoid-regulated kinase 1 as a therapeutic target for NF2-deficient meningiomas.

    PubMed

    Beauchamp, Roberta L; James, Marianne F; DeSouza, Patrick A; Wagh, Vilas; Zhao, Wen-Ning; Jordan, Justin T; Stemmer-Rachamimov, Anat; Plotkin, Scott R; Gusella, James F; Haggarty, Stephen J; Ramesh, Vijaya

    2015-07-10

    Meningiomas are the most common primary intracranial adult tumor. All Neurofibromatosis 2 (NF2)-associated meningiomas and ~60% of sporadic meningiomas show loss of NF2 tumor suppressor protein. There are no effective medical therapies for progressive and recurrent meningiomas. Our previous work demonstrated aberrant activation of mTORC1 signaling that led to ongoing clinical trials with rapamycin analogs for NF2 and sporadic meningioma patients. Here we performed a high-throughput kinome screen to identify kinases responsible for mTORC1 pathway activation in NF2-deficient meningioma cells. Among the emerging top candidates were the mTORC2-specific target serum/glucocorticoid-regulated kinase 1 (SGK1) and p21-activated kinase 1 (PAK1). In NF2-deficient meningioma cells, inhibition of SGK1 rescues mTORC1 activation, and SGK1 activation is sensitive to dual mTORC1/2 inhibitor AZD2014, but not to rapamycin. PAK1 inhibition also leads to attenuated mTORC1 but not mTORC2 signaling, suggesting that mTORC2/SGK1 and Rac1/PAK1 pathways are independently responsible for mTORC1 activation in NF2-deficient meningiomas. Using CRISPR-Cas9 genome editing, we generated isogenic human arachnoidal cell lines (ACs), the origin cell type for meningiomas, expressing or lacking NF2. NF2-null CRISPR ACs recapitulate the signaling of NF2-deficient meningioma cells. Interestingly, we observe increased SGK1 transcription and protein expression in NF2-CRISPR ACs and in primary NF2-negative meningioma lines. Moreover, we demonstrate that the dual mTORC1/mTORC2 inhibitor, AZD2014 is superior to rapamycin and PAK inhibitor FRAX597 in blocking proliferation of meningioma cells. Importantly, AZD2014 is currently in use in several clinical trials of cancer. Therefore, we believe that AZD2014 may provide therapeutic advantage over rapalogs for recurrent and progressive meningiomas. PMID:26219339

  12. Rotorcraft and Enabling Robotic Rescue

    NASA Technical Reports Server (NTRS)

    Young, Larry A.

    2010-01-01

    This paper examines some of the issues underlying potential robotic rescue devices (RRD) in the context where autonomous or manned rotorcraft deployment of such robotic systems is a crucial attribute for their success in supporting future disaster relief and emergency response (DRER) missions. As a part of this discussion, work related to proof-of-concept prototyping of two notional RRD systems is summarized.

  13. Activation of sphingosine kinase-1 in cancer: implications for therapeutic targeting.

    PubMed

    Cuvillier, Olivier; Ader, Isabelle; Bouquerel, Pierre; Brizuela, Leyre; Malavaud, Bernard; Mazerolles, Catherine; Rischmann, Pascal

    2010-06-01

    Sphingolipid metabolites are critical to the regulation of a number of fundamental biological processes including cancer. Whereas ceramide and sphingosine mediate and trigger apoptosis or cell growth arrest, sphingosine 1-phosphate promotes proliferation, cell survival and angiogenesis. The delicate equilibrium between the intracellular levels of each of these sphingolipids is controlled by the enzymes that either produce or degrade these metabolites. Sphingosine kinase-1 is a crucial regulator of this two-pan balance, because its produces the pro-survival and pro-angiogenic sphingosine 1-phosphate and decreases the amount of both ceramide and sphingosine, the pro-apoptotic sphingolipids. Moreover, its gene is oncogenic, its mRNA is overproduced in several solid tumors, its overexpression protects cells from apoptosis, and its activity is down-regulated by anti-cancer treatments. Therefore, the sphingosine kinase-1/sphingosine 1-phosphate signaling pathway appears to be a target of interest for therapeutic manipulation. PMID:20302564

  14. Casein Kinase 1 Epsilon Expression Predicts Poorer Prognosis in Low T-Stage Oral Cancer Patients

    PubMed Central

    Lin, Shu-Hui; Lin, Yueh-Min; Yeh, Chung-Min; Chen, Chih-Jung; Chen, Mei-Wen; Hung, Hsiao-Fang; Yeh, Kun-Tu; Yang, Shun-Fa

    2014-01-01

    Casein kinase 1 is a group of ubiquitous serine/threonine kinases that are involved in normal cellular functions and several pathological conditions, such as DNA repair, cell cycle progression, cytokinesis, differentiation, and apoptosis. Recent studies have indicated that casein kinase 1-epsilon (CK1ɛ) and casein kinase 1-delta (CK1δ) expression has a role in human cancers. We investigated the associations between CK1ɛ and CK1δ expression and the clinical parameters of oral cancer using immunohistochemical study methods on oral squamous cell carcinoma specimens. The results of our immunohistochemical analysis showed that the loss of CK1ɛ expression was greatly associated with a poor four-year survival rate in oral cancer patients (p = 0.002). A Kaplan-Meier analysis showed that patients who had a loss of CK1ɛ expression had a considerably poorer overall survival rate than patients who had positive CK1ɛ expressions (p = 0.022). A univariate analysis revealed that patients who had a loss of CK1ɛ expression had considerably poorer overall survival (OS) than patients who had positive expression (p = 0.024, hazard ratio (HR) = 1.7). In conclusion, our data indicated that the loss of cytoplasmic CK1ɛ expression is greatly associated with poor survival and might be an adverse survival factor. PMID:24557581

  15. Rescues conducted by surfers on Australian beaches.

    PubMed

    Attard, Anna; Brander, Robert W; Shaw, Wendy S

    2015-09-01

    This study describes the demographics, occurrence, location, primary hazards and outcomes involved in rescues performed by surfers on Australian beaches. Conservative estimates suggest that the number of rescues conducted by Australian surfers each year is on par with the number conducted by volunteer surf lifesavers. Surfers perform a considerable number of serious rescues in both lifesaver/lifeguard patrolled (45%) and unpatrolled (53%) beach locations. Rip currents represent the major physical hazard leading to rescue (75%) and the dominant emotional response of people rescued is one of panic (85%). Most surfer rescue events occur during conditions of moderate waves and sunny, fine weather with the highest proportion of rescues occurring on quiet beaches with few people around (26%). Swimming is the activity associated with most rescue events (63%), followed by board riding (25%). Males aged 18-29 represent the largest demographic of people rescued. Surfers with prior water-safety training are more likely to perform a higher number of rescues, however ability to perform rescues is not associated with formal training, but rather number of years' experience surfing. Seventy-eight percent of surfers were happy to help, while 28% expressed feelings of annoyance or inconvenience, generally towards unwary swimmers. Results of this research suggest that 63% of surfers feel they have saved a life. This value may be enhanced through improved training of surfers in basic water safety rescue techniques. PMID:26056968

  16. Overcoming MDR-associated chemoresistance in HL-60 acute myeloid leukemia cells by targeting sphingosine kinase-1.

    PubMed

    Bonhoure, E; Pchejetski, D; Aouali, N; Morjani, H; Levade, T; Kohama, T; Cuvillier, O

    2006-01-01

    We examined the involvement of sphingosine kinase-1, a critical regulator of the sphingolipid balance, in susceptibility to antineoplastic agents of either sensitive or multidrug-resistant acute myeloid leukemia cells. Contrary to parental HL-60 cells, doxorubicin and etoposide failed to trigger apoptosis in chemoresistant HL-60/Doxo and HL-60NP16 cells overexpressing MRP1 and MDR1, respectively. Chemosensitive HL-60 cells displayed sphingosine kinase-1 inhibition coupled with ceramide generation. In contrast, chemoresistant HL-60/ Doxo and HL-60/VP16 had sustained sphingosine kinase-1 activity and did not produce ceramide during treatment. Enforced expression of sphingosine kinase-1 in chemosensitive HL-60 cells resulted in marked inhibition of apoptosis that was mediated by blockade of mitochondrial cytochrome c efflux hence suggesting a control of apoptosis at the pre-mitochondrial level. Incubation with cell-permeable ceramide of chemoresistant cells led to a sphingosine kinase-1 inhibition and apoptosis both prevented by sphingosine kinase-1 over-expression. Furthermore, F-12509a, a new sphingosine kinase inhibitor, led to ceramide accumulation, decrease in sphingosine 1-phosphate content and caused apoptosis equally in chemosensitive and chemoresistant cell lines that is inhibited by adding sphingosine 1-phosphate or overexpressing sphingosine kinase-1. F-12509a induced classical apoptosis hallmarks namely nuclear fragmentation, caspase-3 cleavage as well as downregulation of antiapoptotic XIAP, and release of cytochrome c and SMAC/Diablo. PMID:16281067

  17. Rescue fantasies and the secret benefactor.

    PubMed

    Gillman, R D

    1992-01-01

    The concept of rescue fantasies is traced from Freud's earliest idea of the rescue of the mother as the fallen woman to later ideas of ambivalent rescue of the father, siblings, and children. Clinical vignettes from work with children and adults illustrate these points as well as reparative rescue fantasies in response to trauma and narcissistic hurt. The contemporary family romance myth of the secret benefactor as rescuer is described. An analytic case presentation explores the narcissistic-masochistic and the positive and negative oedipal meanings of the secret benefactor rescue fantasy. Application to countertransference enactments in the analyst is suggested. PMID:1289936

  18. Toward a behavioral ecology of rescue behavior.

    PubMed

    Hollis, Karen L; Nowbahari, Elise

    2013-01-01

    Although the study of helping behavior has revolutionized the field of behavioral ecology, scientific examination of rescue behavior remains extremely rare, except perhaps in ants, having been described as early as 1874. Nonetheless, recent work in our laboratories has revealed several new patterns of rescue behavior that appear to be much more complex than previously studied forms. This precisely-directed rescue behavior bears a remarkable resemblance to what has been labeled empathy in rats, and thus raises numerous philosophical and theoretical questions: How should rescue behavior (or empathy) be defined? What distinguishes rescue from other forms of altruism? In what ways is rescue behavior in ants different from, and similar to, rescue in other non-human animals? What selection pressures dictate its appearance? In this paper, we review our own experimental studies of rescue in both laboratory and field, which, taken together, begin to reveal some of the behavioral ecological conditions that likely have given rise to rescue behavior in ants. Against this background, we also address important theoretical questions involving rescue, including those outlined above. In this way, we hope not only to encourage further experimental analysis of rescue behavior, but also to highlight important similarities and differences in very distant taxa. PMID:23864298

  19. Hermes rescue strategies during launch

    NASA Astrophysics Data System (ADS)

    Cledassou, Rodelphe

    Safety and rescue strategies during the launch of Hermes space plane by Ariane 5 are discussed. Before solid booster separation, the pilots must be ejected by seats which are later recovered. After solid booster separation it becomes possible to extract the plane, which can perform a reentry leading to an available landing site or to sea recovery. When there is no useful landing site, the plane can be injected on a downgraded orbit.

  20. Mitotic regulation of SEPT9 protein by cyclin-dependent kinase 1 (Cdk1) and Pin1 protein is important for the completion of cytokinesis.

    PubMed

    Estey, Mathew P; Di Ciano-Oliveira, Caterina; Froese, Carol D; Fung, Karen Y Y; Steels, Jonathan D; Litchfield, David W; Trimble, William S

    2013-10-18

    Precise cell division is essential for multicellular development, and defects in this process have been linked to cancer. Septins are a family of proteins that are required for mammalian cell division, but their function and mode of regulation during this process are poorly understood. Here, we demonstrate that cyclin-dependent kinase 1 (Cdk1) phosphorylates septin 9 (SEPT9) upon mitotic entry, and this phosphorylation controls association with the proline isomerase, Pin1. Both SEPT9 and Pin1 are critical for mediating the final separation of daughter cells. Expression of mutant SEPT9 that is defective in Pin1 binding was unable to rescue cytokinesis defects caused by SEPT9 depletion but rather induced dominant-negative defects in cytokinesis. However, unlike SEPT9 depletion, Pin1 was not required for the accumulation of the exocyst complex at the midbody. These results suggest that SEPT9 plays multiple roles in abscission, one of which is regulated by the action of Cdk1 and Pin1. PMID:23990466

  1. Cyclin-dependent Kinase 5 (Cdk5)-dependent Phosphorylation of p70 Ribosomal S6 Kinase 1 (S6K) Is Required for Dendritic Spine Morphogenesis.

    PubMed

    Lai, Kwok-On; Liang, Zhuoyi; Fei, Erkang; Huang, Huiqian; Ip, Nancy Y

    2015-06-01

    The maturation and maintenance of dendritic spines depends on neuronal activity and protein synthesis. One potential mechanism involves mammalian target of rapamycin, which promotes protein synthesis through phosphorylation of eIF4E-binding protein and p70 ribosomal S6 kinase 1 (S6K). Upon extracellular stimulation, mammalian target of rapamycin phosphorylates S6K at Thr-389. S6K also undergoes phosphorylation at other sites, including four serine residues in the autoinhibitory domain. Despite extensive biochemical studies, the importance of phosphorylation in the autoinhibitory domain in S6K function remains unresolved, and its role has not been explored in the cellular context. Here we demonstrated that S6K in neuron was phosphorylated at Ser-411 within the autoinhibitory domain by cyclin-dependent kinase 5. Ser-411 phosphorylation was regulated by neuronal activity and brain-derived neurotrophic factor (BDNF). Knockdown of S6K in hippocampal neurons by RNAi led to loss of dendritic spines, an effect that mimics neuronal activity blockade by tetrodotoxin. Notably, coexpression of wild type S6K, but not the phospho-deficient S411A mutant, could rescue the spine defects. These findings reveal the importance of cyclin-dependent kinase 5-mediated phosphorylation of S6K at Ser-411 in spine morphogenesis driven by BDNF and neuronal activity. PMID:25903132

  2. Comparative theoretical study of the binding of potential cancer-treatment drugs to Checkpoint kinase 1

    NASA Astrophysics Data System (ADS)

    Araújo, Pedro M. M.; Pinto da Silva, Luís; Esteves da Silva, Joaquim C. G.

    2014-01-01

    This Letter focuses the binding between Checkpoint kinase 1 and two molecules with known inhibition potential, C39 and C40. In order to find the most relevant residues the structures were submitted to an optimization process. As expected C39 presented the highest inhibitory power towards Chk1, being this inhibition mode highly dependent on the interactions with Lys38 and Glu91. Glu55 and Asp148 exhibit unfavorable interactions to C39. Glu91 was the most important residues in the binding of C40 to Chk1, while interaction with Lys38, Glu55 and Gly90 resulted in repulsion.

  3. Death Associated Protein Kinase 1 (DAPK1): A Regulator of Apoptosis and Autophagy.

    PubMed

    Singh, Pratibha; Ravanan, Palaniyandi; Talwar, Priti

    2016-01-01

    Death-Associated Protein Kinase 1 (DAPK1) belongs to a family of five serine/threonine (Ser/Thr) kinases that possess tumor suppressive function and also mediate a wide range of cellular processes, including apoptosis and autophagy. The loss and gain-of-function of DAPK1 is associated with various cancer and neurodegenerative diseases respectively. In recent years, mechanistic studies have broadened our knowledge of the molecular mechanisms involved in DAPK1-mediated autophagy/apoptosis. In the present review, we have discussed the structural information and various cellular functions of DAPK1 in a comprehensive manner. PMID:27445685

  4. Structural Basis for the Potent and Selective Inhibition of Casein Kinase 1 Epsilon

    SciTech Connect

    Long, Alexander M.; Zhao, Huilin; Huang, Xin

    2012-10-29

    Casein kinase 1 epsilon (CK1ε) and its closest homologue CK1δ are key regulators of diverse cellular processes. We report two crystal structures of PF4800567, a potent and selective inhibitor of CK1ε, bound to the kinase domains of human CK1ε and CK1δ as well as one apo CK1ε crystal structure. These structures provide a molecular basis for the strong and specific inhibitor interactions with CK1ε and suggest clues for further development of CK1δ inhibitors.

  5. Death Associated Protein Kinase 1 (DAPK1): A Regulator of Apoptosis and Autophagy

    PubMed Central

    Singh, Pratibha; Ravanan, Palaniyandi; Talwar, Priti

    2016-01-01

    Death-Associated Protein Kinase 1 (DAPK1) belongs to a family of five serine/threonine (Ser/Thr) kinases that possess tumor suppressive function and also mediate a wide range of cellular processes, including apoptosis and autophagy. The loss and gain-of–function of DAPK1 is associated with various cancer and neurodegenerative diseases respectively. In recent years, mechanistic studies have broadened our knowledge of the molecular mechanisms involved in DAPK1-mediated autophagy/apoptosis. In the present review, we have discussed the structural information and various cellular functions of DAPK1 in a comprehensive manner. PMID:27445685

  6. Quercetin ameliorates ischemia/reperfusion-induced cognitive deficits by inhibiting ASK1/JNK3/caspase-3 by enhancing the Akt signaling pathway.

    PubMed

    Pei, Bing; Yang, Miaomiao; Qi, Xiaoyan; Shen, Xin; Chen, Xing; Zhang, Fayong

    2016-09-01

    Cerebral ischemia/reperfusion (I/R) is a major cause of severe disability and death all worldwide. However, therapeutic options to minimize the detrimental effects of cerebral I/R injury are limited. Recent research has demonstrated that quercetin mediates neuroprotective effects associated with the activation of the Akt signaling pathway in the cerebral I/R brain. Therefore, the aim of this study was to further investigate the mechanisms of cognitive deficits induced by cerebral I/R injury and the effects of quercetin on these mechanisms. First, we assessed anxiety-like behavioral and cognitive impairment using the open field test and the Morris water maze test, respectively. Next, we examined the severity of apoptosis by staining hippocampal neurons by the Cresyl violet method. Third, we used western blot analysis to investigate the expression of total and phosphorylated Akt, ASK1, JNK3, c-Jun and caspase-3 after I/R injury. Our results revealed that mice subjected to bilateral common carotid occlusion exhibited severe anxiety-like behavior, learning and memory impairment, cell damage and apoptosis. These severe effects were attenuated by administration of quercetin. Further, western blot analysis revealed that quercetin increased p-Akt expression and decreased p-ASK1, p-JNK3 and cleaved caspase-3 expression after cerebral I/R injury and led to inhibition of neuronal apoptosis. Conversely, treatment with LY294002 (a selective inhibitor of Akt1) reversed the effects of quercetin. In conclusion, these findings highlight the important role of quercetin in protecting against cognitive deficits and inhibiting neuronal apoptosis via the Akt signaling pathway. We believe that quercetin might prove to be a useful therapeutic component in treating cerebral I/R diseases in the near future. PMID:27450812

  7. Calcium-Ask1-MKK7-JNK2-c-Src Signaling Cascade Mediates Disruption of Intestinal Epithelial Tight Junctions by Dextran Sulfate Sodium

    PubMed Central

    Samak, Geetha; Chaudhry, Kamaljit K.; Gangwar, Ruchika; Narayanan, Damodaran; Jaggar, Jonathan H.; Rao, RadhaKrishna

    2015-01-01

    Disruption of intestinal epithelial tight junctions is an important event in the pathogenesis of ulcerative colitis. Dextran sodium sulfate (DSS) induces colitis in mice with the symptoms similar to ulcerative colitis. However, the mechanism of DSS-induced colitis is unknown. We investigated the mechanism of DSS-induced disruption of intestinal epithelial tight junctions and barrier dysfunction in Caco-2 cell monolayers in vitro and mouse colon in vivo. DSS treatment resulted in disruption of tight junctions, adherens junctions and actin cytoskeleton leading to barrier dysfunction in Caco-2 cell monolayers. DSS induced a rapid activation of c-jun N-terminal kinase (JNK), and the inhibition or knockdown of JNK2 attenuated DSS-induced tight junction disruption and barrier dysfunction. In mice, DSS administration for 4 days caused redistribution of tight junction and adherens junction proteins from the epithelial junctions, which was blocked by JNK inhibitor. In Caco-2 cell monolayers, DSS increased intracellular Ca2+ concentration, and depletion of intracellular Ca2+ by BAPTA or thapsigargin attenuated DSS-induced JNK activation, tight junction disruption and barrier dysfunction. Knockdown of Ask1 or MKK7 blocked DSS-induced tight junction disruption and barrier dysfunction. DSS activated c-Src by a Ca2+ and JNK-dependent mechanism. Inhibition of Src kinase activity or knockdown of c-Src blocked DSS-induced tight junction disruption and barrier dysfunction. DSS increased Tyr-phosphorylation of occludin, ZO-1, E-cadherin and β-catenin. SP600125 abrogated DSS-induced Tyr-phosphorylation of junctional proteins. Recombinant JNK2 induced threonine phosphorylation and auto phosphorylation of c-Src. This study demonstrates that Ca2+-Ask1-MKK7-JNK2-cSrc signaling cascade mediates DSS-induced tight junction disruption and barrier dysfunction. PMID:25377781

  8. Lunar mission safety and rescue: Escape/rescue analysis and plan

    NASA Technical Reports Server (NTRS)

    1971-01-01

    The results are presented of the technical analysis of escape/rescue/survival situations, crew survival techniques, alternate escape/rescue approaches and vehicles, and the advantages and disadvantages of each for advanced lunar exploration. Candidate escape/rescue guidelines are proposed and elements of a rescue plan developed. The areas of discussions include the following: lunar arrival/departure operations, lunar orbiter operations, lunar surface operations, lunar surface base escape/rescue analysis, lander tug location operations, portable airlock, emergency pressure suit, and the effects of no orbiting lunar station, no lunar surface base, and no foreign lunar orbit/surface operations on the escape/rescue plan.

  9. Rescuing neurons in prion disease.

    PubMed

    Verity, Nicholas C; Mallucci, Giovanna R

    2011-01-01

    One of the major current challenges to both medicine and neuroscience is the treatment of neurodegenerative diseases, which pose an ever-increasing medical, social and economic burden in the developed world. These disorders, which include Alzheimer's, Huntington's and Parkinson's diseases, and the rarer prion diseases, are separate entities clinically but have common features, including aggregates of misfolded proteins and varying patterns of neurodegeneration. A key barrier to effective treatment is that patients present clinically with advanced, irreversible, neuronal loss. Critically, mechanisms of neurotoxicity are poorly understood. Prevention of neuronal loss, ideally by targeting underlying pathogenic mechanisms, must be the aim of therapy. The present review describes the rationale and experimental approaches that have allowed such prevention, rescuing neurons in mice with prion disease. This rescue cured animals of a rapidly fatal neurodegenerative condition, resulting in symptom-free survival for their natural lifespan. Early pathological changes were reversed; behavioural, cognitive and neurophysiological deficits were recovered; and there was no neuronal loss. This was achieved by targeting the central pathogenic process in prion disease rather than the presumed toxic species, first by proof-of-principle experiments in transgenic mice and then by treatment using RNA interference for gene knockdown. The results have been a new therapeutic target for prion disease, further insight into mechanisms of prion neurotoxicity and the discovery of a window of reversibility in neuronal damage. Furthermore, the work gives rise to new concepts for treatment strategies for other neurodegenerative disorders, and highlights the need for clinical detection of early neuronal dysfunction, so that similar early rescue can also be achieved for these disorders. PMID:21158739

  10. Structural Basis of Human p70 Ribosomal S6 Kinase-1 Regulation by Activation Loop Phosphorylation

    SciTech Connect

    Sunami, Tomoko; Byrne, Noel; Diehl, Ronald E.; Funabashi, Kaoru; Hall, Dawn L.; Ikuta, Mari; Patel, Sangita B.; Shipman, Jennifer M.; Smith, Robert F.; Takahashi, Ikuko; Zugay-Murphy, Joan; Iwasawa, Yoshikazu; Lumb, Kevin J.; Munshi, Sanjeev K.; Sharma, Sujata

    2010-03-04

    p70 ribosomal S6 kinase (p70S6K) is a downstream effector of the mTOR signaling pathway involved in cell proliferation, cell growth, cell-cycle progression, and glucose homeostasis. Multiple phosphorylation events within the catalytic, autoinhibitory, and hydrophobic motif domains contribute to the regulation of p70S6K. We report the crystal structures of the kinase domain of p70S6K1 bound to staurosporine in both the unphosphorylated state and in the 3{prime}-phosphoinositide-dependent kinase-1-phosphorylated state in which Thr-252 of the activation loop is phosphorylated. Unphosphorylated p70S6K1 exists in two crystal forms, one in which the p70S6K1 kinase domain exists as a monomer and the other as a domain-swapped dimer. The crystal structure of the partially activated kinase domain that is phosphorylated within the activation loop reveals conformational ordering of the activation loop that is consistent with a role in activation. The structures offer insights into the structural basis of the 3{prime}-phosphoinositide-dependent kinase-1-induced activation of p70S6K and provide a platform for the rational structure-guided design of specific p70S6K inhibitors.

  11. Low cytoplasmic casein kinase 1 epsilon expression predicts poor prognosis in patients with hepatocellular carcinoma.

    PubMed

    Lin, Shu-Hui; Yeh, Chung-Min; Hsieh, Ming-Ju; Lin, Yueh-Min; Chen, Mei-Wen; Chen, Chih-Jung; Lin, Cheng-Yu; Hung, Hsiao-Fang; Yeh, Kun-Tu; Yang, Shun-Fa

    2016-03-01

    Casein kinase 1 epsilon (CK1ε) is a member of the casein kinase 1 (CK1) family, which comprises highly conserved and ubiquitous serine/threonine protein kinases. Recent studies have demonstrated that CK1ε plays a role in human cancers; however, the role of CK1ε in hepatocellular carcinoma (HCC) remains unclear. The study used immunohistochemistry to examine CK1ε expression in 230 HCC specimens by tissue microarray (TMA) and assessed the effect of CK1ε knockdown on migration of human hepatoma cells in vitro. The immunohistochemical analyses showed that low CK1ε expression was significantly correlated with tumor differentiation (p = 0.008), T classification (p = 0.016), tumor vascular invasion (p = 0.002), and cancer stage (p = 0.010). The univariate and multivariate analyses showed that patients with low CK1ε expression had a considerably lower OS rate than that of the patients with high CK1ε expression (p = 0.041, hazard ratio = 1.4; p = 0.039, hazard ratio = 1.4). Moreover, CK1ε small interfering RNA (siRNA) treatment exerted an invasion-promoting effect in human hepatoma cells. In conclusion, our data indicated that low CK1ε expression is correlated with a low survival rate and CK1ε may play a role as a tumor suppressor in hepatocarcinogenesis. PMID:26482619

  12. Sphingosine kinase 1/sphingosine 1-phosphate signalling pathway as a potential therapeutic target of pulmonary hypertension

    PubMed Central

    Xing, Xi-Qian; Li, Yan-Li; Zhang, Yu-Xuan; Xiao, Yi; Li, Zhi-Dong; Liu, Li-Qiong; Zhou, Yu-Shan; Zhang, Hong-Yan; Liu, Yan-Hong; Zhang, Li-Hui; Zhuang, Min; Chen, Yan-Ping; Ouyang, Sheng-Rong; Wu, Xu-Wei; Yang, Jiao

    2015-01-01

    Pulmonary hypertension is characterized by extensive vascular remodelling, leading to increased pulmonary vascular resistance and eventual death due to right heart failure. The pathogenesis of pulmonary hypertension involves vascular endothelial dysfunction and disordered vascular smooth muscle cell (VSMC) proliferation and migration, but the exact processes remain unknown. Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid involved in a wide spectrum of biological processes. S1P has been shown to regulate VSMC proliferation and migration and vascular tension via a family of five S1P G-protein-coupled receptors (S1P1-SIP5). S1P has been shown to have both a vasoconstrictive and vasodilating effect. The S1P receptors S1P1 and S1P3 promote, while S1P2 inhibits VSMC proliferation and migration in vitro in response to S1P. Moreover, it has been reported recently that sphingosine kinase 1 and S1P2 inhibitors might be useful therapeutic agents in the treatment of empirical pulmonary hypertension. The sphingosine kinase 1/S1P signalling pathways may play a role in the pathogenesis of pulmonary hypertension. Modulation of this pathway may offer novel therapeutic strategies. PMID:26550106

  13. Sphingosine kinase 1/sphingosine 1-phosphate signalling pathway as a potential therapeutic target of pulmonary hypertension.

    PubMed

    Xing, Xi-Qian; Li, Yan-Li; Zhang, Yu-Xuan; Xiao, Yi; Li, Zhi-Dong; Liu, Li-Qiong; Zhou, Yu-Shan; Zhang, Hong-Yan; Liu, Yan-Hong; Zhang, Li-Hui; Zhuang, Min; Chen, Yan-Ping; Ouyang, Sheng-Rong; Wu, Xu-Wei; Yang, Jiao

    2015-01-01

    Pulmonary hypertension is characterized by extensive vascular remodelling, leading to increased pulmonary vascular resistance and eventual death due to right heart failure. The pathogenesis of pulmonary hypertension involves vascular endothelial dysfunction and disordered vascular smooth muscle cell (VSMC) proliferation and migration, but the exact processes remain unknown. Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid involved in a wide spectrum of biological processes. S1P has been shown to regulate VSMC proliferation and migration and vascular tension via a family of five S1P G-protein-coupled receptors (S1P1-SIP5). S1P has been shown to have both a vasoconstrictive and vasodilating effect. The S1P receptors S1P1 and S1P3 promote, while S1P2 inhibits VSMC proliferation and migration in vitro in response to S1P. Moreover, it has been reported recently that sphingosine kinase 1 and S1P2 inhibitors might be useful therapeutic agents in the treatment of empirical pulmonary hypertension. The sphingosine kinase 1/S1P signalling pathways may play a role in the pathogenesis of pulmonary hypertension. Modulation of this pathway may offer novel therapeutic strategies. PMID:26550106

  14. The Data Rescue @ Home Project

    NASA Astrophysics Data System (ADS)

    Stickler, A.; Allan, R.; Valente, M. A.; Tinz, B.; Brönnimann, S.

    2012-04-01

    Climate science as a whole as well as reanalyses as a special case can significantly profit from the recovery, imaging and digitisation of historical observations. The importance of this fact is reflected in large, global data rescue projects and initiatives such as the Atmospheric Circulation Reconstructions over the Earth (ACRE, www.met-acre.org) or the EU FP7 ERA-CLIM project (www.era-clim.eu). From the time before 1957, there are still large amounts of surface data e.g. from former colonies and from overseas territories of European countries )e.g. Portugal, France and Germany) that need to be rescued. Also in case of the very early upper-air observations before the 1930s, even Europe and North America still hold an important quantity of data to be recovered in digital form. Here, we present the web platform "Data Rescue @ Home" (www.data-rescue-at-home.org), which has been developed at ETH Zurich and Oeschger Centre for Climate Change Research, and which has been designed to take advantage of the voluntary assistance of the thousands of people on the web who are interested in climate or old weather data. On the website, these volunteers can enter meteorological data shown on digital images into entry masks that resemble the original. By registering, the users get access to their personal digitisation statistics and help optimising the project. At the moment, 4 digitisation projects are online: One project is dealing with German upper-air data from the Second World War period. In a second project, station data from Tulagi (Solomon Islands) is being digitised. Finally, two collaborative projects have been included: One in cooperation with the Instituto Dom Luiz (Univ. Lisbon, Portugal), where Portuguese station data from Angra (Azores) is digitised, and a further one in cooperation with the German Meteorological Service (DWD), in which precipitation data from former German colonies is being digitised. On our poster, we will report on the status of the projects

  15. Lightweight Boom For Rescue Helicopter

    NASA Technical Reports Server (NTRS)

    Haslim, Leonard A.

    1993-01-01

    Telescoping boom and associated mechanisms attached to helicopter aid rescue operations by extending lifeline beyond sweep of main rotor. Pilot observes rescuee and control position of helicopter more effectively than if rescuee directly below and hidden from pilot's view. Rescuee outside downdraft of rotor, which is often powerful enough to blow away or submerge someone in water. Used for marine or land operations. Boom thin and lightweight because it need not support weight of rescuee. Lifeline pulls away from boom after secured around rescuee, who is lifted directly into cabin by winch. Potential application for in situ erection of telescopic space structures.

  16. Drug storage temperatures in rescue vehicles.

    PubMed

    DuBois, W C

    2000-04-01

    This study was conducted to determine storage temperatures of drugs carried on rescue vehicles. Recording thermometers were placed inside drug boxes carried on rescue vehicles. Those temperatures were compared with ambient air temperatures, temperatures inside mechanically cooled compartments of the rescue vehicles, and USP-recommended drug storage temperatures. The results indicate that drug storage temperatures in some prehospital rescue vehicles exceed USP guidelines. Mechanical cooling of the storage compartment results in drug storage temperatures within the USP guidelines. Mechanical cooling of drug storage compartments on vehicles is technologically and financially possible. PMID:10729674

  17. Iron mitigates DMT1-mediated manganese cytotoxicity via the ASK1-JNK signaling axis: Implications of iron supplementation for manganese toxicity

    PubMed Central

    Tai, Yee Kit; Chew, Katherine C. M.; Tan, Bryce W. Q.; Lim, Kah-Leong; Soong, Tuck Wah

    2016-01-01

    Manganese (Mn2+) neurotoxicity from occupational exposure is well documented to result in a Parkinson-like syndrome. Although the understanding of Mn2+ cytotoxicity is still incomplete, both Mn2+ and Fe2+ can be transported via the divalent metal transporter 1 (DMT1), suggesting that competitive uptake might disrupt Fe2+ homeostasis. Here, we found that DMT1 overexpression significantly enhanced Mn2+ cytoplasmic accumulation and JNK phosphorylation, leading to a reduction in cell viability. Although a robust activation of autophagy was observed alongside these changes, it did not trigger autophagic cell death, but was instead shown to be essential for the degradation of ferritin, which normally sequesters labile Fe2+. Inhibition of ferritin degradation through the neutralization of lysosomal pH resulted in increased ferritin and enhanced cytoplasmic Fe2+ depletion. Similarly, direct Fe2+ chelation also resulted in aggravated Mn2+-mediated JNK phosphorylation, while Fe2+ repletion protected cells, and this occurs via the ASK1-thioredoxin pathway. Taken together, our study presents the novel findings that Mn2+ cytotoxicity involves the depletion of the cytoplasmic Fe2+ pool, and the increase in autophagy-lysosome activity is important to maintain Fe2+ homeostasis. Thus, Fe2+ supplementation could have potential applications in the prevention and treatment of Mn2+-mediated toxicity. PMID:26878799

  18. Curcumin enhances the antitumor effect of ABT-737 via activation of the ROS-ASK1-JNK pathway in hepatocellular carcinoma cells

    PubMed Central

    ZHENG, RUINIAN; YOU, ZHIJIAN; JIA, JUN; LIN, SHUNHUAN; HAN, SHUAI; LIU, AIXUE; LONG, HUIDONG; WANG, SENMING

    2016-01-01

    At present, the therapeutic treatment strategies for patients with hepatocellular carcinoma (HCC) remain unsatisfactory, and novel methods are urgently required to treat this disease. Members of the B cell lymphoma (Bcl)-2 family are anti-apoptotic proteins, which are commonly expressed at high levels in certain HCC tissues and positively correlate with the treatment resistance of patients with HCC. ABT-737, an inhibitor of Bcl-2 anti-apoptotic proteins, has been demonstrated to exhibit potent antitumor effects in several types of tumor, including HCC. However, treatment with ABT-737 alone also activates certain pro-survival signaling pathways, which attenuate the antitumor validity of ABT-737. Curcumin, which is obtained from Curcuma longa, is also an antitumor potentiator in multiple types of cancer. In the present study, the synergistic effect of curcumin and ABT-737 on HCC cells was investigated for the first time, to the best of our knowledge. It was found that curcumin markedly enhanced the antitumor effects of ABT-737 on HepG2 cells, which was partially dependent on the induction of apoptosis, according to western blot analysis and flow cytometric apoptosis analysis. In addition, the sustained activation of the ROS-ASK1-c-Jun N-terminal kinase pathway may be an important mediator of the synergistic effect of curcumin and ABT-737. Collectively, these results indicated that the combination of curcumin and ABT-737 can efficaciously induce the death of HCC cells, and may offer a potential treatment strategy for patients with HCC. PMID:26707143

  19. Iron mitigates DMT1-mediated manganese cytotoxicity via the ASK1-JNK signaling axis: Implications of iron supplementation for manganese toxicity.

    PubMed

    Tai, Yee Kit; Chew, Katherine C M; Tan, Bryce W Q; Lim, Kah-Leong; Soong, Tuck Wah

    2016-01-01

    Manganese (Mn(2+)) neurotoxicity from occupational exposure is well documented to result in a Parkinson-like syndrome. Although the understanding of Mn(2+) cytotoxicity is still incomplete, both Mn(2+) and Fe(2+) can be transported via the divalent metal transporter 1 (DMT1), suggesting that competitive uptake might disrupt Fe(2+) homeostasis. Here, we found that DMT1 overexpression significantly enhanced Mn(2+) cytoplasmic accumulation and JNK phosphorylation, leading to a reduction in cell viability. Although a robust activation of autophagy was observed alongside these changes, it did not trigger autophagic cell death, but was instead shown to be essential for the degradation of ferritin, which normally sequesters labile Fe(2+). Inhibition of ferritin degradation through the neutralization of lysosomal pH resulted in increased ferritin and enhanced cytoplasmic Fe(2+) depletion. Similarly, direct Fe(2+) chelation also resulted in aggravated Mn(2+)-mediated JNK phosphorylation, while Fe(2+) repletion protected cells, and this occurs via the ASK1-thioredoxin pathway. Taken together, our study presents the novel findings that Mn(2+) cytotoxicity involves the depletion of the cytoplasmic Fe(2+) pool, and the increase in autophagy-lysosome activity is important to maintain Fe(2+) homeostasis. Thus, Fe(2+) supplementation could have potential applications in the prevention and treatment of Mn(2+)-mediated toxicity. PMID:26878799

  20. Downregulation of death-associated protein kinase 1 (DAPK1) in chronic lymphocytic leukemia.

    PubMed

    Raval, Aparna; Tanner, Stephan M; Byrd, John C; Angerman, Elizabeth B; Perko, James D; Chen, Shih-Shih; Hackanson, Björn; Grever, Michael R; Lucas, David M; Matkovic, Jennifer J; Lin, Thomas S; Kipps, Thomas J; Murray, Fiona; Weisenburger, Dennis; Sanger, Warren; Lynch, Jane; Watson, Patrice; Jansen, Mary; Yoshinaga, Yuko; Rosenquist, Richard; de Jong, Pieter J; Coggill, Penny; Beck, Stephan; Lynch, Henry; de la Chapelle, Albert; Plass, Christoph

    2007-06-01

    The heritability of B cell chronic lymphocytic leukemia (CLL) is relatively high; however, no predisposing mutation has been convincingly identified. We show that loss or reduced expression of death-associated protein kinase 1 (DAPK1) underlies cases of heritable predisposition to CLL and the majority of sporadic CLL. Epigenetic silencing of DAPK1 by promoter methylation occurs in almost all sporadic CLL cases. Furthermore, we defined a disease haplotype, which segregates with the CLL phenotype in a large family. DAPK1 expression of the CLL allele is downregulated by 75% in germline cells due to increased HOXB7 binding. In the blood cells from affected family members, promoter methylation results in additional loss of DAPK1 expression. Thus, reduced expression of DAPK1 can result from germline predisposition, as well as epigenetic or somatic events causing or contributing to the CLL phenotype. PMID:17540169

  1. NSun2 Promotes Cell Growth via Elevating Cyclin-Dependent Kinase 1 Translation

    PubMed Central

    Xing, Junyue; Yi, Jie; Cai, Xiaoyu; Tang, Hao; Liu, Zhenyun; Zhang, Xiaotian; Martindale, Jennifer L.; Yang, Xiaoling; Jiang, Bin; Gorospe, Myriam

    2015-01-01

    The tRNA methytransferase NSun2 promotes cell proliferation, but the molecular mechanism has not been elucidated. Here, we report that NSun2 regulates cyclin-dependent kinase 1 (CDK1) expression in a cell cycle-dependent manner. Knockdown of NSun2 decreased the CDK1 protein level, while overexpression of NSun2 elevated it without altering CDK1 mRNA levels. Further studies revealed that NSun2 methylated CDK1 mRNA in vitro and in cells and that methylation by NSun2 enhanced CDK1 translation. Importantly, NSun2-mediated regulation of CDK1 expression had an impact on the cell division cycle. These results provide new insight into the regulation of CDK1 during the cell division cycle. PMID:26391950

  2. Dynamic conformational ensembles regulate casein kinase-1 isoforms: Insights from molecular dynamics and molecular docking studies.

    PubMed

    Singh, Surya Pratap; Gupta, Dwijendra K

    2016-04-01

    Casein kinase-1 (CK1) isoforms actively participate in the down-regulation of canonical Wnt signaling pathway; however recent studies have shown their active roles in oncogenesis of various tissues through this pathway. Functional loss of two isoforms (CK1-α/ε) has been shown to activate the carcinogenic pathway which involves the stabilization of of cytoplasmic β-catenin. Development of anticancer therapeutics is very laborious task and depends upon the structural and conformational details of the target. This study focuses on, how the structural dynamics and conformational changes of two CK1 isoforms are synchronized in carcinogenic pathway. The conformational dynamics in kinases is the responsible for their action as has been supported by the molecular docking experiments. PMID:26788877

  3. Tyrosine phosphorylation of mitochondrial pyruvate dehydrogenase kinase 1 is important for cancer metabolism

    PubMed Central

    Hitosugi, Taro; Fan, Jun; Chung, Tae-Wook; Lythgoe, Katherine; Wang, Xu; Xie, Jianxin; Ge, Qingyuan; Gu, Ting-Lei; Polakiewicz, Roberto D.; Roesel, Johannes L.; Chen, Zhuo (Georgia); Boggon, Titus J.; Lonial, Sagar; Fu, Haian; Khuri, Fadlo R.; Kang, Sumin; Chen, Jing

    2011-01-01

    SUMMARY Many tumor cells rely on aerobic glycolysis instead of oxidative phosphorylation for their continued proliferation and survival. Myc and HIF-1 are believed to promote such a metabolic switch by, in part, upregulating gene expression of pyruvate dehydrogenase (PDH) kinase 1 (PDHK1), which phosphorylates and inactivates mitochondrial PDH and consequently pyruvate dehydrogenase complex (PDC). Here we report that tyrosine phosphorylation enhances PDHK1 kinase activity by promoting ATP and PDC binding. Functional PDC can form in mitochondria outside of matrix in some cancer cells and PDHK1 is commonly tyrosine phosphorylated in human cancers by diverse oncogenic tyrosine kinases localized to different mitochondrial compartments. Expression of phosphorylation-deficient, catalytic hypomorph PDHK1 mutants in cancer cells leads to decreased cell proliferation under hypoxia and increased oxidative phosphorylation with enhanced mitochondrial utilization of pyruvate, and reduced tumor growth in xenograft nude mice. Together, tyrosine phosphorylation activates PDHK1 to promote the Warburg effect and tumor growth. PMID:22195962

  4. [Peculiarities of phosphoglycerate kinase-1 pseudogene evolution in Schrenck salamander (Salamandrella schrenckii Strauch, 1870)].

    PubMed

    Malyarchuk, B A; Denisova, G A; Derenko, M V

    2013-07-01

    Processed copies of genes generally evolve in neutral mode as pseudogenes, however, some of them might be important sources of new functional genes. The psiPGK1 pseudogene has been discovered in Schrenck salamander (Salamandrella schrenckii, Amphibia, Caudata, Hynobiidae) via polymerase chain reaction used to amplify the phosphoglycerate kinase 1 gene (PGK1). This pseudogene is an intronless copy of PGK1 gene absent of exon 6. Analysis of psiPGK1 pseudogene polymorphism has demonstrated that it lacks mutations, which results in shifts in the stop codons and reading frames, as well as that the interspecies variation of this pseudogene was inconsistent with the neutral model of evolution. In addition, the pattern of phylogeographic differentiation of the psiPGK1 variants mainly coincides with that observed in mitochondrial DNA. These observations allow it to be suggested that the psiPGK1 pseudogene is a new functional gene in the Schrenck salamander. PMID:24450152

  5. Design and synthesis of orally bioavailable serum and glucocorticoid-regulated kinase 1 (SGK1) inhibitors

    SciTech Connect

    Hammond, Marlys; Washburn, David G.; Hoang, Tram H.; Manns, Sharada; Frazee, James S.; Nakamura, Hiroko; Patterson, Jaclyn R.; Trizna, Walter; Wu, Charlene; Azzarano, Leonard M.; Nagilla, Rakesh; Nord, Melanie; Trejo, Rebecca; Head, Martha S.; Zhao, Baoguang; Smallwood, Angela M.; Hightower, Kendra; Laping, Nicholas J.; Schnackenberg, Christine G.; Thompson, Scott K.

    2010-09-27

    The lead serum and glucocorticoid-related kinase 1 (SGK1) inhibitors 4-(5-phenyl-1H-pyrrolo[2,3-b]pyridin-3-yl)benzoic acid (1) and {l_brace}4-[5-(2-naphthalenyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]phenyl{r_brace}acetic acid (2) suffer from low DNAUC values in rat, due in part to formation and excretion of glucuronic acid conjugates. These PK/glucuronidation issues were addressed either by incorporating a substituent on the 3-phenyl ring ortho to the key carboxylate functionality of 1 or by substituting on the group in between the carboxylate and phenyl ring of 2. Three of these analogs have been identified as having good SGK1 inhibition potency and have DNAUC values suitable for in vivo testing.

  6. p21-Activated Kinase 1 Plays a Critical Role in Cellular Activation by Nef

    PubMed Central

    Fackler, Oliver T.; Lu, Xiaobin; Frost, Jeffrey A.; Geyer, Matthias; Jiang, Bing; Luo, Wen; Abo, Arie; Alberts, Arthur S.; Peterlin, B. Matija

    2000-01-01

    The activation of Nef-associated kinase (NAK) by Nef from human and simian immunodeficiency viruses is critical for efficient viral replication and pathogenesis. This induction occurs via the guanine nucleotide exchange factor Vav and the small GTPases Rac1 and Cdc42. In this study, we identified NAK as p21-activated kinase 1 (PAK1). PAK1 bound to Nef in vitro and in vivo. Moreover, the induction of cytoskeletal rearrangements such as the formation of trichopodia, the activation of Jun N-terminal kinase, and the increase of viral production were blocked by an inhibitory peptide that targets the kinase activity of PAK1 (PAK1 83-149). These results identify NAK as PAK1 and emphasize the central role its kinase activity plays in cytoskeletal rearrangements and cellular signaling by Nef. PMID:10713183

  7. 3-Phosphoinositide-Dependent protein Kinase-1 (PDK1) inhibitors: A review from patent literature

    PubMed Central

    Barile, Elisa; De, Surya K.; Pellecchia, Maurizio

    2016-01-01

    PDK1 (3-Phosphoinositide-dependent kinase 1) is a key member of the AGC protein kinase family. It plays an important role in a variety of cellular functions, leading to the activation of the PI3K signaling pathway, an event often associated with the onset and progression of several human cancers. Numerous recent observations suggest that PDK1 inhibitors may provide novel opportunities for the development of effective classes of therapeutics. On these premises, recent years have witnessed an increased effort by medicinal chemists to develop novel scaffolds to derive potent and selective PDK1 inhibitors. The intent of this review is to update the reader on the recent patent literature covering applications published between June 2008 and September 2011 that report on PDK1 inhibitors. PMID:24236780

  8. Teaching Holocaust Rescue: A Problematic Pedagogy

    ERIC Educational Resources Information Center

    Lindquist, David H.

    2008-01-01

    Determining how to teach about rescue during the Holocaust presents many dilemmas to teachers as they plan Holocaust curricula. Rescue is often overemphasized, and faulty perspectives about rescuers and their actions may cause students to develop distorted views about this aspect of Holocaust history. This article explores several factors that…

  9. Fire Service Training. Rescue Practices. (Revised).

    ERIC Educational Resources Information Center

    North Carolina State Dept. of Community Colleges, Raleigh.

    One of a set of fourteen instructional outlines for use in a course to train novice firemen, this guide covers the topic of rescue operations. Two types of rescue functions are recognized: the primary one consists of locating and saving trapped victims, and the secondary one of recovering bodies and making the area safe for other workers and…

  10. Rescue coronary stenting in acute myocardial infarction

    NASA Astrophysics Data System (ADS)

    Barbieri, Enrico; Meneghetti, Paolo; Molinari, Gionata; Zardini, Piero

    1996-01-01

    Failed rescue coronary angioplasty is a high risk situation because of high mortality. Coronary stent has given us the chance of improving and maintaining the patency of the artery. We report our preliminary experience of rescue stenting after unsuccessful coronary angioplasty.

  11. 46 CFR 169.517 - Rescue boat.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 7 2010-10-01 2010-10-01 false Rescue boat. 169.517 Section 169.517 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) NAUTICAL SCHOOLS SAILING SCHOOL VESSELS Lifesaving and Firefighting Equipment Primary Lifesaving Equipment § 169.517 Rescue boat. All vessels certificated for exposed or partially protected...

  12. Simplified Aid For Crew Rescue (SAFR)

    NASA Technical Reports Server (NTRS)

    Fisher, H. Thomas

    1990-01-01

    Viewgraphs and discussion of a Crew Emergency Rescue System (CERS) are presented. Topics covered include: functional description; operational description; interfaces with other subsystems/elements; simplified aid for crew rescue (SACR) characteristics; potential resource requirements; logistics, repair, and resupply; potential performance improvements; and automation impact.

  13. Satellite search and rescue analysis

    NASA Astrophysics Data System (ADS)

    Bailey, J. T.

    The success of rescue operations in the case of the survivors of aircraft crashes depends crucially on the rapid detection of the aircraft location. Similar considerations apply in the case of marine distress. For this reason, the U.S. is currently participating in a program called Cospas/Sarsat, an international cooperative humanitarian effort designed to assist in saving the lives of aviators and mariners in distress. The other original participants in the program include France, Canada, and the Soviet Union. The program began as an experiment with the launch of the first spacecraft, Cospas I, in June 1982. The Cospas/Sarsat partners are engaged in work concerning a second experiment, involving a new generation distress beacon operating on a frequency of 406 MHz. Details regarding the Cospas/Sarsat constellation are discussed, and attention is given to the immediate and the long-term outlook.

  14. Hubble Space Telescope Crew Rescue Analysis

    NASA Technical Reports Server (NTRS)

    Hamlin, Teri L.; Canga, Michael; Boyer, Roger; Thigpen, Eric

    2009-01-01

    In the aftermath of the 2003 Columbia accident NASA removed the Hubble Space Telescope (HST) Servicing Mission 4 (SM4) from the Space Shuttle manifest. Reasons cited included concerns that the risk of flying the mission would be too high. There was at the time no viable technique to repair the orbiter s thermal protection system if it were to be damaged by debris during ascent. Furthermore in the event of damage, since the mission was not to the International Space Station, there was no safe haven for the crew to wait for an extended period of time for a rescue. The HST servicing mission was reconsidered because of improvements in the ascent debris environment, the development of techniques for the astronauts to perform on orbit repairs to damage thermal protection, and the development of a strategy to provide a crew rescue capability. However, leading up to the launch of servicing mission, the HST crew rescue capability was a recurring topic. For HST there was a limited amount of time available to perform a crew rescue because of the limited consumables available on the Orbiter. The success of crew rescue depends upon several factors including when a problem is identified, when and to what extent power down procedures are begun, and where the rescue vehicle is in its ground processing cycle. Severe power downs maximize crew rescue success but would eliminate the option for the orbiter servicing the HST to attempt a landing. Therefore, crew rescue success needed to be weighed against preserving the ability of the orbiter to have landing option in case there was a problem with the rescue vehicle. This paper focuses on quantification of the HST mission loss of crew rescue capability using Shuttle historical data and various power down capabilities. That work supported NASA s decision to proceed with the HST service mission, which was successfully completed on May 24th 2009.

  15. Spermiogenesis initiation in Caenorhabditis elegans involves a casein kinase 1 encoded by the spe-6 gene.

    PubMed Central

    Muhlrad, Paul J; Ward, Samuel

    2002-01-01

    Immature spermatids from Caenorhabditis elegans are stimulated by an external activation signal to reorganize their membranes and cytoskeleton to form crawling spermatozoa. This rapid maturation, termed spermiogenesis, occurs without any new gene expression. To better understand this signal transduction pathway, we isolated suppressors of a mutation in the spe-27 gene, which is part of the pathway. The suppressors bypass the requirement for spe-27, as well as three other genes that act in this pathway, spe-8, spe-12, and spe-29. Eighteen of the suppressor mutations are new alleles of spe-6, a previously identified gene required for an early stage of spermatogenesis. The original spe-6 mutations are loss-of-function alleles that prevent major sperm protein (MSP) assembly in the fibrous bodies of spermatocytes and arrest development in meiosis. We have isolated the spe-6 gene and find that it encodes a predicted protein-serine/threonine kinase in the casein kinase 1 family. The suppressor mutations appear to be reduction-of-function alleles. We propose a model whereby SPE-6, in addition to its early role in spermatocyte development, inhibits spermiogenesis until the activation signal is received. The activation signal is transduced through SPE-8, SPE-12, SPE-27, and SPE-29 to relieve SPE-6 repression, thus triggering the formation of crawling spermatozoa. PMID:12019230

  16. Mitogen and stress response kinase-1 (MSK1) mediates excitotoxic induced death of hippocampal neurones.

    PubMed

    Hughes, Jane P; Staton, Penny C; Wilkinson, Marc G; Strijbos, Paul J L M; Skaper, Stephen D; Arthur, J Simon C; Reith, Alastair D

    2003-07-01

    Activation of the mitogen-activated protein kinase (MAPK/ERK) signal transduction pathway may mediate excitotoxic neuronal cell death in vitro and during ischemic brain injury in vivo. However, little is known, of the upstream regulation or downstream consequences of ERK activation under these conditions. Magnesium removal has been described to induce hyperexcitability and degeneration in cultured hippocampal neurones. Here, we show that neurotoxicity evoked by Mg2+ removal in primary hippocampal neurones stimulates ERK, but not p38, phosphorylation. Removal of Mg2+ also resulted in induction of the MAPK/ERK substrate mitogen- and stress-response kinase 1 (MSK1) and induced phosphorylation of the MSK1 substrate, the transcription factor cAMP response element binding protein (CREB). Neuronal death and phosphorylation of components in this cascade were inhibited by the Raf inhibitor SB-386023, by the MEK inhibitor U0126, or by the MSK1 inhibitors H89 and Ro318220. Importantly, this form of cell death was inhibited in hippocampal neurones cultured from MSK1-/- mice and inhibitors of Raf or MEK had no additive neuroprotective effect. Together, these data indicate that MSK1 is a physiological kinase for CREB and that this activity is an essential component of activity-dependent neuronal cell death. PMID:12807421

  17. Mumps Virus Nucleoprotein Enhances Phosphorylation of the Phosphoprotein by Polo-Like Kinase 1

    PubMed Central

    Pickar, Adrian; Zengel, James; Xu, Pei; Li, Zhuo

    2015-01-01

    ABSTRACT The viral RNA-dependent RNA polymerases (vRdRps) of nonsegmented, negative-sense viruses (NNSVs) consist of the enzymatic large protein (L) and the phosphoprotein (P). P is heavily phosphorylated, and its phosphorylation plays a critical role in viral RNA synthesis. Since NNSVs do not encode kinases, P is phosphorylated by host kinases. In this study, we investigate the roles that viral proteins play in the phosphorylation of mumps virus (MuV) P. We found that nucleoprotein (NP) enhances the phosphorylation of P. We have identified the serine/threonine kinase Polo-like kinase 1 (PLK1) as a host kinase that phosphorylates P and have found that phosphorylation of P by PLK1 is enhanced by NP. The PLK1 binding site in MuV P was mapped to residues 146 to 148 within the S(pS/T)P motif, and the phosphorylation site was identified as residues S292 and S294. IMPORTANCE It has previously been shown that P acts as a chaperone for NP, which encapsidates viral genomic RNA to form the NP-RNA complex, the functional template for viral RNA synthesis. Thus, it is assumed that phosphorylation of P may regulate NP's ability to form the NP-RNA complex, thereby regulating viral RNA synthesis. Our work demonstrates that MuV NP affects phosphorylation of P, suggesting that NP can regulate viral RNA synthesis by regulating phosphorylation of P. PMID:26608325

  18. Checkpoint kinase 1 is activated and promotes cell survival after exposure to sulphur mustard.

    PubMed

    Jowsey, Paul A; Blain, Peter G

    2015-01-22

    Sulphur mustard (SM) is a vesicating agent that has been used several times as a weapon during military conflict and continues to pose a threat as an agent of warfare/terrorism. After exposure, SM exerts both acute and delayed long-term toxic effects principally to the skin, eyes and respiratory system. These effects are thought to be mediated, at least in part, by direct interaction of SM with DNA, forming a myriad of DNA lesions and initiating effects on cell cycle and cell death pathways. Previous studies have demonstrated that a complex network of cellular DNA damage response pathways are utilised in cells exposed to SM, consistent with SM causing multiple forms of DNA damage. The present study focused on the role of Checkpoint kinase 1 (CHK1), a protein with putative roles in homologous recombination repair, p53 activation and the initiation of cell cycle checkpoints after certain forms of DNA damage. The data showed that SM caused robust activation of CHK1, monitored by multi-site phosphorylation analysis and that this activation was dependent on the ataxia telangiectasia and Rad3-related (ATR) protein kinase. Furthermore, specific inhibition of CHK1 increased SM toxicity in multiple human cell lines, with concomitant increases in markers of apoptosis, DNA damage and mitosis. Finally, the effect of CHK1 inhibition on SM toxicity was much more marked in cells with non-functional p53. PMID:25448276

  19. Checkpoint kinase 1 inhibitors as targeted molecular agents for clear cell carcinoma of the ovary

    PubMed Central

    KOBAYASHI, HIROSHI; SHIGETOMI, HIROSHI; YOSHIMOTO, CHIHARU

    2015-01-01

    In clear cell carcinoma of the ovary, chemoresistance frequently results in treatment failure. The present study aimed to review the potential association of transcription factor hepatocyte nuclear factor (HNF)-1β with cell cycle checkpoint machinery, as a mechanism for chemoresistance. The English-language literature on the subject was reviewed to identify genomic alterations and aberrant molecular pathways interacting with chemoresistance in clear cell carcinoma. Oxidative stress induced by repeated hemorrhage induces greater susceptibility of endometriotic cells to DNA damage, and subsequent malignant transformation results in endometriosis-associated ovarian cancer. Molecular changes, including those in HNF-1β and checkpoint kinase 1 (Chk1), may be a manifestation of essential alterations in cell cycle regulation, detoxification and chemoresistance in clear cell carcinoma. Chk1 is a critical signal transducer in the cell cycle checkpoint machinery. DNA damage, in turn, increases persistent phosphorylation of Chk1 and induction of G2/M phase cell cycle arrest in cells overexpressing HNF-1β. HNF-1β deletion induces apoptosis, suggesting that enhanced levels of HNF-1β may be associated with chemoresistance. Targeted therapy with Chk1 inhibitors may be explored as a potential treatment modality for patients with clear cell carcinoma. This provides a novel direction for combination therapy, including targeting of Chk1, which may overcome drug resistance and improve treatment efficacy. PMID:26622535

  20. Role of checkpoint kinase 1 (Chk1) in the mechanisms of resistance to histone deacetylase inhibitors.

    PubMed

    Lee, Ju-Hee; Choy, Megan L; Ngo, Lang; Venta-Perez, Gisela; Marks, Paul A

    2011-12-01

    Histone deacetylase inhibitors (HDACi) are a new group of anticancer drugs with tumor selective toxicity. Normal cells are relatively resistant to HDACi-induced cell death compared with cancer cells. Previously, we found that vorinostat induces DNA breaks in normal and transformed cells, which normal but not cancer cells can repair. In this study, we found that checkpoint kinase 1 (Chk1), a component of the G2 DNA damage checkpoint, is important in the resistance of normal cells to HDACi in vitro and in vivo. Inhibition of Chk1 activity with Chk1 inhibitor (UCN-01, AZD7762, or CHIR-124) in normal cells increases their sensitivity to HDACi (vorinostat, romidepsin, or entinostat) induced cell death, associated with extensive mitotic disruption. Mitotic abnormalities included loss of sister chromatid cohesion and chromosomal disruption. Inhibition of Chk1 did increase HDACi-induced cell death of transformed cells. Thus, Chk1 is an important factor in the resistance of normal cells, and some transformed cells, to HDACi-induced cell death. Use of Chk1 inhibitors in combination with anticancer agents to treat cancers may be associated with substantial toxicity. PMID:22106282

  1. Molecular interactions of polo-like kinase 1 in human cancers.

    PubMed

    Weng Ng, Wayne Tiong; Shin, Joo-Shik; Roberts, Tara Laurine; Wang, Bin; Lee, Cheok Soon

    2016-07-01

    Polo-like kinase 1 (PLK1) is an essential protein in communicating cell-cycle progression and DNA damage. Overexpression of PLK1 has been validated as a marker for poor prognosis in many cancers. PLK1 knockdown decreases the survival of cancer cells. PLK1 is therefore an attractive target for anticancer treatments. Several inhibitors have been developed, and some have been clinically tested to show additive effects with conventional therapies. Upstream regulation of PLK1 involves multiple interactions of proteins such as FoxM1, E2F and p21. Other cancer-related proteins such as pRB and p53 also indirectly influence PLK1 expression. With the high mutation rates of these genes seen in cancers, they may be associated with PLK1 deregulation. This raises the question of whether PLK1 overexpression is a cause or a consequence of oncogenesis. In addition, hypomethylation of the CpG island of the PLK1 promoter region contributes to its upregulation. PLK1 expression can be affected by many factors; thus, it is possible that PLK1 deregulation in each individual patient tumours could be due to different underlying mechanisms. PMID:26941182

  2. Doublecortin-like kinase 1 expression associates with breast cancer with neuroendocrine differentiation.

    PubMed

    Liu, Yu-Hong; Tsang, Julia Y S; Ni, Yun-Bi; Hlaing, Thazin; Chan, Siu-Ki; Chan, Kui-Fat; Ko, Chun-Wai; Mujtaba, S Shafaq; Tse, Gary M

    2016-01-12

    Doublecortin-like kinase 1 (DCLK1), a microtubule associated kinase, has recently been proposed to be a putative marker for stemness and adverse prognosis in gastrointestinal cancers. However, it is not clear whether the protein also plays similar roles in breast cancer. Here, the expression of DCLK1 was analyzed in a large cohort of invasive breast cancers (IBC) by immunohistochemistry. DCKL1 was associated with favorable clinico-pathologic features, namely lower histologic grade, absence of lymphovascular invasion, fibrotic focus, necrosis and lower pN stage (p≤0.045). Additionally, independent significant correlations were found with estrogen receptor and neuroendocrine markers (p ≤0.019), implicating its relationship with IBC with neuroendocrine differentiation (IBC-NED). In the current cohort, IBC-NED showed worse outcome than luminal cancers without NED (hazard ratio=1.756, p=0.041). Interestingly, within the IBC-NED group, DCLK1 was found to be a good prognostic factor (hazard ratio =0.288, p=0.011). These findings were in contrast to those in gastrointestinal cancers, suggesting different functional roles of DCLK1 in different types of cancers. In clinical practice, NED is not routinely assessed; thus IBC-NED are not well studied. Its poor outcome and significant heterogeneity warrants more attention. DCLK1 expression could aid in the prognostication and management of this special cancer subtype. PMID:26621833

  3. The Impact of Sphingosine Kinase-1 in Head and Neck Cancer

    PubMed Central

    Tamashiro, Paulette M.; Furuya, Hideki; Shimizu, Yoshiko; Iino, Kayoko; Kawamori, Toshihiko

    2013-01-01

    Head and neck squamous cell carcinoma (HNSCC) has a high reoccurrence rate and an extremely low survival rate. There is limited availability of effective therapies to reduce the rate of recurrence, resulting in high morbidity and mortality of advanced cases. Late presentation, delay in detection of lesions, and a high rate of metastasis make HNSCC a devastating disease. This review offers insight into the role of sphingosine kinase-1 (SphK1), a key enzyme in sphingolipid metabolism, in HNSCC. Sphingolipids not only play a structural role in cellular membranes, but also modulate cell signal transduction pathways to influence biological outcomes such as senescence, differentiation, apoptosis, migration, proliferation, and angiogenesis. SphK1 is a critical regulator of the delicate balance between proliferation and apoptosis. The highest expression of SphK1 is found in the advanced stage of disease, and there is a positive correlation between SphK1 expression and recurrent tumors. On the other hand, silencing SphK1 reduces HNSCC tumor growth and sensitizes tumors to radiation-induced death. Thus, SphK1 plays an important and influential role in determining HNSCC proliferation and metastasis. We discuss roles of SphK1 and other sphingolipids in HNSCC development and therapeutic strategies against HNSCC. PMID:24970177

  4. Structure and ubiquitination-dependent activation of TANK-binding kinase 1.

    PubMed

    Tu, Daqi; Zhu, Zehua; Zhou, Alicia Y; Yun, Cai-hong; Lee, Kyung-Eun; Toms, Angela V; Li, Yiqun; Dunn, Gavin P; Chan, Edmond; Thai, Tran; Yang, Shenghong; Ficarro, Scott B; Marto, Jarrod A; Jeon, Hyesung; Hahn, William C; Barbie, David A; Eck, Michael J

    2013-03-28

    Upon stimulation by pathogen-associated inflammatory signals, TANK-binding kinase 1 (TBK1) induces type I interferon expression and modulates nuclear factor κB (NF-κB) signaling. Here, we describe the 2.4 Å-resolution crystal structure of nearly full-length TBK1 in complex with specific inhibitors. The structure reveals a dimeric assembly created by an extensive network of interactions among the kinase, ubiquitin-like, and scaffold/dimerization domains. An intact TBK1 dimer undergoes K63-linked polyubiquitination on lysines 30 and 401, and these modifications are required for TBK1 activity. The ubiquitination sites and dimer contacts are conserved in the close homolog inhibitor of κB kinase ε (IKKε) but not in IKKβ, a canonical IKK that assembles in an unrelated manner. The multidomain architecture of TBK1 provides a structural platform for integrating ubiquitination with kinase activation and IRF3 phosphorylation. The structure of TBK1 will facilitate studies of the atypical IKKs in normal and disease physiology and further the development of more specific inhibitors that may be useful as anticancer or anti-inflammatory agents. PMID:23453972

  5. Adipocyte lipolysis-stimulated interleukin-6 production requires sphingosine kinase 1 activity.

    PubMed

    Zhang, Wenliang; Mottillo, Emilio P; Zhao, Jiawei; Gartung, Allison; VanHecke, Garrett C; Lee, Jen-Fu; Maddipati, Krishna R; Xu, Haiyan; Ahn, Young-Hoon; Proia, Richard L; Granneman, James G; Lee, Menq-Jer

    2014-11-14

    Adipocyte lipolysis can increase the production of inflammatory cytokines such as interleukin-6 (IL-6) that promote insulin resistance. However, the mechanisms that link lipolysis with inflammation remain elusive. Acute activation of β3-adrenergic receptors (ADRB3) triggers lipolysis and up-regulates production of IL-6 in adipocytes, and both of these effects are blocked by pharmacological inhibition of hormone-sensitive lipase. We report that stimulation of ADRB3 induces expression of sphingosine kinase 1 (SphK1) and increases sphingosine 1-phosphate production in adipocytes in a manner that also depends on hormone-sensitive lipase activity. Mechanistically, we found that adipose lipolysis-induced SphK1 up-regulation is mediated by the c-Jun N-terminal kinase (JNK)/activating protein-1 signaling pathway. Inhibition of SphK1 by sphingosine kinase inhibitor 2 diminished the ADRB3-induced IL-6 production both in vitro and in vivo. Induction of IL-6 by ADRB3 activation was suppressed by siRNA knockdown of Sphk1 in cultured adipocytes and was severely attenuated in Sphk1 null mice. Conversely, ectopic expression of SphK1 increased IL-6 expression in adipocytes. Collectively, these data demonstrate that SphK1 is a critical mediator in lipolysis-triggered inflammation in adipocytes. PMID:25253697

  6. Sphingosine kinase 1 inhibition sensitizes hormone-resistant prostate cancer to docetaxel.

    PubMed

    Sauer, Lysann; Nunes, Joao; Salunkhe, Vishal; Skalska, Lenka; Kohama, Takafumi; Cuvillier, Olivier; Waxman, Jonathan; Pchejetski, Dmitry

    2009-12-01

    It has recently been shown that docetaxel chemotherapy is effective in prolonging life in patients with prostate cancer (PCa). We have investigated potential ways of increasing the effectiveness of chemotherapy in this disease. We have previously reported that sphingosine kinase 1 (SphK1) inhibition is a key step in docetaxel-induced apoptosis in the PC-3 PCa cell line and that pharmacologicalSphK1 inhibition is chemosensitizing in the docetaxel-resistant PCa LNCaP cell line. In this study we have addressed the mechanism of docetaxel-induced apoptosis of PC-3 cells and identified SphK1-dependent and -independent components. We have shown that SphK1 inhibition by docetaxel is a two-step process involving an initial loss of enzyme activity followed by a decrease in SphK1 gene expression. Using hormoneresistant PC-3 and DU145 PCa cells we have demonstrated that both pharmacological and siRNA-mediated SphK1 inhibition leads to a four-fold decrease in the docetaxel IC50 dose. This work points out to potential ways of increasing the effectiveness of chemotherapy for PCa by SphK1 inhibition. PMID:19521959

  7. Chemosensitizing effects of sphingosine kinase-1 inhibition in prostate cancer cell and animal models.

    PubMed

    Pchejetski, Dimitri; Doumerc, Nicolas; Golzio, Muriel; Naymark, Maria; Teissié, Justin; Kohama, Takafumi; Waxman, Jonathan; Malavaud, Bernard; Cuvillier, Olivier

    2008-07-01

    We have previously reported that, in prostate cancer, inhibition of the oncogenic sphingosine kinase-1/sphingosine 1-phosphate (SphK1/S1P) pathway is a key element in chemotherapy-induced apoptosis. Here, we show that selective pharmacologic inhibition of SphK1 triggers apoptosis in LNCaP and PC-3 prostate cancer cells, an effect that is reversed by SphK1 enforced expression. More importantly, we show for the first time that the up-regulation of the SphK1/S1P pathway plays a crucial role in the resistance of prostate cancer cells to chemotherapy. Importantly, pharmacologic SphK1 inhibition with the B-5354c compound sensitizes LNCaP and PC-3 cells to docetaxel and camptothecin, respectively. In vivo, camptothecin and B-5354c alone display a limited effect on tumor growth in PC-3 cells, whereas in combination there is a synergy of effect on tumor size with a significant increase in the ceramide to S1P sphingolipid ratio. To conclude, our study highlights the notion that drugs specifically designed to inhibit SphK1 could provide a means of enhancing the effects of conventional treatment through the prosurvival antiapoptotic SphK1/S1P pathway. PMID:18644996

  8. Sphingosine kinase-1 as a chemotherapy sensor in prostate adenocarcinoma cell and mouse models.

    PubMed

    Pchejetski, Dimitri; Golzio, Muriel; Bonhoure, Elisabeth; Calvet, Cyril; Doumerc, Nicolas; Garcia, Virginie; Mazerolles, Catherine; Rischmann, Pascal; Teissié, Justin; Malavaud, Bernard; Cuvillier, Olivier

    2005-12-15

    Systemic chemotherapy was considered of modest efficacy in prostate cancer until the recent introduction of taxanes. We took advantage of the known differential effect of camptothecin and docetaxel on human PC-3 and LNCaP prostate cancer cells to determine their effect on sphingosine kinase-1 (SphK1) activity and subsequent ceramide/sphingosine 1-phosphate (S1P) balance in relation with cell survival. In vitro, docetaxel and camptothecin induced strong inhibition of SphK1 and elevation of the ceramide/S1P ratio only in cell lines sensitive to these drugs. SphK1 overexpression in both cell lines impaired the efficacy of chemotherapy by decreasing the ceramide/S1P ratio. Alternatively, silencing SphK1 by RNA interference or pharmacologic inhibition induced apoptosis coupled with ceramide elevation and loss of S1P. The differential effect of both chemotherapeutics was confirmed in an orthotopic PC-3/green fluorescent protein model established in nude mice. Docetaxel induced a stronger SphK1 inhibition and ceramide/S1P ratio elevation than camptothecin. This was accompanied by a smaller tumor volume and the reduced occurrence and number of metastases. SphK1-overexpressing PC-3 cells implanted in animals developed remarkably larger tumors and resistance to docetaxel treatment. These results provide the first in vivo demonstration of SphK1 as a sensor of chemotherapy. PMID:16357178

  9. FTY720 (fingolimod) sensitizes prostate cancer cells to radiotherapy by inhibition of sphingosine kinase-1.

    PubMed

    Pchejetski, Dmitri; Bohler, Torsten; Brizuela, Leyre; Sauer, Lysann; Doumerc, Nicolas; Golzio, Muriel; Salunkhe, Vishal; Teissié, Justin; Malavaud, Bernard; Waxman, Jonathan; Cuvillier, Olivier

    2010-11-01

    Radiotherapy is widely used as a radical treatment for prostate cancer, but curative treatments are elusive for poorly differentiated tumors where survival is just 15% at 15 years. Dose escalation improves local response rates but is limited by tolerance in normal tissues. A sphingosine analogue, FTY720 (fingolimod), a drug currently in phase III studies for treatment of multiple sclerosis, has been found to be a potent apoptosis inducer in prostate cancer cells. Using in vitro and in vivo approaches, we analyzed the impact of FTY720 on sphingolipid metabolism in hormone-refractory metastatic prostate cancer cells and evaluated its potential as a radiosensitizer on cell lines and prostate tumor xenografts. In prostate cancer cell lines, FTY720 acted as a sphingosine kinase 1 (SphK1) inhibitor that induced prostate cancer cell apoptosis in a manner independent of sphingosine-1-phosphate receptors. In contrast, γ irradiation did not affect SphK1 activity in prostate cancer cells yet synergized with FTY720 to inhibit SphK1. In mice bearing orthotopic or s.c. prostate cancer tumors, we show that FTY720 dramatically increased radiotherapeutic sensitivity, reducing tumor growth and metastasis without toxic side effects. Our findings suggest that low, well-tolerated doses of FTY720 could offer significant improvement to the clinical treatment of prostate cancer. PMID:20959468

  10. Multiple faces of protein interacting with C kinase 1 (PICK1): Structure, function, and diseases.

    PubMed

    Li, Yun-Hong; Zhang, Nan; Wang, Ya-Nan; Shen, Ying; Wang, Yin

    2016-09-01

    Protein interacting with C-kinase 1 (PICK1) has received considerable attention because it is the only protein that contains both PSD-95/DlgA/ZO-1 (PDZ) domain and Bin-Amphiphysin-Rvs (BAR) domain. Through PDZ and BAR domains, PICK1 binds to a large number of membrane proteins and lipid molecules, and is thereby of multiple functions. PICK1 is widely expressed in various tissues, particularly abundant in the brain and testis. In the central nervous system (CNS), PICK1 interacts with numerous neurotransmitters receptors, transporters, ion channels, and enzymes, and controls their trafficking. The best characterized function of PICK1 is that it regulates trafficking of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR) subunit GluA2 during long-term depression and long-term potentiation. Recent evidence shows that PICK1 participates in various diseases including neurobiological disorders, such as chronic pain, epilepsy, oxidative stress, stroke, Parkinson's disease, amyotrophic lateral sclerosis, schizophrenia, and non-neurological disorders, such as globozoospermia, breast cancer, and heart failure. In this review, we will summarize recent advances focusing on the structure and regulation of PICK1 and its functions in protein trafficking, neurological and non-neurological diseases. PMID:26970394

  11. Quantitative phospho-proteomics to investigate the polo-like kinase 1-dependent phospho-proteome.

    PubMed

    Grosstessner-Hain, Karin; Hegemann, Björn; Novatchkova, Maria; Rameseder, Jonathan; Joughin, Brian A; Hudecz, Otto; Roitinger, Elisabeth; Pichler, Peter; Kraut, Norbert; Yaffe, Michael B; Peters, Jan-Michael; Mechtler, Karl

    2011-11-01

    Polo-like kinase 1 (PLK1) is a key regulator of mitotic progression and cell division, and small molecule inhibitors of PLK1 are undergoing clinical trials to evaluate their utility in cancer therapy. Despite this importance, current knowledge about the identity of PLK1 substrates is limited. Here we present the results of a proteome-wide analysis of PLK1-regulated phosphorylation sites in mitotic human cells. We compared phosphorylation sites in HeLa cells that were or were not treated with the PLK1-inhibitor BI 4834, by labeling peptides via methyl esterification, fractionation of peptides by strong cation exchange chromatography, and phosphopeptide enrichment via immobilized metal affinity chromatography. Analysis by quantitative mass spectrometry identified 4070 unique mitotic phosphorylation sites on 2069 proteins. Of these, 401 proteins contained one or multiple phosphorylation sites whose abundance was decreased by PLK1 inhibition. These include proteins implicated in PLK1-regulated processes such as DNA damage, mitotic spindle formation, spindle assembly checkpoint signaling, and chromosome segregation, but also numerous proteins that were not suspected to be regulated by PLK1. Analysis of amino acid sequence motifs among phosphorylation sites down-regulated under PLK1 inhibition in this data set identified two potential novel variants of the PLK1 consensus motif. PMID:21857030

  12. Serum and glucocorticoid-regulated kinase 1 regulates neutrophil clearance during inflammation resolution.

    PubMed

    Burgon, Joseph; Robertson, Anne L; Sadiku, Pranvera; Wang, Xingang; Hooper-Greenhill, Edward; Prince, Lynne R; Walker, Paul; Hoggett, Emily E; Ward, Jonathan R; Farrow, Stuart N; Zuercher, William J; Jeffrey, Philip; Savage, Caroline O; Ingham, Philip W; Hurlstone, Adam F; Whyte, Moira K B; Renshaw, Stephen A

    2014-02-15

    The inflammatory response is integral to maintaining health by functioning to resist microbial infection and repair tissue damage. Large numbers of neutrophils are recruited to inflammatory sites to neutralize invading bacteria through phagocytosis and the release of proteases and reactive oxygen species into the extracellular environment. Removal of the original inflammatory stimulus must be accompanied by resolution of the inflammatory response, including neutrophil clearance, to prevent inadvertent tissue damage. Neutrophil apoptosis and its temporary inhibition by survival signals provides a target for anti-inflammatory therapeutics, making it essential to better understand this process. GM-CSF, a neutrophil survival factor, causes a significant increase in mRNA levels for the known anti-apoptotic protein serum and glucocorticoid-regulated kinase 1 (SGK1). We have characterized the expression patterns and regulation of SGK family members in human neutrophils and shown that inhibition of SGK activity completely abrogates the antiapoptotic effect of GM-CSF. Using a transgenic zebrafish model, we have disrupted sgk1 gene function and shown this specifically delays inflammation resolution, without altering neutrophil recruitment to inflammatory sites in vivo. These data suggest SGK1 plays a key role in regulating neutrophil survival signaling and thus may prove a valuable therapeutic target for the treatment of inflammatory disease. PMID:24431232

  13. Serum and Glucocorticoid Regulated Kinase 1 (SGK1) Regulates Neutrophil Clearance During Inflammation Resolution

    PubMed Central

    Burgon, Joseph; Robertson, Anne L.; Sadiku, Pranvera; Wang, Xingang; Hooper-Greenhill, Edward; Prince, Lynne R.; Walker, Paul; Hoggett, Emily E.; Ward, Jonathan R.; Farrow, Stuart N.; Zuercher, William J.; Jeffrey, Philip; Savage, Caroline O.; Ingham, Philip W.; Hurlstone, Adam F.; Whyte, Moira K. B.; Renshaw, Stephen A.

    2013-01-01

    The inflammatory response is integral to maintaining health, by functioning to resist microbial infection and repair tissue damage. Large numbers of neutrophils are recruited to inflammatory sites to neutralise invading bacteria through phagocytosis and the release of proteases and reactive oxygen species into the extracellular environment. Removal of the original inflammatory stimulus must be accompanied by resolution of the inflammatory response, including neutrophil clearance, to prevent inadvertent tissue damage. Neutrophil apoptosis and its temporary inhibition by survival signals provides a target for anti-inflammatory therapeutics, making it essential to better understand this process. GM-CSF, a neutrophil survival factor, causes a significant increase in mRNA levels for the known anti-apoptotic protein Serum and Glucocorticoid Regulated Kinase 1 (SGK1). We have characterised the expression patterns and regulation of SGK family members in human neutrophils, and shown that inhibition of SGK activity completely abrogates the anti-apoptotic effect of GM-CSF. Using a transgenic zebrafish model, we have disrupted sgk1 gene function and shown this specifically delays inflammation resolution, without altering neutrophil recruitment to inflammatory sites in vivo. These data suggest SGK1 plays a key role in regulating neutrophil survival signalling, and thus may prove a valuable therapeutic target for the treatment of inflammatory disease. PMID:24431232

  14. Arabidopsis SOMATIC EMBRYOGENESIS RECEPTOR KINASES1 and 2 are essential for tapetum development and microspore maturation.

    PubMed

    Colcombet, Jean; Boisson-Dernier, Aurélien; Ros-Palau, Roc; Vera, Carlos E; Schroeder, Julian I

    2005-12-01

    Among the >200 members of the leucine-rich repeat receptor kinase family in Arabidopsis thaliana, only a few have been functionally characterized. Here, we report a critical function in anther development for the SOMATIC EMBRYOGENESIS RECEPTOR KINASE1 (SERK1) and SERK2 genes. Both SERK1 and SERK2 are expressed widely in locules until stage 6 anthers and are more concentrated in the tapetal cell layer later. Whereas serk1 and serk2 single insertion mutants did not show developmental phenotypes, serk1 serk2 double mutants were not able to produce seeds because of a lack of pollen development in mutant anthers. In young buds, double mutant anthers developed normally, but serk1 serk2 microsporangia produced more sporogenous cells that were unable to develop beyond meiosis. Furthermore, serk1 serk2 double mutants developed only three cell layers surrounding the sporogenous cell mass, whereas wild-type anthers developed four cell layers. Further confocal microscopic and molecular analyses showed that serk1 serk2 double mutant anthers lack development of the tapetal cell layer, which accounts for the microspore abortion and male sterility. Taken together, these findings demonstrate that the SERK1 and SERK2 receptor kinases function redundantly as an important control point for sporophytic development controlling male gametophyte production. PMID:16284306

  15. Calcium-independent activation of extracellular signal-regulated kinases 1 and 2 by cyclic strain

    NASA Technical Reports Server (NTRS)

    Ikeda, M.; Takei, T.; Mills, I.; Sumpio, B. E.

    1998-01-01

    We have previously demonstrated that cyclic strain induces extracellular signal-regulated kinases 1 and 2 (ERK1/2) activation in endothelial cells (EC). The aim of this study was to investigate the effect of Ca2+ on the activation of ERK1/2. Bovine aortic EC were pretreated with a chelator of extracellular Ca2+, ethylaneglycol-bis(aminoethylether)-tetra-acetate (EGTA), a depleter of Ca2+ pools, 2,5-Di-(tert-butyl)-1,4-benzohydroquinone (BHQ), or a Ca2+ channel blocker, GdCl3, and subjected to an average 10 % strain at a rate of 60 cycles/min for 10 min. BHQ and GdCl3 did not inhibit the strain-induced ERK1/2 activation. Chelation of normal extracellular Ca2+ (1.8 mM) medium with EGTA (3 mM) acutely stimulated baseline phosphorylation and activation of ERK1/2, thereby obscuring any strain-induced activation of ERK1/2. However, in EC preincubated for 24 hours in Ca2+-free medium, elevated baseline phosphorylation was minimally activated by EGTA (200 microM) such that cyclic strain stimulated ERK1/2 in the presence or absence of BHQ. These results suggest a Ca2+ independence of the ERK1/2 signaling pathway by cyclic strain. Copyright 1998 Academic Press.

  16. Inositol hexakisphosphate kinase-1 interacts with perilipin1 to modulate lipolysis.

    PubMed

    Ghoshal, Sarbani; Tyagi, Richa; Zhu, Qingzhang; Chakraborty, Anutosh

    2016-09-01

    Lipolysis leads to the breakdown of stored triglycerides (TAG) to release free fatty acids (FFA) and glycerol which is utilized by energy expenditure pathways to generate energy. Therefore, a decrease in lipolysis augments fat accumulation in adipocytes which promotes weight gain. Conversely, if lipolysis is not complemented by energy expenditure, it leads to FFA induced insulin resistance and type-2 diabetes. Thus, lipolysis is under stringent physiological regulation, although the precise mechanism of the regulation is not known. Deletion of inositol hexakisphosphate kinase-1 (IP6K1), the major inositol pyrophosphate biosynthetic enzyme, protects mice from high fat diet (HFD) induced obesity and insulin resistance. IP6K1-KO mice are lean due to enhanced energy expenditure. Therefore, IP6K1 is a target in obesity and type-2 diabetes. However, the mechanism/s by which IP6K1 regulates adipose tissue lipid metabolism is yet to be understood. Here, we demonstrate that IP6K1-KO mice display enhanced basal lipolysis. IP6K1 modulates lipolysis via its interaction with the lipolytic regulator protein perilipin1 (PLIN1). Furthermore, phosphorylation of IP6K1 at a PKC/PKA motif modulates its interaction with PLIN1 and lipolysis. Thus, IP6K1 is a novel regulator of PLIN1 mediated lipolysis. PMID:27373682

  17. Polo-like kinase 1 licenses CENP-A deposition at centromeres.

    PubMed

    McKinley, Kara L; Cheeseman, Iain M

    2014-07-17

    To ensure the stable transmission of the genome during vertebrate cell division, the mitotic spindle must attach to a single locus on each chromosome, termed the centromere. The fundamental requirement for faithful centromere inheritance is the controlled deposition of the centromere-specifying histone, CENP-A. However, the regulatory mechanisms that ensure the precise control of CENP-A deposition have proven elusive. Here, we identify polo-like kinase 1 (Plk1) as a centromere-localized regulator required to initiate CENP-A deposition in human cells. We demonstrate that faithful CENP-A deposition requires integrated signals from Plk1 and cyclin-dependent kinase (CDK), with Plk1 promoting the localization of the key CENP-A deposition factor, the Mis18 complex, and CDK inhibiting Mis18 complex assembly. By bypassing these regulated steps, we uncoupled CENP-A deposition from cell-cycle progression, resulting in mitotic defects. Thus, CENP-A deposition is controlled by a two-step regulatory paradigm comprised of Plk1 and CDK that is crucial for genomic integrity. PMID:25036634

  18. Polo-like kinase 1 licenses CENP-A deposition at centromeres

    PubMed Central

    McKinley, Kara L.; Cheeseman, Iain M.

    2014-01-01

    Summary: To ensure the stable transmission of the genome during vertebrate cell division, the mitotic spindle must attach to a single locus on each chromosome, termed the centromere. The fundamental requirement for faithful centromere inheritance is the controlled deposition of the centromere-specifying histone, CENP-A. However, the regulatory mechanisms that ensure the precise control of CENP-A deposition have proved elusive. Here, we identify Polo-like kinase 1 (Plk1) as a centromere-localized regulator required to initiate CENP-A deposition in human cells. We demonstrate that faithful CENP-A deposition requires integrated signals from Plk1 and cyclin-dependent kinase (CDK), with Plk1 promoting the localization of the key CENP-A assembly factor, the Mis18 complex, and CDK inhibiting Mis18 complex assembly. By bypassing these regulated steps, we uncoupled CENP-A deposition from cell cycle progression, resulting in mitotic defects. Thus, CENP-A deposition is controlled by a two-step regulatory paradigm comprised of Plk1 and CDK that is crucial for genomic integrity. PMID:25036634

  19. SUMOylation regulates polo-like kinase 1-interacting checkpoint helicase (PICH) during mitosis.

    PubMed

    Sridharan, Vinidhra; Park, Hyewon; Ryu, Hyunju; Azuma, Yoshiaki

    2015-02-01

    Mitotic SUMOylation has an essential role in faithful chromosome segregation in eukaryotes, although its molecular consequences are not yet fully understood. In Xenopus egg extract assays, we showed that poly(ADP-ribose) polymerase 1 (PARP1) is modified by SUMO2/3 at mitotic centromeres and that its enzymatic activity could be regulated by SUMOylation. To determine the molecular consequence of mitotic SUMOylation, we analyzed SUMOylated PARP1-specific binding proteins. We identified Polo-like kinase 1-interacting checkpoint helicase (PICH) as an interaction partner of SUMOylated PARP1 in Xenopus egg extract. Interestingly, PICH also bound to SUMOylated topoisomerase IIα (TopoIIα), a major centromeric small ubiquitin-like modifier (SUMO) substrate. Purified recombinant human PICH interacted with SUMOylated substrates, indicating that PICH directly interacts with SUMO, and this interaction is conserved among species. Further analysis of mitotic chromosomes revealed that PICH localized to the centromere independent of mitotic SUMOylation. Additionally, we found that PICH is modified by SUMO2/3 on mitotic chromosomes and in vitro. PICH SUMOylation is highly dependent on protein inhibitor of activated STAT, PIASy, consistent with other mitotic chromosomal SUMO substrates. Finally, the SUMOylation of PICH significantly reduced its DNA binding capability, indicating that SUMOylation might regulate its DNA-dependent ATPase activity. Collectively, our findings suggest a novel SUMO-mediated regulation of the function of PICH at mitotic centromeres. PMID:25564610

  20. Pharmacophore modeling and virtual screening studies of checkpoint kinase 1 inhibitors.

    PubMed

    Chen, Jin-Juan; Liu, Ting-Lin; Yang, Li-Jun; Li, Lin-Li; Wei, Yu-Quan; Yang, Sheng-Yong

    2009-07-01

    In this study, chemical feature-based 3-dimensional (3D) pharmacophore models of Checkpoint kinase 1 (Chk1) inhibitors were developed based on the known inhibitors of Chk1. The best pharmacophore model Hypo1 was characterized by the best correlation coefficient (0.9577), and the lowest root mean square deviation (0.8871). Hypo1 consists of one hydrogen-bond acceptor, one hydrogen-bond donor, and two hydrophobic features, as well as one excluded volume. This pharmacophore model was further validated by both test set and cross validation methods. A comparison analysis of Hypo1 with chemical features in the active site of Chk1 indicates that the pharmacophore model Hypo1 can correctly reflect the interactions between Chk1 and its ligands. Then Hypo1 was used to screen chemical databases, including Specs and Chinese Nature Product Database (CNPD) for potential lead compounds. The hit compounds were subsequently subjected to filtering by Lipinski's rule of five and docking study to refine the retrieved hits. Finally some of the most potent (estimated) compounds were selected from the final refined hits and suggested for further experimental investigation. PMID:19571415

  1. Serum- and glucocorticoid-inducible kinase 1 in the regulation of renal and extrarenal potassium transport.

    PubMed

    Lang, Florian; Vallon, Volker

    2012-02-01

    Serum- and glucocorticoid inducible-kinase 1 (SGK1) is an early gene transcriptionally upregulated by cell stress such as cell shrinkage and hypoxia and several hormones including gluco- and mineralocorticoids. It is activated by insulin and growth factors. SGK1 is a powerful regulator of a wide variety of channels and transporters. The present review describes the role of SGK1 in the regulation of potassium (K(+)) channels, K(+) transporters and K(+) homeostasis. SGK1-regulated K(+) channels include renal outer medullary K+ channel, Kv1.3, Kv1.5, KCNE1/KCNQ1, KCNQ4 and, via regulation of calcium (Ca(2+)) entry, Ca(2+)-sensitive K(+) channels. SGK1-sensitive transporters include sodium-potassium-chloride cotransporter 2 and sodium/potassium-adenosine triphosphatase. SGK1-dependent regulation of K(+) channels and K(+) transport contributes to the stimulation of renal K(+) excretion following high K(+) intake, to insulin-induced cellular K(+) uptake and hypokalemia, to inhibition of insulin release by glucocorticoids, to stimulation of mast cell degranulation and gastric acid secretion, and to cardiac repolarization. Thus, SGK1 has a profound effect on K(+) homeostasis and on a multitude of K(+)-sensitive cellular functions. PMID:22038256

  2. Death-associated protein kinase 1 promotes growth of p53-mutant cancers.

    PubMed

    Zhao, Jing; Zhao, Dekuang; Poage, Graham M; Mazumdar, Abhijit; Zhang, Yun; Hill, Jamal L; Hartman, Zachary C; Savage, Michelle I; Mills, Gordon B; Brown, Powel H

    2015-07-01

    Estrogen receptor-negative (ER-negative) breast cancers are extremely aggressive and associated with poor prognosis. In particular, effective treatment strategies are limited for patients diagnosed with triple receptor-negative breast cancer (TNBC), which also carries the worst prognosis of all forms of breast cancer; therefore, extensive studies have focused on the identification of molecularly targeted therapies for this tumor subtype. Here, we sought to identify molecular targets that are capable of suppressing tumorigenesis in TNBCs. Specifically, we found that death-associated protein kinase 1 (DAPK1) is essential for growth of p53-mutant cancers, which account for over 80% of TNBCs. Depletion or inhibition of DAPK1 suppressed growth of p53-mutant but not p53-WT breast cancer cells. Moreover, DAPK1 inhibition limited growth of other p53-mutant cancers, including pancreatic and ovarian cancers. DAPK1 mediated the disruption of the TSC1/TSC2 complex, resulting in activation of the mTOR pathway. Our studies demonstrated that high DAPK1 expression causes increased cancer cell growth and enhanced signaling through the mTOR/S6K pathway; evaluation of multiple breast cancer patient data sets revealed that high DAPK1 expression associates with worse outcomes in individuals with p53-mutant cancers. Together, our data support targeting DAPK1 as a potential therapeutic strategy for p53-mutant cancers. PMID:26075823

  3. Cellular trafficking of the IL-1RI-associated kinase-1 requires intact kinase activity

    SciTech Connect

    Boel, Gaby-Fleur . E-mail: boel@mail.dife.de; Jurrmann, Nadine; Brigelius-Flohe, Regina

    2005-06-24

    Upon stimulation of cells with interleukin-1 (IL-1) the IL-1 receptor type I (IL-1RI) associated kinase-1 (IRAK-1) transiently associates to and dissociates from the IL-1RI and thereafter translocates into the nucleus. Here we show that nuclear translocation of IRAK-1 depends on its kinase activity since translocation was not observed in EL-4 cells overexpressing a kinase negative IRAK-1 mutant (EL-4{sup IRAK-1-K239S}). IRAK-1 itself, an endogenous substrate with an apparent molecular weight of 24 kDa (p24), and exogenous substrates like histone and myelin basic protein are phosphorylated by nuclear located IRAK-1. Phosphorylation of p24 cannot be detected in EL-4{sup IRAK-1-K239S} cells. IL-1-dependent recruitment of IRAK-1 to the IL-1RI and subsequent phosphorylation of IRAK-1 is a prerequisite for nuclear translocation of IRAK-1. It is therefore concluded that intracellular localization of IRAK-1 depends on its kinase activity and that IRAK-1 may also function as a kinase in the nucleus as shown by a new putative endogenous substrate.

  4. Genetic depletion of Polo-like kinase 1 leads to embryonic lethality due to mitotic aberrancies.

    PubMed

    Wachowicz, Paulina; Fernández-Miranda, Gonzalo; Marugán, Carlos; Escobar, Beatriz; de Cárcer, Guillermo

    2016-07-01

    Polo-like kinase 1 (PLK1) is a serine/threonine kinase that plays multiple and essential roles during the cell division cycle. Its inhibition in cultured cells leads to severe mitotic aberrancies and cell death. Whereas previous reports suggested that Plk1 depletion in mice leads to a non-mitotic arrest in early embryos, we show here that the bi-allelic Plk1 depletion in mice certainly results in embryonic lethality due to extensive mitotic aberrations at the morula stage, including multi- and mono-polar spindles, impaired chromosome segregation and cytokinesis failure. In addition, the conditional depletion of Plk1 during mid-gestation leads also to severe mitotic aberrancies. Our data also confirms that Plk1 is completely dispensable for mitotic entry in vivo. On the other hand, Plk1 haploinsufficient mice are viable, and Plk1-heterozygous fibroblasts do not harbor any cell cycle alterations. Plk1 is overexpressed in many human tumors, suggesting a therapeutic benefit of inhibiting Plk1, and specific small-molecule inhibitors for this kinase are now being evaluated in clinical trials. Therefore, the different Plk1 mouse models here presented are a valuable tool to reexamine the relevance of the mitotic kinase Plk1 during mammalian development and animal physiology. PMID:27417127

  5. Design of Targeted Inhibitors of Polo-like Kinase 1 (Plk1)

    NASA Astrophysics Data System (ADS)

    Dalafave, D. S.

    2011-03-01

    Computational design of small molecule inhibitors of Polo-like Kinase 1 (Plk1) is presented. Plk1, which regulates cell cycle, is often overexpressed in cancers. Its downregulation was shown to inhibit cancer progression. Most inhibitors of kinases' interact with the highly conserved ATP binding site. This makes the development of Plk1-specific inhibitors challenging, since different kinases have similar ATP sites. However, Plk1 also contains the polo-box domain (PBD), which is absent from other kinases. In this study, the PBD site was used as a target for designed Plk1 inhibitors. Common structural features of experimentally known Plk1 ligands were first identified. The information was used to design putative small molecules that specifically bonded Plk1. Druglikeness and possible toxicities of the designed molecules were determined. Molecules with no implied toxicities and optimal druglikeness were used for docking studies. The docking studies identified several molecules that made stable complexes with the Plk1 PBD site. Possible utilization of the designed molecules in drugs against cancers with overexpressed Plk1 is discussed.

  6. Casein kinase 1α–dependent feedback loop controls autophagy in RAS-driven cancers

    PubMed Central

    Cheong, Jit Kong; Zhang, Fuquan; Chua, Pei Jou; Bay, Boon Huat; Thorburn, Andrew; Virshup, David M.

    2015-01-01

    Activating mutations in the RAS oncogene are common in cancer but are difficult to therapeutically target. RAS activation promotes autophagy, a highly regulated catabolic process that metabolically buffers cells in response to diverse stresses. Here we report that casein kinase 1α (CK1α), a ubiquitously expressed serine/threonine kinase, is a key negative regulator of oncogenic RAS–induced autophagy. Depletion or pharmacologic inhibition of CK1α enhanced autophagic flux in oncogenic RAS–driven human fibroblasts and multiple cancer cell lines. FOXO3A, a master longevity mediator that transcriptionally regulates diverse autophagy genes, was a critical target of CK1α, as depletion of CK1α reduced levels of phosphorylated FOXO3A and increased expression of FOXO3A-responsive genes. Oncogenic RAS increased CK1α protein abundance via activation of the PI3K/AKT/mTOR pathway. In turn, elevated levels of CK1α increased phosphorylation of nuclear FOXO3A, thereby inhibiting transactivation of genes critical for RAS-induced autophagy. In both RAS-driven cancer cells and murine xenograft models, pharmacologic CK1α inactivation synergized with lysosomotropic agents to inhibit growth and promote tumor cell death. Together, our results identify a kinase feedback loop that influences RAS-dependent autophagy and suggest that targeting CK1α-regulated autophagy offers a potential therapeutic opportunity to treat oncogenic RAS–driven cancers. PMID:25798617

  7. Soluble fms-like tyrosine kinase 1 promotes angiotensin II sensitivity in preeclampsia.

    PubMed

    Burke, Suzanne D; Zsengellér, Zsuzsanna K; Khankin, Eliyahu V; Lo, Agnes S; Rajakumar, Augustine; DuPont, Jennifer J; McCurley, Amy; Moss, Mary E; Zhang, Dongsheng; Clark, Christopher D; Wang, Alice; Seely, Ellen W; Kang, Peter M; Stillman, Isaac E; Jaffe, Iris Z; Karumanchi, S Ananth

    2016-07-01

    Preeclampsia is a hypertensive disorder of pregnancy in which patients develop profound sensitivity to vasopressors, such as angiotensin II, and is associated with substantial morbidity for the mother and fetus. Enhanced vasoconstrictor sensitivity and elevations in soluble fms-like tyrosine kinase 1 (sFLT1), a circulating antiangiogenic protein, precede clinical signs and symptoms of preeclampsia. Here, we report that overexpression of sFlt1 in pregnant mice induced angiotensin II sensitivity and hypertension by impairing endothelial nitric oxide synthase (eNOS) phosphorylation and promoting oxidative stress in the vasculature. Administration of the NOS inhibitor l-NAME to pregnant mice recapitulated the angiotensin sensitivity and oxidative stress observed with sFlt1 overexpression. Sildenafil, an FDA-approved phosphodiesterase 5 inhibitor that enhances NO signaling, reversed sFlt1-induced hypertension and angiotensin II sensitivity in the preeclampsia mouse model. Sildenafil treatment also improved uterine blood flow, decreased uterine vascular resistance, and improved fetal weights in comparison with untreated sFlt1-expressing mice. Finally, sFLT1 protein expression inversely correlated with reductions in eNOS phosphorylation in placental tissue of human preeclampsia patients. These data support the concept that endothelial dysfunction due to high circulating sFLT1 may be the primary event leading to enhanced vasoconstrictor sensitivity that is characteristic of preeclampsia and suggest that targeting sFLT1-induced pathways may be an avenue for treating preeclampsia and improving fetal outcomes. PMID:27270170

  8. Circadian Metabolic Regulation through Crosstalk between Casein Kinase 1δ and Transcriptional Coactivator PGC-1α

    PubMed Central

    Li, Siming; Chen, Xiao-Wei; Yu, Lei; Saltiel, Alan R.

    2011-01-01

    Circadian clock coordinates behavior and physiology in mammals in response to light and feeding cycles. Disruption of normal clock function is associated with increased risk for cardiovascular and metabolic diseases, underscoring the emerging concept that temporal regulation of tissue metabolism is a fundamental aspect of energy homeostasis. We have previously demonstrated that transcriptional coactivator, peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), coordinates circadian metabolic rhythms through simultaneous regulation of metabolic and clock gene expression. In this study, we found that PGC-1α physically interacts with, and is phosphorylated by, casein kinase 1δ (CK1δ), a core component of the circadian pacemaker. CK1δ represses the transcriptional function of PGC-1α in cultured hepatocytes, resulting in decreased gluconeogenic gene expression and glucose secretion. At the molecular level, CK1δ phosphorylation of PGC-1α within its arginine/serine-rich domain enhances its degradation through the proteasome system. Together, these results elucidate a novel mechanism through which circadian pacemaker transduces timing signals to the metabolic regulatory network that controls hepatic energy metabolism. PMID:22052997

  9. Partitioning of casein kinase 1-like 6 to late endosome-like vesicles.

    PubMed

    Ben-Nissan, Gili; Yang, Yaodong; Lee, Jung-Youn

    2010-04-01

    Members of the casein kinase 1 family are highly conserved protein Ser/Thr kinases found in all eukaryotes. They are involved in various cellular, physiological, and developmental processes, but the role of this family of kinase in plants is not well known. By localization studies employing fluorescent live cell imaging and biochemical membrane fractionation, here we showed that Arabidopsis casein kinase-like 6 (CKL6) localizes to motile vesicle-like structures that cofractionate with prevacuolar markers. They were found both in the cytoplasm and at the cell periphery and were motile within the cell. Apparently, this motility was dependent on actin filaments and CKL6-positive vesicles partially colocalized with a late endosomal compartment. However, CKL6-positive structures were not sensitive to brefeldin A nor wortmannin treatment, suggesting that they may belong to a novel compartment. Association of CKL6-positive structures with the cell periphery at the cellular junctions was detected after separation of the protoplasts by plasmolysis. Collectively, these data led us to propose that CKL6 is associated with late endosomal-like compartments that are not fully characterized and may play a role in cellular processes important for regulating components in membrane trafficking. PMID:19941015

  10. Doublecortin-like kinase 1 expression associates with breast cancer with neuroendocrine differentiation

    PubMed Central

    Liu, Yu-Hong; Tsang, Julia Y.S.; Ni, Yun-Bi; Hlaing, Thazin; Chan, Siu-Ki; Chan, Kui-Fat; Ko, Chun-Wai; Mujtaba, S. Shafaq; Tse, Gary M.

    2016-01-01

    Doublecortin-like kinase 1 (DCLK1), a microtubule associated kinase, has recently been proposed to be a putative marker for stemness and adverse prognosis in gastrointestinal cancers. However, it is not clear whether the protein also plays similar roles in breast cancer. Here, the expression of DCLK1 was analyzed in a large cohort of invasive breast cancers (IBC) by immunohistochemistry. DCKL1 was associated with favorable clinico-pathologic features, namely lower histologic grade, absence of lymphovascular invasion, fibrotic focus, necrosis and lower pN stage (p≤0.045). Additionally, independent significant correlations were found with estrogen receptor and neuroendocrine markers (p ≤0.019), implicating its relationship with IBC with neuroendocrine differentiation (IBC-NED). In the current cohort, IBC-NED showed worse outcome than luminal cancers without NED (hazard ratio=1.756, p=0.041). Interestingly, within the IBC-NED group, DCLK1 was found to be a good prognostic factor (hazard ratio =0.288, p=0.011). These findings were in contrast to those in gastrointestinal cancers, suggesting different functional roles of DCLK1 in different types of cancers. In clinical practice, NED is not routinely assessed; thus IBC-NED are not well studied. Its poor outcome and significant heterogeneity warrants more attention. DCLK1 expression could aid in the prognostication and management of this special cancer subtype. PMID:26621833

  11. Death-associated protein kinase 1 promotes growth of p53-mutant cancers

    PubMed Central

    Zhao, Jing; Zhao, Dekuang; Poage, Graham M.; Mazumdar, Abhijit; Zhang, Yun; Hill, Jamal L.; Hartman, Zachary C.; Savage, Michelle I.; Mills, Gordon B.; Brown, Powel H.

    2015-01-01

    Estrogen receptor–negative (ER-negative) breast cancers are extremely aggressive and associated with poor prognosis. In particular, effective treatment strategies are limited for patients diagnosed with triple receptor–negative breast cancer (TNBC), which also carries the worst prognosis of all forms of breast cancer; therefore, extensive studies have focused on the identification of molecularly targeted therapies for this tumor subtype. Here, we sought to identify molecular targets that are capable of suppressing tumorigenesis in TNBCs. Specifically, we found that death-associated protein kinase 1 (DAPK1) is essential for growth of p53-mutant cancers, which account for over 80% of TNBCs. Depletion or inhibition of DAPK1 suppressed growth of p53-mutant but not p53-WT breast cancer cells. Moreover, DAPK1 inhibition limited growth of other p53-mutant cancers, including pancreatic and ovarian cancers. DAPK1 mediated the disruption of the TSC1/TSC2 complex, resulting in activation of the mTOR pathway. Our studies demonstrated that high DAPK1 expression causes increased cancer cell growth and enhanced signaling through the mTOR/S6K pathway; evaluation of multiple breast cancer patient data sets revealed that high DAPK1 expression associates with worse outcomes in individuals with p53-mutant cancers. Together, our data support targeting DAPK1 as a potential therapeutic strategy for p53-mutant cancers. PMID:26075823

  12. Tank binding kinase 1 is a centrosome-associated kinase necessary for microtubule dynamics and mitosis

    PubMed Central

    Pillai, Smitha; Nguyen, Jonathan; Johnson, Joseph; Haura, Eric; Coppola, Domenico; Chellappan, Srikumar

    2015-01-01

    TANK Binding Kinase 1 (TBK1) is a non-canonical IκB kinase that contributes to KRAS-driven lung cancer. Here we report that TBK1 plays essential roles in mammalian cell division. Specifically, levels of active phospho-TBK1 increase during mitosis and localize to centrosomes, mitotic spindles and midbody, and selective inhibition or silencing of TBK1 triggers defects in spindle assembly and prevents mitotic progression. TBK1 binds to the centrosomal protein CEP170 and to the mitotic apparatus protein NuMA, and both CEP170 and NuMA are TBK1 substrates. Further, TBK1 is necessary for CEP170 centrosomal localization and binding to the microtubule depolymerase Kif2b, and for NuMA binding to dynein. Finally, selective disruption of the TBK1–CEP170 complex augments microtubule stability and triggers defects in mitosis, suggesting that TBK1 functions as a mitotic kinase necessary for microtubule dynamics and mitosis. PMID:26656453

  13. Casein Kinase 1 α Phosphorylates the Wnt Regulator Jade-1 and Modulates Its Activity*

    PubMed Central

    Borgal, Lori; Rinschen, Markus M.; Dafinger, Claudia; Hoff, Sylvia; Reinert, Matthäus J.; Lamkemeyer, Tobias; Lienkamp, Soeren S.; Benzing, Thomas; Schermer, Bernhard

    2014-01-01

    Tight regulation of Wnt/β-catenin signaling is critical for vertebrate development and tissue maintenance, and deregulation can lead to a host of disease phenotypes, including developmental disorders and cancer. Proteins associated with primary cilia and centrosomes have been demonstrated to negatively regulate canonical Wnt signaling in interphase cells. The plant homeodomain zinc finger protein Jade-1 can act as an E3 ubiquitin ligase-targeting β-catenin for proteasomal degradation and concentrates at the centrosome and ciliary basal body in addition to the nucleus in interphase cells. We demonstrate that the destruction complex component casein kinase 1α (CK1α) phosphorylates Jade-1 at a conserved SLS motif and reduces the ability of Jade-1 to inhibit β-catenin signaling. Consistently, Jade-1 lacking the SLS motif is more effective than wild-type Jade-1 in reducing β-catenin-induced secondary axis formation in Xenopus laevis embryos in vivo. Interestingly, CK1α also phosphorylates β-catenin and the destruction complex component adenomatous polyposis coli at a similar SLS motif to the effect that β-catenin is targeted for degradation. The opposing effect of Jade-1 phosphorylation by CK1α suggests a novel example of the dual functions of CK1α activity to either oppose or promote canonical Wnt signaling in a context-dependent manner. PMID:25100726

  14. The Indispensable Role of Cyclin-Dependent Kinase 1 in Skeletal Development

    PubMed Central

    Saito, Masanori; Mulati, Mieradili; Talib, S. Zakiah A.; Kaldis, Philipp; Takeda, Shu; Okawa, Atsushi; Inose, Hiroyuki

    2016-01-01

    Skeletal development is tightly regulated through the processes of chondrocyte proliferation and differentiation. Although the involvement of transcription and growth factors on the regulation of skeletal development has been extensively studied, the role of cell cycle regulatory proteins in this process remains elusive. To date, through cell-specific loss-of-function experiments in vivo, no cell cycle regulatory proteins have yet been conclusively shown to regulate skeletal development. Here, we demonstrate that cyclin-dependent kinase 1 (Cdk1) regulates skeletal development based on chondrocyte-specific loss-of-function experiments performed in a mouse model. Cdk1 is highly expressed in columnar proliferative chondrocytes and is greatly downregulated upon differentiation into hypertrophic chondrocytes. Cdk1 is essential for proper chondrocyte proliferation and deletion of Cdk1 resulted in accelerated differentiation of chondrocytes. In vitro and ex vivo analyses revealed that Cdk1 is an essential cell cycle regulatory protein for parathyroid hormone-related peptide (PTHrP) signaling pathway, which is critical to chondrocyte proliferation and differentiation. These results demonstrate that Cdk1 functions as a molecular switch from proliferation to hypertrophic differentiation of chondrocytes and thus is indispensable for skeletal development. Given the availability of inhibitors of Cdk1 activity, our results could provide insight for the treatment of diseases involving abnormal chondrocyte proliferation, such as osteoarthritis. PMID:26860366

  15. Protein kinase Cδ regulates vaccinia-related kinase 1 in DNA damage–induced apoptosis

    PubMed Central

    Park, Choon-Ho; Choi, Bo-Hwa; Jeong, Min-Woo; Kim, Sangjune; Kim, Wanil; Song, Yun Seon; Kim, Kyong-Tai

    2011-01-01

    Vaccinia-related kinase 1 (VRK1) is a novel serine/threonine kinase that plays an important role in cell proliferation. However, little is known about the upstream regulators of VRK1 activity. Here we provide evidence for a role of protein kinase Cδ (PKCδ) in the regulation of murine VRK1. We show that PKCδ interacts with VRK1, phosphorylates the Ser-355 residue in the putative regulatory region, and negatively regulates its kinase activity in vitro. Intriguingly, PKCδ-induced cell death was facilitated by phosphorylation of VRK1 when cells were exposed to a DNA-damaging agent. In addition, p53 played a critical role in the regulation of DNA damage–induced cell death accompanied by PKCδ-mediated modulation of VRK1. In p53-deficient cells, PKCδ-mediated phosphorylation of VRK1 had no effect on cell viability. However, cells overexpressing p53 exhibited significant reduction of cell viability when cotransfected with both VRK1 and PKCδ. Taken together, these results indicate that PKCδ regulates phosphorylation and down-regulation of VRK1, thereby contributing to cell cycle arrest and apoptotic cell death in a p53-dependent manner. PMID:21346188

  16. A New Antibiotic to the Rescue?

    MedlinePlus

    ... to the Rescue? Experimental drug shows promise against MRSA superbug in animal trials To use the sharing ... treated animals infected with the so-called "superbug" MRSA -- methicillin-resistant staphylococcus aureus . The results are "important ...

  17. Fire and Rescue Technology. Resources in Technology.

    ERIC Educational Resources Information Center

    Valesey, Brigitte G.

    1997-01-01

    Provides occupational information about fire and rescue operations personnel, such as fire science, fire protection engineering, emergency medical technicians, and firefighters. Provides information about organizations in these fields. (JOW)

  18. A New Antibiotic to the Rescue?

    MedlinePlus

    ... gov/medlineplus/news/fullstory_159417.html A New Antibiotic to the Rescue? Experimental drug shows promise against ... THURSDAY, June 16, 2016 (HealthDay News) -- An experimental antibiotic has shown promise against a dangerous drug-resistant ...

  19. NASA's Search-and-Rescue Technology

    NASA Video Gallery

    This animation depicts the next-generation search and rescue system, the DASS. Under this system, instruments used to relay emergency beacon signals will be installed on GPS satellites. When one em...

  20. Hubble Space Telescope Crew Rescue Analysis

    NASA Technical Reports Server (NTRS)

    Hamlin, Teri L.; Canga, Michael A.; Cates, Grant R.

    2010-01-01

    In the aftermath of the 2003 Columbia accident, NASA removed the Hubble Space Telescope (HST) Servicing Mission 4 (SM4) from the Space Shuttle manifest. Reasons cited included concerns that the risk of flying the mission would be too high. The HST SM4 was subsequently reinstated and flown as Space Transportation System (STS)-125 because of improvements in the ascent debris environment, the development of techniques for astronauts to perform on orbit repairs to damaged thermal protection, and the development of a strategy to provide a viable crew rescue capability. However, leading up to the launch of STS-125, the viability of the HST crew rescue capability was a recurring topic. For STS-125, there was a limited amount of time available to perform a crew rescue due to limited consumables (power, oxygen, etc.) available on the Orbiter. The success of crew rescue depended upon several factors, including when a problem was identified; when and what actions, such as powering down, were begun to conserve consumables; and where the Launch on Need (LON) vehicle was in its ground processing cycle. Crew rescue success also needed to be weighed against preserving the Orbiter s ability to have a landing option in case there was a problem with the LON vehicle. This paper focuses on quantifying the HST mission loss of crew rescue capability using Shuttle historical data and various power down strategies. Results from this effort supported NASA s decision to proceed with STS-125, which was successfully completed on May 24th 2009.

  1. Factors influencing mine rescue team behaviors.

    PubMed

    Jansky, Jacqueline H; Kowalski-Trakofler, K M; Brnich, M J; Vaught, C

    2016-01-01

    A focus group study of the first moments in an underground mine emergency response was conducted by the National Institute for Occupational Safety and Health (NIOSH), Office for Mine Safety and Health Research. Participants in the study included mine rescue team members, team trainers, mine officials, state mining personnel, and individual mine managers. A subset of the data consists of responses from participants with mine rescue backgrounds. These responses were noticeably different from those given by on-site emergency personnel who were at the mine and involved with decisions made during the first moments of an event. As a result, mine rescue team behavior data were separated in the analysis and are reported in this article. By considering the responses from mine rescue team members and trainers, it was possible to sort the data and identify seven key areas of importance to them. On the basis of the responses from the focus group participants with a mine rescue background, the authors concluded that accurate and complete information and a unity of purpose among all command center personnel are two of the key conditions needed for an effective mine rescue operation. PMID:26963229

  2. 30 CFR 49.15 - Mine rescue station.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Mine rescue station. 49.15 Section 49.15 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Coal Mines § 49.15 Mine rescue station. (a) Every...

  3. 30 CFR 49.5 - Mine rescue station.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 30 Mineral Resources 1 2013-07-01 2013-07-01 false Mine rescue station. 49.5 Section 49.5 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Metal and Nonmetal Mines § 49.5 Mine rescue station. (a)...

  4. 30 CFR 49.5 - Mine rescue station.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 30 Mineral Resources 1 2014-07-01 2014-07-01 false Mine rescue station. 49.5 Section 49.5 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Metal and Nonmetal Mines § 49.5 Mine rescue station. (a)...

  5. 30 CFR 49.15 - Mine rescue station.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 30 Mineral Resources 1 2012-07-01 2012-07-01 false Mine rescue station. 49.15 Section 49.15 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Coal Mines § 49.15 Mine rescue station. (a) Every...

  6. 30 CFR 49.15 - Mine rescue station.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Mine rescue station. 49.15 Section 49.15 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Coal Mines § 49.15 Mine rescue station. (a) Every...

  7. 30 CFR 49.15 - Mine rescue station.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 30 Mineral Resources 1 2013-07-01 2013-07-01 false Mine rescue station. 49.15 Section 49.15 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Coal Mines § 49.15 Mine rescue station. (a) Every...

  8. 30 CFR 49.5 - Mine rescue station.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 30 Mineral Resources 1 2012-07-01 2012-07-01 false Mine rescue station. 49.5 Section 49.5 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Metal and Nonmetal Mines § 49.5 Mine rescue station. (a)...

  9. 30 CFR 49.15 - Mine rescue station.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 30 Mineral Resources 1 2014-07-01 2014-07-01 false Mine rescue station. 49.15 Section 49.15 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING MINE RESCUE TEAMS Mine Rescue Teams for Underground Coal Mines § 49.15 Mine rescue station. (a) Every...

  10. Targeting polo-like kinase 1 suppresses essential functions of alloreactive T cells.

    PubMed

    Berges, Carsten; Chatterjee, Manik; Topp, Max S; Einsele, Hermann

    2016-06-01

    Acute graft-versus-host disease (aGvHD) is still a major cause of transplant-related mortality after allogeneic stem cell transplantation (ASCT). It requires immunosuppressive treatments that broadly abrogate T cell responses including beneficial ones directed against tumor cells or infective pathogens. Polo-like kinase 1 (PLK1) is overexpressed in many cancer types including leukemia, and clinical studies demonstrated that targeting PLK1 using selective PLK1 inhibitors resulted in inhibition of proliferation and induction of apoptosis predominantly in tumor cells, supporting the feasibility of PLK1 as target for anticancer therapy. Here, we show that activation of alloreactive T cells (Tallo) up-regulate expression of PLK1, suggesting that PLK1 is a potential new candidate for dual therapy of aGvHD and leukemia after ASCT. Inhibition of PLK1, using PLK1-specific inhibitor GSK461364A selectively depletes Tallo by preventing activation and by inducing apoptosis in already activated Tallo, while memory T cells are preserved. Activated Tallo cells which survive exposure to PLK1 undergo inhibition of proliferation by induction of G2/M cell cycle arrest, which is accompanied by accumulation of cell cycle regulator proteins p21(WAF/CIP1), p27(Kip1), p53 and cyclin B1, whereas abundance of CDK4 decreased. We also show that suppressive effects of PLK1 inhibition on Tallo were synergistically enhanced by concomitant inhibition of molecular chaperone Hsp90. Taken together, our data suggest that PLK1 inhibition represents a reasonable dual strategy to suppress residual tumor growth and efficiently deplete Tallo, and thus provide a rationale to selectively prevent and treat aGvHD. PMID:26724940

  11. TANK-binding kinase-1 broadly affects oyster immune response to bacteria and viruses.

    PubMed

    Tang, Xueying; Huang, Baoyu; Zhang, Linlin; Li, Li; Zhang, Guofan

    2016-09-01

    As a benthic filter feeder of estuaries, the immune system of oysters provides one of the best models for studying the genetic and molecular basis of the innate immune pathway in marine invertebrates and examining the influence of environmental factors on the immune system. Here, the molecular function of molluscan TANK-binding kinase-1 (TBK1) (which we named CgTBK1) was studied in the Pacific oyster, Crassostrea gigas. Compared with known TBK1 proteins in other model organisms, CgTBK1 contains a conserved S-TKc domain and a coiled coil domain at the N- and C-terminals but lacks an important ubiquitin domain. Quantitative real-time PCR analysis revealed that the expression level of CgTBK1 was ubiquitous in all selected tissues, with highest expression in the gills. CgTBK1 expression was significantly upregulated in response to infections with Vibrio alginolyticus, ostreid herpesvirus 1 (OsHV-1 reference strain and μvar), and polyinosinic:polycytidylic acid sodium salt, suggesting its broad function in immune response. Subcellular localization showed the presence of CgTBK1 in the cytoplasm of HeLa cells, suggesting its potential function as the signal transducer between the receptor and transcription factor. We further demonstrated that CgTBK1 interacted with CgSTING in HEK293T cells, providing evidence that CgTBK1 could be activated by direct binding to CgSTING. In summary, we characterized the TBK1 gene in C. gigas and demonstrated its role in the innate immune response to pathogen infections. PMID:27422757

  12. Sphingosine Kinase 1 Deficiency Confers Protection against Hyperoxia-Induced Bronchopulmonary Dysplasia in a Murine Model

    PubMed Central

    Harijith, Anantha; Pendyala, Srikanth; Reddy, Narsa M.; Bai, Tao; Usatyuk, Peter V.; Berdyshev, Evgeny; Gorshkova, Irina; Huang, Long Shuang; Mohan, Vijay; Garzon, Steve; Kanteti, Prasad; Reddy, Sekhar P.; Raj, J. Usha; Natarajan, Viswanathan

    2014-01-01

    Bronchopulmonary dysplasia of the premature newborn is characterized by lung injury, resulting in alveolar simplification and reduced pulmonary function. Exposure of neonatal mice to hyperoxia enhanced sphingosine-1-phosphate (S1P) levels in lung tissues; however, the role of increased S1P in the pathobiological characteristics of bronchopulmonary dysplasia has not been investigated. We hypothesized that an altered S1P signaling axis, in part, is responsible for neonatal lung injury leading to bronchopulmonary dysplasia. To validate this hypothesis, newborn wild-type, sphingosine kinase1−/− (Sphk1−/−), sphingosine kinase 2−/− (Sphk2−/−), and S1P lyase+/− (Sgpl1+/−) mice were exposed to hyperoxia (75%) from postnatal day 1 to 7. Sphk1−/−, but not Sphk2−/− or Sgpl1+/−, mice offered protection against hyperoxia-induced lung injury, with improved alveolarization and alveolar integrity compared with wild type. Furthermore, SphK1 deficiency attenuated hyperoxia-induced accumulation of IL-6 in bronchoalveolar lavage fluids and NADPH oxidase (NOX) 2 and NOX4 protein expression in lung tissue. In vitro experiments using human lung microvascular endothelial cells showed that exogenous S1P stimulated intracellular reactive oxygen species (ROS) generation, whereas SphK1 siRNA, or inhibitor against SphK1, attenuated hyperoxia-induced S1P generation. Knockdown of NOX2 and NOX4, using specific siRNA, reduced both basal and S1P-induced ROS formation. These results suggest an important role for SphK1-mediated S1P signaling–regulated ROS in the development of hyperoxia-induced lung injury in a murine neonatal model of bronchopulmonary dysplasia. PMID:23933064

  13. Skeletal muscle salt inducible kinase 1 promotes insulin resistance in obesity

    PubMed Central

    Nixon, Mark; Stewart-Fitzgibbon, Randi; Fu, Jingqi; Akhmedov, Dmitry; Rajendran, Kavitha; Mendoza-Rodriguez, Maria G.; Rivera-Molina, Yisel A.; Gibson, Micah; Berglund, Eric D.; Justice, Nicholas J.; Berdeaux, Rebecca

    2015-01-01

    Objective Insulin resistance causes type 2 diabetes mellitus and hyperglycemia due to excessive hepatic glucose production and inadequate peripheral glucose uptake. Our objectives were to test the hypothesis that the proposed CREB/CRTC2 inhibitor salt inducible kinase 1 (SIK1) contributes to whole body glucose homeostasis in vivo by regulating hepatic transcription of gluconeogenic genes and also to identify novel SIK1 actions on glucose metabolism. Methods We created conditional (floxed) SIK1-knockout mice and studied glucose metabolism in animals with global, liver, adipose or skeletal muscle Sik1 deletion. We examined cAMP-dependent regulation of SIK1 and the consequences of SIK1 depletion on primary mouse hepatocytes. We probed metabolic phenotypes in tissue-specific SIK1 knockout mice fed high fat diet through hyperinsulinemic-euglycemic clamps and biochemical analysis of insulin signaling. Results SIK1 knockout mice are viable and largely normoglycemic on chow diet. On high fat diet, global SIK1 knockout animals are strikingly protected from glucose intolerance, with both increased plasma insulin and enhanced peripheral insulin sensitivity. Surprisingly, liver SIK1 is not required for regulation of CRTC2 and gluconeogenesis, despite contributions of SIK1 to hepatocyte CRTC2 and gluconeogenesis regulation ex vivo. Sik1 mRNA accumulates in skeletal muscle of obese high fat diet-fed mice, and knockout of SIK1 in skeletal muscle, but not liver or adipose tissue, improves insulin sensitivity and muscle glucose uptake on high fat diet. Conclusions SIK1 is dispensable for glycemic control on chow diet. SIK1 promotes insulin resistance on high fat diet by a cell-autonomous mechanism in skeletal muscle. Our study establishes SIK1 as a promising therapeutic target to improve skeletal muscle insulin sensitivity in obese individuals without deleterious effects on hepatic glucose production. PMID:26844205

  14. Identification of Candidate Cyclin-dependent kinase 1 (Cdk1) Substrates in Mitosis by Quantitative Phosphoproteomics.

    PubMed

    Petrone, Adam; Adamo, Mark E; Cheng, Chao; Kettenbach, Arminja N

    2016-07-01

    Cyclin-dependent kinase 1 (Cdk1) is an essential regulator of many mitotic processes including the reorganization of the cytoskeleton, chromosome segregation, and formation and separation of daughter cells. Deregulation of Cdk1 activity results in severe defects in these processes. Although the role of Cdk1 in mitosis is well established, only a limited number of Cdk1 substrates have been identified in mammalian cells. To increase our understanding of Cdk1-dependent phosphorylation pathways in mitosis, we conducted a quantitative phosphoproteomics analysis in mitotic HeLa cells using two small molecule inhibitors of Cdk1, Flavopiridol and RO-3306. In these analyses, we identified a total of 24,840 phosphopeptides on 4,273 proteins, of which 1,215 phosphopeptides on 551 proteins were significantly reduced by 2.5-fold or more upon Cdk1 inhibitor addition. Comparison of phosphopeptide quantification upon either inhibitor treatment revealed a high degree of correlation (R(2) value of 0.87) between the different datasets. Motif enrichment analysis of significantly regulated phosphopeptides revealed enrichment of canonical Cdk1 kinase motifs. Interestingly, the majority of proteins identified in this analysis contained two or more Cdk1 inhibitor-sensitive phosphorylation sites, were highly connected with other candidate Cdk1 substrates, were enriched at specific subcellular structures, or were part of protein complexes as identified by the CORUM database. Furthermore, candidate Cdk1 substrates were enriched in G2 and M phase-specific genes. Finally, we validated a subset of candidate Cdk1 substrates by in vitro kinase assays. Our findings provide a valuable resource for the cell signaling and mitosis research communities and greatly increase our knowledge of Cdk1 substrates and Cdk1-dependent signaling pathways. PMID:27134283

  15. Functional Dynamics of Polo-Like Kinase 1 at the Centrosome▿ †

    PubMed Central

    Kishi, Kazuhiro; van Vugt, Marcel A. T. M.; Okamoto, Ken-ichi; Hayashi, Yasunori; Yaffe, Michael B.

    2009-01-01

    Polo-like kinase 1 (Plk1) functions as a key regulator of mitotic events by phosphorylating substrate proteins on centrosomes, kinetochores, the mitotic spindle, and the midbody. Through mechanisms that are incompletely understood, Plk1 is released from and relocalizes to different mitotic structures as cells proceed through mitosis. We used fluorescence recovery after photobleaching to examine the kinetics of this process in more detail. We observed that Plk1 displayed a range of different recovery rates that differ at each mitotic substructure and depend on both the Polo-box domain and a functional kinase domain. Upon mitotic entry, centrosomal Plk1 becomes more dynamic, a process that is directly enhanced by Plk1 kinase activity. In contrast, Plk1 displays little dynamic exchange at the midbody, a process that again is modulated by the kinase activity of Plk1. Our findings suggest that the intrinsic kinase activity of Plk1 triggers its release from early mitotic structures and its relocalization to late mitotic structures. To assess the importance of Plk1 dynamic relocalization, Plk1 was persistently tethered to the centrosome. This resulted in a G2 delay, followed by a prominent prometaphase arrest, as a consequence of defective spindle formation and activation of the spindle checkpoint. The dynamic release of Plk1 from early mitotic structures is thus crucial for mid- to late-stage mitotic events and demonstrates the importance of a fully dynamic Plk1 at the centrosome for proper cell cycle progression. This dependence on dynamic Plk1 was further observed during the mitotic reentry of cells after a DNA damage G2 checkpoint, as this process was significantly delayed upon centrosomal tethering of Plk1. These results indicate that mitotic progression and control of mitotic reentry after DNA damage resides, at least in part, on the dynamic behavior of Plk1. PMID:19307309

  16. Polo-like kinase 1 inhibition sensitizes neuroblastoma cells for vinca alkaloid-induced apoptosis

    PubMed Central

    Czaplinski, Sebastian; Hugle, Manuela; Stiehl, Valerie; Fulda, Simone

    2016-01-01

    High polo-like kinase 1 (PLK1) expression has been linked to poor outcome in neuroblastoma (NB), indicating that it represents a relevant therapeutic target in this malignancy. Here, we identify a synergistic induction of apoptosis by the PLK1 inhibitor BI 2536 and vinca alkaloids in NB cells. Synergistic drug interaction of BI 2536 together with vincristine (VCR), vinblastine (VBL) or vinorelbine (VNR) is confirmed by calculation of combination index (CI). Also, BI 2536 and VCR act in concert to reduce long-term clonogenic survival. Importantly, BI 2536 significantly enhances the antitumor activity of VCR in an in vivo model of NB. Mechanistically, BI 2536/VCR co-treatment triggers prolonged mitotic arrest, which is necessary for BI 2536/VCR-mediated apoptosis, since pharmacological inhibition of mitotic arrest by the CDK1 inhibitor RO-3306 significantly reduces cell death. Prolonged mitotic arrest leads to phosphorylation-mediated inactivation of BCL-2 and BCL-XL as well as downregulation of MCL-1, since inhibition of mitotic arrest by RO-3306 also prevents phosphorylation of BCL-2 and BCL-XL and MCL-1 downregulation. This inactivation of antiapoptotic BCL-2 proteins promotes activation of BAX and BAK, cleavage of caspase-9 and -3 and caspase-dependent apoptosis. Engagement of the mitochondrial pathway of apoptosis is critically required for BI 2536/VCR-induced apoptosis, since ectopic expression of a non-degradable MCL-1 phospho-mutant, BCL-2 overexpression or BAK knockdown significantly reduce BI 2536/VCR-mediated apoptosis. Thus, PLK1 inhibitors may open new perspectives for chemosensitization of NB. PMID:26046302

  17. Removal of Soluble Fms-Like Tyrosine Kinase-1 by Dextran Sulfate Apheresis in Preeclampsia.

    PubMed

    Thadhani, Ravi; Hagmann, Henning; Schaarschmidt, Wiebke; Roth, Bernhard; Cingoez, Tuelay; Karumanchi, S Ananth; Wenger, Julia; Lucchesi, Kathryn J; Tamez, Hector; Lindner, Tom; Fridman, Alexander; Thome, Ulrich; Kribs, Angela; Danner, Marco; Hamacher, Stefanie; Mallmann, Peter; Stepan, Holger; Benzing, Thomas

    2016-03-01

    Preeclampsia is a devastating complication of pregnancy. Soluble Fms-like tyrosine kinase-1 (sFlt-1) is an antiangiogenic protein believed to mediate the signs and symptoms of preeclampsia. We conducted an open pilot study to evaluate the safety and potential efficacy of therapeutic apheresis with a plasma-specific dextran sulfate column to remove circulating sFlt-1 in 11 pregnant women (20-38 years of age) with very preterm preeclampsia (23-32 weeks of gestation, systolic BP ≥140 mmHg or diastolic BP ≥90 mmHg, new onset protein/creatinine ratio >0.30 g/g, and sFlt-1/placental growth factor ratio >85). We evaluated the extent of sFlt-1 removal, proteinuria reduction, pregnancy continuation, and neonatal and fetal safety of apheresis after one (n=6), two (n=4), or three (n=1) apheresis treatments. Mean sFlt-1 levels were reduced by 18% (range 7%-28%) with concomitant reductions of 44% in protein/creatinine ratios. Pregnancy continued for 8 days (range 2-11) and 15 days (range 11-21) in women treated once and multiple times, respectively, compared with 3 days (range 0-14) in untreated contemporaneous preeclampsia controls (n=22). Transient maternal BP reduction during apheresis was managed by withholding pre-apheresis antihypertensive therapy, saline prehydration, and reducing blood flow through the apheresis column. Compared with infants born prematurely to untreated women with and without preeclampsia (n=22 per group), no adverse effects of apheresis were observed. In conclusion, therapeutic apheresis reduced circulating sFlt-1 and proteinuria in women with very preterm preeclampsia and appeared to prolong pregnancy without major adverse maternal or fetal consequences. A controlled trial is warranted to confirm these findings. PMID:26405111

  18. Extracellular signal-regulated kinases 1 and 2 activation in endothelial cells exposed to cyclic strain

    NASA Technical Reports Server (NTRS)

    Ikeda, M.; Takei, T.; Mills, I.; Kito, H.; Sumpio, B. E.

    1999-01-01

    The aim of this study was to determine whether extracellular signal-regulated kinases 1/2 (ERK1/ERK2) are activated and might play a role in enhanced proliferation and morphological change induced by strain. Bovine aortic endothelial cells (BAEC) were subjected to an average of 6 or 10% strain at a rate of 60 cycles/min for up to 4 h. Cyclic strain caused strain- and time-dependent phosphorylation and activation of ERK1/ERK2. Peak phosphorylation and activation of ERK1/ERK2 induced by 10% strain were at 10 min. A specific ERK1/ERK2 kinase inhibitor, PD-98059, inhibited phosphorylation and activation of ERK1/ERK2 but did not inhibit the increased cell proliferation and cell alignment induced by strain. Treatment of BAEC with 2,5-di-tert-butyl-1, 4-benzohydroquinone, to deplete inositol trisphosphate-sensitive calcium storage, and gadolinium chloride, a Ca2+ channel blocker, did not inhibit the activation of ERK1/ERK2. Strain-induced ERK1/ERK2 activation was partly inhibited by the protein kinase C inhibitor calphostin C and completely inhibited by the tyrosine kinase inhibitor genistein. These data suggest that 1) ERK1/ERK2 are not critically involved in the strain-induced cell proliferation and orientation, 2) strain-dependent activation of ERK1/ERK2 is independent of intracellular and extracellular calcium mobilization, and 3) protein kinase C activation and tyrosine kinase regulate strain-induced activation of ERK1/ERK2.

  19. Impairment of Angiogenic Sphingosine Kinase-1/Sphingosine-1-Phosphate Receptors Pathway in Preeclampsia

    PubMed Central

    Dobierzewska, Aneta; Palominos, Macarena; Sanchez, Marianela; Dyhr, Michael; Helgert, Katja; Venegas-Araneda, Pia; Tong, Stephen; Illanes, Sebastian E.

    2016-01-01

    Preeclampsia (PE), is a serious pregnancy disorder characterized in the early gestation by shallow trophoblast invasion, impaired placental neo-angiogenesis, placental hypoxia and ischemia, which leads to maternal and fetal morbidity and mortality. Here we hypothesized that angiogenic sphingosine kinase-1 (SPHK1)/sphingosine-1-phosphate (S1P) receptors pathway is impaired in PE. We found that SPHK1 mRNA and protein expression are down-regulated in term placentae and term chorionic villous explants from patients with PE or severe PE (PES), compared with controls. Moreover, mRNA expression of angiogenic S1PR1 and S1PR3 receptors were decreased in placental samples of PE and PES patients, whereas anti-angiogenic S1PR2 was up-regulated in chorionic villous tissue of PES subjects, pointing to its potential atherogenic and inflammatory properties. Furthermore, in in vitro (JAR cells) and ex vivo (chorionic villous explants) models of placental hypoxia, SPHK1 mRNA and protein were strongly up-regulated under low oxygen tension (1% 02). In contrast, there was no change in SPHK1 expression under the conditions of placental physiological hypoxia (8% 02). In both models, nuclear protein levels of HIF1A were increased at 1% 02 during the time course, but there was no up-regulation at 8% 02, suggesting that SPHK1 and HIF1A might be the part of the same canonical pathway during hypoxia and that both contribute to placental neovascularization during early gestation. Taken together, this study suggest the SPHK1 pathway may play a role in the human early placentation process and may be involved in the pathogenesis of PE. PMID:27284992

  20. Concurrent targeting Akt and sphingosine kinase 1 by A-674563 in acute myeloid leukemia cells.

    PubMed

    Xu, Lin; Zhang, Yanan; Gao, Meng; Wang, Guangping; Fu, Yunfeng

    2016-04-15

    Akt signaling plays a pivotal role in acute myeloid leukemia (AML) development and progression. In the present study, we evaluated the potential anti-AML activity by a novel Akt kinase inhibitor A-674563. Our results showed that A-674563 dose-dependently inhibited survival and proliferation of U937 AML cells and six lines of human AML progenitor cells, yet sparing human peripheral blood mononuclear leukocytes (PBMCs). A-674563 activated caspase-3/9 and apoptosis in the AML cells. Reversely, the pan-caspase inhibitor z-VAD-CHO dramatically alleviated A-674563-induced AML cell apoptosis and cytotoxicity. For the molecular study, we showed that A-674563 blocked Akt activation in U937 cells and human AML progenitor cells. Further, A-674563 decreased sphingosine kinase 1 (SphK1) activity in above AML cells to deplete pro-survival sphingosine-1-phosphate (S1P) and boost pro-apoptotic ceramide production. Such an effect on SphK1 signaling by A-674563 appeared independent of Akt blockage. Significantly, K6PC-5, a novel SphK1 activator, or supplement with S1P attenuated A-674563-induced ceramide production, and subsequent U937 cell death and apoptosis. Importantly, intraperitoneal injection of A-674563 at well-tolerated doses suppressed U937 leukemic xenograft tumor growth in nude mice, whiling significantly improving the animal survival. The results of the current study demonstrate that A-674563 exerts potent anti-leukemic activity in vitro and in vivo, possibly via concurrent targeting Akt and SphK1 signalings. PMID:26920060

  1. Salt-inducible kinase 1 regulates E-cadherin expression and intercellular junction stability.

    PubMed

    Eneling, Kristina; Brion, Laura; Pinto, Vanda; Pinho, Maria J; Sznajder, Jacob I; Mochizuki, Naoki; Emoto, Kazuo; Soares-da-Silva, Patricio; Bertorello, Alejandro M

    2012-08-01

    The protein kinase liver kinase B1 (LKB1) regulates cell polarity and intercellular junction stability. Also, LKB1 controls the activity of salt-inducible kinase 1 (SIK1). The role and relevance of SIK1 and its downstream effectors in linking the LKB1 signals within these processes are partially understood. We hypothesize that SIK1 may link LKB1 signals to the maintenance of epithelial junction stability by regulating E-cadherin expression. Results from our studies using a mouse lung alveolar epithelial (MLE-12) cell line or human renal proximal tubule (HK2) cell line transiently or stably lacking the expression of SIK1 (using SIK1 siRNAs or shRNAs), or with its expression abrogated (sik1(+/+) vs. sik1(-/-) mice), indicate that suppression of SIK1 (∼40%) increases the expression of the transcriptional repressors Snail2 (∼12-fold), Zeb1 (∼100%), Zeb2 (∼50%), and TWIST (∼20-fold) by activating cAMP-response element binding protein. The lack of SIK1 and activation of transcriptional repressors decreases the availability of E-cadherin (mRNA and protein expression by ∼100 and 80%, respectively) and the stability of intercellular junctions in epithelia (decreases in transepithelial resistance). Furthermore, LKB1-mediated increases in E-cadherin expression are impaired in cells where SIK1 has been disabled. We conclude that SIK1 is a key regulator of E-cadherin expression, and thereby contributes to the stability of intercellular junctions. PMID:22522110

  2. Bioinformatic Analysis of Pathogenic Missense Mutations of Activin Receptor Like Kinase 1 Ectodomain

    PubMed Central

    Scotti, Claudia; Olivieri, Carla; Boeri, Laura; Canzonieri, Cecilia; Ornati, Federica; Buscarini, Elisabetta; Pagella, Fabio; Danesino, Cesare

    2011-01-01

    Activin A receptor, type II-like kinase 1 (also called ALK1), is a serine-threonine kinase predominantly expressed on endothelial cells surface. Mutations in its ACVRL1 encoding gene (12q11-14) cause type 2 Hereditary Haemorrhagic Telangiectasia (HHT2), an autosomal dominant multisystem vascular dysplasia. The study of the structural effects of mutations is crucial to understand their pathogenic mechanism. However, while an X-ray structure of ALK1 intracellular domain has recently become available (PDB ID: 3MY0), structure determination of ALK1 ectodomain (ALK1EC) has been elusive so far. We here describe the building of a homology model for ALK1EC, followed by an extensive bioinformatic analysis, based on a set of 38 methods, of the effect of missense mutations at the sequence and structural level. ALK1EC potential interaction mode with its ligand BMP9 was then predicted combining modelling and docking data. The calculated model of the ALK1EC allowed mapping and a preliminary characterization of HHT2 associated mutations. Major structural changes and loss of stability of the protein were predicted for several mutations, while others were found to interfere mainly with binding to BMP9 or other interactors, like Endoglin (CD105), whose encoding ENG gene (9q34) mutations are known to cause type 1 HHT. This study gives a preliminary insight into the potential structure of ALK1EC and into the structural effects of HHT2 associated mutations, which can be useful to predict the potential effect of each single mutation, to devise new biological experiments and to interpret the biological significance of new mutations, private mutations, or non-synonymous polymorphisms. PMID:22028876

  3. PPARδ Activation Acts Cooperatively with 3-Phosphoinositide-Dependent Protein Kinase-1 to Enhance Mammary Tumorigenesis

    PubMed Central

    Pollock, Claire B.; Yin, Yuzhi; Yuan, Hongyan; Zeng, Xiao; King, Sruthi; Li, Xin; Kopelovich, Levy; Albanese, Chris; Glazer, Robert I.

    2011-01-01

    Peroxisome proliferator-activated receptorδ (PPARδ) is a transcription factor that is associated with metabolic gene regulation and inflammation. It has been implicated in tumor promotion and in the regulation of 3-phosphoinositide-dependent kinase-1 (PDK1). PDK1 is a key regulator of the AGC protein kinase family, which includes the proto-oncogene AKT/PKB implicated in several malignancies, including breast cancer. To assess the role of PDK1 in mammary tumorigenesis and its interaction with PPARδ, transgenic mice were generated in which PDK1 was expressed in mammary epithelium under the control of the MMTV enhancer/promoter region. Transgene expression increased pT308AKT and pS9GSK3β, but did not alter phosphorylation of mTOR, 4EBP1, ribosomal protein S6 and PKCα. The transgenic mammary gland also expressed higher levels of PPARδ and a gene expression profile resembling wild-type mice maintained on a diet containing the PPARδ agonist, GW501516. Both wild-type and transgenic mice treated with GW501516 exhibited accelerated rates of tumor formation that were more pronounced in transgenic animals. GW501516 treatment was accompanied by a distinct metabolic gene expression and metabolomic signature that was not present in untreated animals. GW501516-treated transgenic mice expressed higher levels of fatty acid and phospholipid metabolites than treated wild-type mice, suggesting the involvement of PDK1 in enhancing PPARδ-driven energy metabolism. These results reveal that PPARδ activation elicits a distinct metabolic and metabolomic profile in tumors that is in part related to PDK1 and AKT signaling. PMID:21297860

  4. Ribosomal protein S6 kinase 1 signaling in prefrontal cortex controls depressive behavior

    PubMed Central

    Dwyer, Jason M.; Maldonado-Avilés, Jaime G.; Lepack, Ashley E.; DiLeone, Ralph J.; Duman, Ronald S.

    2015-01-01

    Current treatments for major depressive disorder (MDD) have a time lag and are ineffective for a large number of patients. Development of novel pharmacological therapies requires a comprehensive understanding of the molecular events that contribute to MDD pathophysiology. Recent evidence points toward aberrant activity of synaptic proteins as a critical contributing factor. In the present studies, we used viral-mediated gene transfer to target a key mediator of activity-dependent synaptic protein synthesis downstream of mechanistic target of rapamycin complex 1 (mTORC1) known as p70 S6 kinase 1 (S6K1). Targeted delivery of two mutants of S6K1, constitutively active or dominant-negative, to the medial prefrontal cortex (mPFC) of rats allowed control of the mTORC1/S6K1 translational pathway. Our results demonstrate that increased expression of S6K1 in the mPFC produces antidepressant effects in the forced swim test without altering locomotor activity. Moreover, expression of active S6K1 in the mPFC blocked the anhedonia caused by chronic stress, resulting in a state of stress resilience. This antidepressant response was associated with increased neuronal complexity caused by enhanced S6K1 activity. Conversely, expression of dominant-negative S6K1 in the mPFC resulted in prodepressive behavior in the forced swim test and was sufficient to cause anhedonia in the absence of chronic stress exposure. Together, these data demonstrate a critical role for S6K1 activity in depressive behaviors, and suggest that pathways downstream of mTORC1 may underlie the pathophysiology and treatment of MDD. PMID:25918363

  5. Metabolic labeling of leucine rich repeat kinases 1 and 2 with radioactive phosphate.

    PubMed

    Taymans, Jean-Marc; Gao, Fangye; Baekelandt, Veerle

    2013-01-01

    Leucine rich repeat kinases 1 and 2 (LRRK1 and LRRK2) are paralogs which share a similar domain organization, including a serine-threonine kinase domain, a Ras of complex proteins domain (ROC), a C-terminal of ROC domain (COR), and leucine-rich and ankyrin-like repeats at the N-terminus. The precise cellular roles of LRRK1 and LRRK2 have yet to be elucidated, however LRRK1 has been implicated in tyrosine kinase receptor signaling, while LRRK2 is implicated in the pathogenesis of Parkinson's disease. In this report, we present a protocol to label the LRRK1 and LRRK2 proteins in cells with (32)P orthophosphate, thereby providing a means to measure the overall phosphorylation levels of these 2 proteins in cells. In brief, affinity tagged LRRK proteins are expressed in HEK293T cells which are exposed to medium containing (32)P-orthophosphate. The (32)P-orthophosphate is assimilated by the cells after only a few hours of incubation and all molecules in the cell containing phosphates are thereby radioactively labeled. Via the affinity tag (3xflag) the LRRK proteins are isolated from other cellular components by immunoprecipitation. Immunoprecipitates are then separated via SDS-PAGE, blotted to PVDF membranes and analysis of the incorporated phosphates is performed by autoradiography ((32)P signal) and western detection (protein signal) of the proteins on the blots. The protocol can readily be adapted to monitor phosphorylation of any other protein that can be expressed in cells and isolated by immunoprecipitation. PMID:24084685

  6. Disrupting Cyclin Dependent Kinase 1 in Spermatocytes Causes Late Meiotic Arrest and Infertility in Mice.

    PubMed

    Clement, Tracy M; Inselman, Amy L; Goulding, Eugenia H; Willis, William D; Eddy, Edward M

    2015-12-01

    While cyclin dependent kinase 1 (CDK1) has a critical role in controlling resumption of meiosis in oocytes, its role has not been investigated directly in spermatocytes. Unique aspects of male meiosis led us to hypothesize that its role is different in male meiosis than in female meiosis. We generated a conditional knockout (cKO) of the Cdk1 gene in mouse spermatocytes to test this hypothesis. We found that CDK1-null spermatocytes undergo synapsis, chiasmata formation, and desynapsis as is seen in oocytes. Additionally, CDK1-null spermatocytes relocalize SYCP3 to centromeric foci, express H3pSer10, and initiate chromosome condensation. However, CDK1-null spermatocytes fail to form condensed bivalent chromosomes in prophase of meiosis I and instead are arrested at prometaphase. Thus, CDK1 has an essential role in male meiosis that is consistent with what is known about the role of CDK1 in female meiosis, where it is required for formation of condensed bivalent metaphase chromosomes and progression to the first meiotic division. We found that cKO spermatocytes formed fully condensed bivalent chromosomes in the presence of okadaic acid, suggesting that cKO chromosomes are competent to condense, although they do not do so in vivo. Additionally, arrested cKO spermatocytes exhibited irregular cell shape, irregular large nuclei, and large distinctive nucleoli. These cells persist in the seminiferous epithelium through the next seminiferous epithelial cycle with a lack of stage XII checkpoint-associated cell death. This indicates that CDK1 is required upstream of a checkpoint-associated cell death as well as meiotic metaphase progression in mouse spermatocytes. PMID:26490841

  7. Structural and energetic basis of protein kinetic destabilization in human phosphoglycerate kinase 1 deficiency.

    PubMed

    Pey, Angel L; Mesa-Torres, Noel; Chiarelli, Laurent R; Valentini, Giovanna

    2013-02-19

    Protein kinetic destabilization is a common feature of many human genetic diseases. Human phosphoglycerate kinase 1 (PGK1) deficiency is a rare genetic disease caused by mutations in the PGK1 protein, which often shows reduced kinetic stability. In this work, we have performed an in-depth characterization of the thermal stability of the wild type and four disease-causing mutants (I47N, L89P, E252A, and T378P) of human PGK1. PGK1 thermal denaturation is a process under kinetic control, and it is described well by a two-state irreversible denaturation model. Kinetic analysis of differential scanning calorimetry profiles shows that the disease-causing mutations decrease PGK1 kinetic stability from ~5-fold (E252A) to ~100000-fold (L89P) compared to that of wild-type PGK1, and in some cases, mutant enzymes are denatured on a time scale of a few minutes at physiological temperature. We show that changes in protein kinetic stability are associated with large differences in enthalpic and entropic contributions to denaturation free energy barriers. It is also shown that the denaturation transition state becomes more nativelike in terms of solvent exposure as the protein is destabilized by mutations (Hammond effect). Unfolding experiments with urea further suggest a scenario in which the thermodynamic stability of PGK1 at least partly determines its kinetic stability. ATP and ADP kinetically stabilize PGK1 enzymes, and kinetic stabilization is nucleotide- and mutant-selective. Overall, our data provide insight into the structural and energetic basis underlying the low kinetic stability displayed by some mutants causing human PGK1 deficiency, which may have important implications for the development of native state kinetic stabilizers for the treatment of this disease. PMID:23336698

  8. Impairment of Angiogenic Sphingosine Kinase-1/Sphingosine-1-Phosphate Receptors Pathway in Preeclampsia.

    PubMed

    Dobierzewska, Aneta; Palominos, Macarena; Sanchez, Marianela; Dyhr, Michael; Helgert, Katja; Venegas-Araneda, Pia; Tong, Stephen; Illanes, Sebastian E

    2016-01-01

    Preeclampsia (PE), is a serious pregnancy disorder characterized in the early gestation by shallow trophoblast invasion, impaired placental neo-angiogenesis, placental hypoxia and ischemia, which leads to maternal and fetal morbidity and mortality. Here we hypothesized that angiogenic sphingosine kinase-1 (SPHK1)/sphingosine-1-phosphate (S1P) receptors pathway is impaired in PE. We found that SPHK1 mRNA and protein expression are down-regulated in term placentae and term chorionic villous explants from patients with PE or severe PE (PES), compared with controls. Moreover, mRNA expression of angiogenic S1PR1 and S1PR3 receptors were decreased in placental samples of PE and PES patients, whereas anti-angiogenic S1PR2 was up-regulated in chorionic villous tissue of PES subjects, pointing to its potential atherogenic and inflammatory properties. Furthermore, in in vitro (JAR cells) and ex vivo (chorionic villous explants) models of placental hypoxia, SPHK1 mRNA and protein were strongly up-regulated under low oxygen tension (1% 02). In contrast, there was no change in SPHK1 expression under the conditions of placental physiological hypoxia (8% 02). In both models, nuclear protein levels of HIF1A were increased at 1% 02 during the time course, but there was no up-regulation at 8% 02, suggesting that SPHK1 and HIF1A might be the part of the same canonical pathway during hypoxia and that both contribute to placental neovascularization during early gestation. Taken together, this study suggest the SPHK1 pathway may play a role in the human early placentation process and may be involved in the pathogenesis of PE. PMID:27284992

  9. Polo-like kinase 1 inhibition sensitizes neuroblastoma cells for vinca alkaloid-induced apoptosis.

    PubMed

    Czaplinski, Sebastian; Hugle, Manuela; Stiehl, Valerie; Fulda, Simone

    2016-02-23

    High polo-like kinase 1 (PLK1) expression has been linked to poor outcome in neuroblastoma (NB), indicating that it represents a relevant therapeutic target in this malignancy. Here, we identify a synergistic induction of apoptosis by the PLK1 inhibitor BI 2536 and vinca alkaloids in NB cells. Synergistic drug interaction of BI 2536 together with vincristine (VCR), vinblastine (VBL) or vinorelbine (VNR) is confirmed by calculation of combination index (CI). Also, BI 2536 and VCR act in concert to reduce long-term clonogenic survival. Importantly, BI 2536 significantly enhances the antitumor activity of VCR in an in vivo model of NB. Mechanistically, BI 2536/VCR co-treatment triggers prolonged mitotic arrest, which is necessary for BI 2536/VCR-mediated apoptosis, since pharmacological inhibition of mitotic arrest by the CDK1 inhibitor RO-3306 significantly reduces cell death. Prolonged mitotic arrest leads to phosphorylation-mediated inactivation of BCL-2 and BCL-XL as well as downregulation of MCL-1, since inhibition of mitotic arrest by RO-3306 also prevents phosphorylation of BCL-2 and BCL-XL and MCL-1 downregulation. This inactivation of antiapoptotic BCL-2 proteins promotes activation of BAX and BAK, cleavage of caspase-9 and -3 and caspase-dependent apoptosis. Engagement of the mitochondrial pathway of apoptosis is critically required for BI 2536/VCR-induced apoptosis, since ectopic expression of a non-degradable MCL-1 phospho-mutant, BCL-2 overexpression or BAK knockdown significantly reduce BI 2536/VCR-mediated apoptosis. Thus, PLK1 inhibitors may open new perspectives for chemosensitization of NB. PMID:26046302

  10. Luteolin Suppresses Cancer Cell Proliferation by Targeting Vaccinia-Related Kinase 1

    PubMed Central

    Shin, Joon; Harikishore, Amaravadhi; Lim, Jong-Kwan; Jung, Youngseob; Lyu, Ha-Na; Baek, Nam-In; Choi, Kwan Yong; Yoon, Ho Sup; Kim, Kyong-Tai

    2014-01-01

    Uncontrolled proliferation, a major feature of cancer cells, is often triggered by the malfunction of cell cycle regulators such as protein kinases. Recently, cell cycle-related protein kinases have become attractive targets for anti-cancer therapy, because they play fundamental roles in cellular proliferation. However, the protein kinase-targeted drugs that have been developed so far do not show impressive clinical results and also display severe side effects; therefore, there is undoubtedly a need to investigate new drugs targeting other protein kinases that are critical in cell cycle progression. Vaccinia-related kinase 1 (VRK1) is a mitotic kinase that functions in cell cycle regulation by phosphorylating cell cycle-related substrates such as barrier-to-autointegration factor (BAF), histone H3, and the cAMP response element (CRE)-binding protein (CREB). In our study, we identified luteolin as the inhibitor of VRK1 by screening a small-molecule natural compound library. Here, we evaluated the efficacy of luteolin as a VRK1-targeted inhibitor for developing an effective anti-cancer strategy. We confirmed that luteolin significantly reduces VRK1-mediated phosphorylation of the cell cycle-related substrates BAF and histone H3, and directly interacts with the catalytic domain of VRK1. In addition, luteolin regulates cell cycle progression by modulating VRK1 activity, leading to the suppression of cancer cell proliferation and the induction of apoptosis. Therefore, our study suggests that luteolin-induced VRK1 inhibition may contribute to establish a novel cell cycle-targeted strategy for anti-cancer therapy. PMID:25310002

  11. Plant thymidine kinase 1: a novel efficient suicide gene for malignant glioma therapy.

    PubMed

    Khan, Zahidul; Knecht, Wolfgang; Willer, Mette; Rozpedowska, Elzbieta; Kristoffersen, Peter; Clausen, Anders Ranegaard; Munch-Petersen, Birgitte; Almqvist, Per M; Gojkovic, Zoran; Piskur, Jure; Ekström, Tomas J

    2010-06-01

    The prognosis for malignant gliomas remains poor, and new treatments are urgently needed. Targeted suicide gene therapy exploits the enzymatic conversion of a prodrug, such as a nucleoside analog, into a cytotoxic compound. Although this therapeutic strategy has been considered a promising regimen for central nervous system (CNS) tumors, several obstacles have been encountered such as inefficient gene transfer to the tumor cells, limited prodrug penetration into the CNS, and inefficient enzymatic activity of the suicide gene. We report here the cloning and successful application of a novel thymidine kinase 1 (TK1) from the tomato plant, with favorable characteristics in vitro and in vivo. This enzyme (toTK1) is highly specific for the nucleoside analog prodrug zidovudine (azidothymidine, AZT), which is known to penetrate the blood-brain barrier. An important feature of toTK1 is that it efficiently phosphorylates its substrate AZT not only to AZT monophosphate, but also to AZT diphosphate, with excellent kinetics. The efficiency of the toTK1/AZT system was confirmed when toTK1-transduced human glioblastoma (GBM) cells displayed a 500-fold increased sensitivity to AZT compared with wild-type cells. In addition, when neural progenitor cells were used as delivery vectors for toTK1 in intracranial GBM xenografts in nude rats, substantial attenuation of tumor growth was achieved in animals exposed to AZT, and survival of the animals was significantly improved compared with controls. The novel toTK1/AZT suicide gene therapy system in combination with stem cell-mediated gene delivery promises new treatment of malignant gliomas. PMID:20154339

  12. Targeting colorectal cancer cells by a novel sphingosine kinase 1 inhibitor PF-543.

    PubMed

    Ju, TongFa; Gao, DaQuan; Fang, Zheng-yu

    2016-02-12

    In this study, we showed that PF-543, a novel sphingosine kinase 1 (SphK1) inhibitor, exerted potent anti-proliferative and cytotoxic effects against a panel of established (HCT-116, HT-29 and DLD-1) and primary human colorectal cancer (CRC) cells. Its sensitivity was negatively associated with SphK1 expression level in the CRC cells. Surprisingly, PF-543 mainly induced programmed necrosis, but not apoptosis, in the CRC cells. CRC cell necrotic death was detected by lactate dehydrogenase (LDH) release, mitochondrial membrane potential (MMP) collapse and mitochondrial P53-cyclophilin-D (Cyp-D) complexation. Correspondingly, the necrosis inhibitor necrostatin-1 largely attenuated PF-543-induced cytotoxicity against CRC cells. Meanwhile, the Cyp-D inhibitors (sanglifehrin A and cyclosporin A), or shRNA-mediated knockdown of Cyp-D, remarkably alleviated PF-543-induced CRC cell necrotic death. Reversely, over-expression of wild-type Cyp-D in HCT-116 cells significantly increased PF-543's sensitivity. In vivo, PF-543 intravenous injection significantly suppressed HCT-116 xenograft growth in severe combined immunodeficient (SCID) mice, whiling remarkably improving the mice survival. The in vivo activity by PF-543 was largely attenuated when combined with the Cyp-D inhibitor cyclosporin A. Collectively, our results demonstrate that PF-543 exerts potent anti-CRC activity in vitro and in vivo. Mitochondrial programmed necrosis pathway is likely the key mechanism responsible for PF-543's actions in CRC cells. PMID:26775841

  13. Effect of selenite on T-cell mitogenesis: contribution of ROS production and apoptosis signal-regulating kinase 1.

    PubMed

    Ueno, Hitoshi; Kajihara, Hitomi; Nakamura, Hajime; Okuno, Tomofumi; Sakazaki, Fumitoshi; Arakawa, Tomohiro; Ogino, Hirofumi; Nakamuro, Katsuhiko; Yodoi, Junji

    2014-01-01

    Although supplementation with the selenocompound, sodium selenite has been shown to stimulate the concanavalin A-induced T-cell mitogenic response, the mechanisms responsible remain unclear. This study was conducted to evaluate the relationships between the induction of apoptosis, formation of tumor necrosis factor (TNF)-alpha and reactive oxygen species (ROS), activation of apoptosis signal-regulating kinase (ASK) 1 and the thioredoxin (Trx) system when mitogenesis was stimulated by selenite. TNF-alpha was dose-dependently released by mouse splenocytes treated with selenite, and apoptosis was induced when TNF-alpha was added at the indicated concentrations. However, supplementation with selenite at low concentrations inhibited the accumulation of ROS with the increased expression of Trx reductase 1 and induction of apoptosis in wild-type splenocytes, and also at high concentrations in Trx-1-transgenic mouse splenocytes. The suppression of apoptosis was accompanied by a decrease in the expression of phospho-ASK1. These results suggest that the stimulation of T-cell mitogenesis by selenite may be partly attributed to the inhibited accumulation of ROS due to a reduced Trx-1/TR1 system, the inactivation of ASK1, and the suppression of apoptosis. PMID:25087957

  14. Rescue Medicine for Epilepsy in Education Settings.

    PubMed

    Hartman, Adam L; Devore, Cynthia Di Laura; Doerrer, Sarah C

    2016-01-01

    Children and adolescents with epilepsy may experience prolonged seizures in school-associated settings (eg, during transportation, in the classroom, or during sports activities). Prolonged seizures may evolve into status epilepticus. Administering a seizure rescue medication can abort the seizure and may obviate the need for emergency medical services and subsequent care in an emergency department. In turn, this may save patients from the morbidity of more invasive interventions and the cost of escalated care. There are significant variations in prescribing practices for seizure rescue medications, partly because of inconsistencies between jurisdictions in legislation and professional practice guidelines among potential first responders (including school staff). There also are potential liability issues for prescribers, school districts, and unlicensed assistive personnel who might administer the seizure rescue medications. This clinical report highlights issues that providers may consider when prescribing seizure rescue medications and creating school medical orders and/or action plans for students with epilepsy. Collaboration among prescribing providers, families, and schools may be useful in developing plans for the use of seizure rescue medications. PMID:26712862

  15. Amorphous silicon bolometer for fire/rescue

    NASA Astrophysics Data System (ADS)

    Francisco, Glenn L.

    2001-03-01

    Thermal imaging sensors have completely changed the way the world views fire and rescue applications. Recently, in the uncooled infrared camera and microbolometer detector areas, major strides have been made in manufacturing personal fire and rescue sensors. A family of new amorphous silicon microbolometers are being produced utilizing low cost, low weight, ultra low power, small size, high volume vacuum packaged silicon wafer-level focal plane array technologies. These bolometers contain no choppers or thermoelectric coolers, require no manual calibration and use readily available commercial off-the-shelf components. Manufacturing and packaging discoveries have allowed infrared sensitive silicon arrays to be produced with the same methods that have driven the rapidly advancing digital wireless telecommunications industries. Fire and rescue professionals are now able to conduct minimum time thermal imaging penetration, surveillance, detection, recognition, rescue and egress while maintaining situational awareness in a manner consistent with the modern technological applications. The purpose of this paper is to describe an uncooled micro bolometer infrared camera approach for meeting fire/rescue wants, needs and requirements, with application of recent technology advancements. This paper also details advances in bolometric focal plane arrays, optical and circuit card technologies, while providing a glimpse into the future of micro sensor growth. Technical barriers are addressed in light of constraints and lessons learned around this technology.

  16. Different approaches of teaching in chemistry. Membrane targeting mechanism of human sphingosine kinase 1

    NASA Astrophysics Data System (ADS)

    Hwang, Jeong-Hye

    Part 1. Biochemistry research involves elucidating the mechanism of membrane targeting of human sphingosine kinase 1 (hSK1). Sphingosine kinase (SK) is an enzyme that catalyzes phosphorylation of sphingosine to sphingosine-1-phosphate (S-1-P). hSK1 can be activated by its agonists resulting in rapid and transient increased production of S-1-P, resulting in enhancement of apoptosis. Upon activation by PMA, SK translocates to the plasma membrane. In vitro measurement demonstrated hSK1 selectively bound phosphatidylserine over anionic lipids and showed strong preference for the plasma membrane-mimetics. Mutational analysis of conserved Thr54 and Asn89 on putative membrane-binding surface from the model structure showed both in vivo and in vitro that these two residues are important for the membrane selectivity of hSK1. Part 2. Chemical education research focuses on three different ways of scientific learning and teaching. First, inquiry teaching that involves a writing method called the Science Writing Heuristic (SWH) is analyzed using grounded theory. This is done in a general education course for pre-service teachers. As a result, (1) students experience understanding of concept through mastery of their own methods and experience different aspects of inquiry processes; (2) SWH method allows instructors to detect misconceptions generated by students' incorrect, but logical interpretations of their data, and helps instructors to make changes to guide students; (3) as students experience meta-cognition, students gain understanding of concepts by relating mathematical progression to different parts of the experiments and also by applying what they learn into other situations. Second, statistical analysis on the long term effects of a combined math/chemistry program is analyzed through multiple linear regression and discriminant function analysis. The results demonstrate the program was beneficial to the underrepresented students when the college success was measured

  17. Sphingosine Kinase-1 Is Central to Androgen-Regulated Prostate Cancer Growth and Survival

    PubMed Central

    Dayon, Audrey; Brizuela, Leyre; Martin, Claire; Mazerolles, Catherine; Pirot, Nelly; Doumerc, Nicolas; Nogueira, Leonor; Golzio, Muriel; Teissié, Justin; Serre, Guy; Rischmann, Pascal; Malavaud, Bernard; Cuvillier, Olivier

    2009-01-01

    Background Sphingosine kinase-1 (SphK1) is an oncogenic lipid kinase notably involved in response to anticancer therapies in prostate cancer. Androgens regulate prostate cancer cell proliferation, and androgen deprivation therapy is the standard of care in the management of patients with advanced disease. Here, we explored the role of SphK1 in the regulation of androgen-dependent prostate cancer cell growth and survival. Methodology/Principal Findings Short-term androgen removal induced a rapid and transient SphK1 inhibition associated with a reduced cell growth in vitro and in vivo, an event that was not observed in the hormono-insensitive PC-3 cells. Supporting the critical role of SphK1 inhibition in the rapid effect of androgen depletion, its overexpression could impair the cell growth decrease. Similarly, the addition of dihydrotestosterone (DHT) to androgen-deprived LNCaP cells re-established cell proliferation, through an androgen receptor/PI3K/Akt dependent stimulation of SphK1, and inhibition of SphK1 could markedly impede the effects of DHT. Conversely, long-term removal of androgen support in LNCaP and C4-2B cells resulted in a progressive increase in SphK1 expression and activity throughout the progression to androgen-independence state, which was characterized by the acquisition of a neuroendocrine (NE)-like cell phenotype. Importantly, inhibition of the PI3K/Akt pathway—by negatively impacting SphK1 activity—could prevent NE differentiation in both cell models, an event that could be mimicked by SphK1 inhibitors. Fascinatingly, the reversability of the NE phenotype by exposure to normal medium was linked with a pronounced inhibition of SphK1 activity. Conclusions/Significance We report the first evidence that androgen deprivation induces a differential effect on SphK1 activity in hormone-sensitive prostate cancer cell models. These results also suggest that SphK1 activation upon chronic androgen deprivation may serve as a compensatory mechanism

  18. Eribulin synergizes with Polo-like kinase 1 inhibitors to induce apoptosis in rhabdomyosarcoma.

    PubMed

    Stehle, Angelika; Hugle, Manuela; Fulda, Simone

    2015-08-28

    Eribulin, a novel microtubule-interfering drug, was recently shown to exhibit high antitumor activity in vivo against various pediatric cancers. Here, we identify a novel synthetic lethal interaction of Eribulin together with Polo-like kinase 1 (PLK1) inhibitors against rhabdomyosarcoma (RMS) in vitro and in vivo. Eribulin and the PLK1 inhibitor BI 2536 at subtoxic concentrations synergize to induce apoptosis in RMS cells as confirmed by calculation of combination index (CI). Also, Eribulin/BI 2536 co-treatment is significantly more effective than monotherapy to reduce cell viability and inhibit colony formation of RMS cells. Similarly, Eribulin and BI 2536 act in concert to trigger apoptosis in a primary, patient-derived ARMS culture, underscoring the clinical relevance of this combination. Importantly, Eribulin and BI 2536 cooperate to suppress tumor growth in an in vivo model of RMS. On molecular grounds, Eribulin/BI 2536 co-treatment causes profound mitotic arrest, which is critically required for synergism, since inhibition of mitotic arrest by CDK1 inhibitor RO-3306 abolishes Eribulin/BI 2536-mediated apoptosis. Eribulin and BI 2536 cooperate to activate caspase-9, -3 and -8, which is necessary for apoptosis induction, since the broad-range caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD.fmk) reduces Eribulin/BI 2536-induced apoptosis significantly, yet partially. Intriguingly, knockdown of endonuclease G (ENDOG) also significantly inhibits Eribulin/BI 2536-triggered apoptosis, demonstrating the involvement of both caspase-dependent and -independent effector pathways. Synergistic induction of apoptosis is similarly found for Eribulin/BI 2536 co-treatment in neuroblastoma cells and for the combination of vincristine (another antimicrotubule chemotherapeutic) with Poloxin (another PLK1 inhibitor), thus pointing to a broader significance of this concomitant microtubule- and PLK1-targeting strategy for pediatric oncology. In

  19. Atypical Regulation of a Green Lineage-Specific B-Type Cyclin-Dependent Kinase1

    PubMed Central

    Corellou, Florence; Camasses, Alain; Ligat, Laetitia; Peaucellier, Gérard; Bouget, François-Yves

    2005-01-01

    Cyclin-dependent kinases (CDKs) are the main regulators of cell cycle progression in eukaryotes. The role and regulation of canonical CDKs, such as the yeast (Saccharomyces cerevisiae) Cdc2 or plant CDKA, have been extensively characterized. However, the function of the plant-specific CDKB is not as well understood. Besides being involved in cell cycle control, Arabidopsis (Arabidopsis thaliana) CDKB would integrate developmental processes to cell cycle progression. We investigated the role of CDKB in Ostreococcus (Ostreococcus tauri), a unicellular green algae with a minimal set of cell cycle genes. In this primitive alga, at the basis of the green lineage, CDKB has integrated two levels of regulations: It is regulated by Tyr phosphorylation like cdc2/CDKA and at the level of synthesis-like B-type CDKs. Furthermore, Ostreococcus CDKB/cyclin B accounts for the main peak of mitotic activity, and CDKB is able to rescue a yeast cdc28ts mutant. By contrast, Ostreococcus CDKA is not regulated by Tyr phosphorylation, and it exhibits a low and steady-state activity from DNA replication to exit of mitosis. This suggests that from a major role in the control of mitosis in green algae, CDKB has evolved in higher plants to assume other functions outside the cell cycle. PMID:15965018

  20. Suppression of Chemically Induced and Spontaneous Mouse Oocyte Activation by AMP-Activated Protein Kinase1

    PubMed Central

    Ya, Ru; Downs, Stephen M.

    2013-01-01

    ABSTRACT Oocyte activation is an important process triggered by fertilization that initiates embryonic development. However, parthenogenetic activation can occur either spontaneously or with chemical treatments. The LT/Sv mouse strain is genetically predisposed to spontaneous activation. LT oocytes have a cell cycle defect and are ovulated at the metaphase I stage instead of metaphase II. A thorough understanding of the female meiosis defects in this strain remains elusive. We have reported that AMP-activated protein kinase (PRKA) has an important role in stimulating meiotic resumption and promoting completion of meiosis I while suppressing premature parthenogenetic activation. Here we show that early activation of PRKA during the oocyte maturation period blocked chemically induced activation in B6SJL oocytes and spontaneous activation in LT/SvEiJ oocytes. This inhibitory effect was associated with high levels of MAPK1/3 activity. Furthermore, stimulation of PRKA partially rescued the meiotic defects of LT/Sv mouse oocytes in concert with correction of abnormal spindle pole localization of PRKA and loss of prolonged spindle assembly checkpoint activity. Altogether, these results confirm a role for PRKA in helping sustain the MII arrest in mature oocytes and suggest that dysfunctional PRKA contributes to meiotic defects in LT/SvEiJ oocytes. PMID:23390161

  1. Checkpoint Kinase 1 Activation Enhances Intestinal Epithelial Barrier Function via Regulation of Claudin-5 Expression

    PubMed Central

    Watari, Akihiro; Hasegawa, Maki; Yagi, Kiyohito; Kondoh, Masuo

    2016-01-01

    Several stressors are known to influence epithelial tight junction (TJ) integrity, but the association between DNA damage and TJ integrity remains unclear. Here we examined the effects of daunorubicin and rebeccamycin, two anti-tumor chemicals that induce DNA damage, on TJ integrity in human intestinal epithelial cells. Daunorubicin and rebeccamycin dose-dependently enhanced transepithelial electrical resistance (TER) and decreased flux of the 4 kDa FITC-dextran in Caco-2 cell monolayer. Daunorubicin- or rebeccamycin-induced enhancement of the TJ barrier function partly rescued attenuation of the barrier function by the inflammatory cytokines TNF-α and IFN-γ. Daunorubicin and rebeccamycin increased claudin-5 expression and the product was distributed in the actin cytoskeleton fraction, which was enriched with TJ proteins. Caffeine, which is an inhibitor of ataxia telangiectasia mutated protein (ATM) and ataxia telangiectasia mutated and Rad3-related protein (ATR), and the Chk1 inhibitor inhibited the TER increases induced by daunorubicin and rebeccamycin, whereas a Chk2 inhibitor did not. Treatment with Chk1 siRNA also significantly inhibited the TER increases. Induction of claudin-5 expression was inhibited by Chk1 inhibitor and by siRNA treatment. Our results suggest that Chk1 activation by daunorubicin and rebeccamycin induced claudin-5 expression and enhanced TJ barrier function in Caco-2 cell monolayer, which suggests a link between DNA damage and TJ integrity in the human intestine. PMID:26727128

  2. [Air rescue: current significance and practical issues].

    PubMed

    Schellhaaß, A; Popp, E

    2014-12-01

    Germany has a nationwide and powerful helicopter emergency medical services system (HEMS), which executes primary rescue missions and interhospital transfer of intensive care patients. In recent years the range of HEMS missions has become modified due to demographic changes and structural changes in the healthcare system. Furthermore, the number of HEMS missions is steadily increasing. If reasonably used air rescue contributes to desired reductions in overall preclinical time. Moreover, it facilitates prompt transport of patients to a hospital suitable for definitive medical care and treatment can be initiated earlier which is a particular advantage for severely injured and critically ill patients. Because of complex challenges during air rescue missions the qualifications of the HEMS personnel have to be considerably higher in comparison with ground based emergency medical services. PMID:25430664

  3. RESCU: A real space electronic structure method

    NASA Astrophysics Data System (ADS)

    Michaud-Rioux, Vincent; Zhang, Lei; Guo, Hong

    2016-02-01

    In this work we present RESCU, a powerful MATLAB-based Kohn-Sham density functional theory (KS-DFT) solver. We demonstrate that RESCU can compute the electronic structure properties of systems comprising many thousands of atoms using modest computer resources, e.g. 16 to 256 cores. Its computational efficiency is achieved from exploiting four routes. First, we use numerical atomic orbital (NAO) techniques to efficiently generate a good quality initial subspace which is crucially required by Chebyshev filtering methods. Second, we exploit the fact that only a subspace spanning the occupied Kohn-Sham states is required, and solving accurately the KS equation using eigensolvers can generally be avoided. Third, by judiciously analyzing and optimizing various parts of the procedure in RESCU, we delay the O (N3) scaling to large N, and our tests show that RESCU scales consistently as O (N2.3) from a few hundred atoms to more than 5000 atoms when using a real space grid discretization. The scaling is better or comparable in a NAO basis up to the 14,000 atoms level. Fourth, we exploit various numerical algorithms and, in particular, we introduce a partial Rayleigh-Ritz algorithm to achieve efficiency gains for systems comprising more than 10,000 electrons. We demonstrate the power of RESCU in solving KS-DFT problems using many examples running on 16, 64 and/or 256 cores: a 5832 Si atoms supercell; a 8788 Al atoms supercell; a 5324 Cu atoms supercell and a small DNA molecule submerged in 1713 water molecules for a total 5399 atoms. The KS-DFT is entirely converged in a few hours in all cases. Our results suggest that the RESCU method has reached a milestone of solving thousands of atoms by KS-DFT on a modest computer cluster.

  4. Spatiotemporal rescue of memory dysfunction in Drosophila.

    PubMed

    McGuire, Sean E; Le, Phuong T; Osborn, Alexander J; Matsumoto, Kunihiro; Davis, Ronald L

    2003-12-01

    We have developed a method for temporal and regional gene expression targeting (TARGET) in Drosophila and show the simultaneous spatial and temporal rescue of a memory defect. The transient expression of the rutabaga-encoded adenylyl cyclase in the mushroom bodies of the adult brain was necessary and sufficient to rescue the rutabaga memory deficit, which rules out a developmental brain defect in the etiology of this deficit and demonstrates an acute role for rutabaga in memory formation in these neurons. The TARGET system offers general utility in simultaneously addressing issues of when and where gene products are required. PMID:14657498

  5. p90 ribosomal S6 kinase 1 (RSK1) isoenzyme specifically regulates cytokinesis progression.

    PubMed

    Nam, Hyun-Ja; Lee, In Jeong; Jang, Seunghoon; Bae, Chang-Dae; Kwak, Sahng-June; Lee, Jae-Ho

    2014-02-01

    The p90 ribosomal S6 kinase family (RSK1-4) of Ser/Thr kinases is a downstream component of the Ras-MAPK cascade responsible for regulating various cellular processes. Here, we examined the potential involvement of RSKs in regulating mitosis by transfecting HeLa cells with siRNAs targeting RSK1 and -2, which are the major isoforms. Depletion of RSK1 but not RSK2 triggered a significant accumulation of binucleated cells compared to control cells (0.5% vs. 10.5%, respectively); this was rescued by expression of exogenous RSK1 but not a kinase-defective mutant. Monitoring of cell division by time-lapse imaging revealed that the observed binucleation mainly stemmed from a failure to form and ingress the cleavage furrow during early cytokinesis. Immunocytochemical analysis of RhoA and anillin, the two principal regulators of cleavage furrow formation and ingression, showed that these proteins were abnormally localized during anaphase in RSK1-depleted cells. Furthermore, RSK1-depleted cells seemed to have impairments in midzone microtubule formation, as suggested by morphological changes and lengthening of the midzone (15.2 ± 1.7 μm vs. 17.4 ± 1.7 μm in control cells). We also observed shortening of the pole-to-polar-cortex distance in RSK1-depleted cells (4.30 ± 1.37 μm vs. 2.80 ± 0.84 μm in control cells) and scanty distribution of microtubules at the periphery of the equatorial region during anaphase, suggesting an aberrant distribution of astral microtubules. Taken together, these results suggest that RSK1 is specifically required for cleavage furrow formation and ingression during cytokinesis. This may occur via the involvement of RSK1 in proper midzone and astral microtubule structure formation during anaphase, which is essential for the correct localization of anillin and RhoA. PMID:24269382

  6. Kinetic Mechanism and Rate-Limiting Steps of Focal Adhesion Kinase-1

    SciTech Connect

    Schneck, Jessica L.; Briand, Jacques; Chen, Stephanie; Lehr, Ruth; McDevitt, Patrick; Zhao, Baoguang; Smallwood, Angela; Concha, Nestor; Oza, Khyati; Kirkpatrick, Robert; Yan, Kang; Villa, James P.; Meek, Thomas D.; Thrall, Sara H.

    2010-12-07

    Steady-state kinetic analysis of focal adhesion kinase-1 (FAK1) was performed using radiometric measurement of phosphorylation of a synthetic peptide substrate (Ac-RRRRRRSETDDYAEIID-NH{sub 2}, FAK-tide) which corresponds to the sequence of an autophosphorylation site in FAK1. Initial velocity studies were consistent with a sequential kinetic mechanism, for which apparent kinetic values k{sub cat} (0.052 {+-} 0.001 s{sup -1}), K{sub MgATP} (1.2 {+-} 0.1 {micro}M), K{sub iMgATP} (1.3 {+-} 0.2 {micro}M), K{sub FAK-tide} (5.6 {+-} 0.4 {micro}M), and K{sub iFAK-tide} (6.1 {+-} 1.1 {micro}M) were obtained. Product and dead-end inhibition data indicated that enzymatic phosphorylation of FAK-tide by FAK1 was best described by a random bi bi kinetic mechanism, for which both E-MgADP-FAK-tide and E-MgATP-P-FAK-tide dead-end complexes form. FAK1 catalyzed the {beta}{gamma}-bridge:{beta}-nonbridge positional oxygen exchange of [{gamma}-{sup 18}O{sub 4}]ATP in the presence of 1 mM [{gamma}-{sup 18}O{sub 4}]ATP and 1.5 mM FAK-tide with a progressive time course which was commensurate with catalysis, resulting in a rate of exchange to catalysis of k{sub x}/k{sub cat} = 0.14 {+-} 0.01. These results indicate that phosphoryl transfer is reversible and that a slow kinetic step follows formation of the E-MgADP-P-FAK-tide complex. Further kinetic studies performed in the presence of the microscopic viscosogen sucrose revealed that solvent viscosity had no effect on k{sub cat}/K{sub FAK-tide}, while k{sub cat} and k{sub cat}/K{sub MgATP} were both decreased linearly at increasing solvent viscosity. Crystallographic characterization of inactive versus AMP-PNP-liganded structures of FAK1 showed that a large conformational motion of the activation loop upon ATP binding may be an essential step during catalysis and would explain the viscosity effect observed on k{sub cat}/K{sub m} for MgATP but not on k{sub cat}/K{sub m} for FAK-tide. From the positional isotope exchange, viscosity, and

  7. EMERGENCY VICTIM CARE AND RESCUE, INSTRUCTOR'S MANUAL.

    ERIC Educational Resources Information Center

    MORANDO, ROCCO V.; STOVER, WILBUR F.

    DEVELOPED AT THE STATE LEVEL BY SQUADMEN AND TRADE AND INDUSTRIAL PERSONNEL, THIS MANUAL IS FOR USE BY A QUALIFIED SQUADMAN IN TEACHING FULL-TIME AND VOLUNTEER EMERGENCY AND RESCUE WORKERS IN AN EMERGENCY SQUAD STATION OR TRAINING CENTER. TEACHING GUIDES ARE PROVIDED FOR A 30-HOUR COURSE ON EMERGENCY VICTIM CARE AND A 20-HOUR COURSE ON VICTIM…

  8. Emergency Medical Rescue in a Radiation Environment

    SciTech Connect

    Briesmeister, L.; Ellington, Y.; Hollis, R.; Kunzman, J.; McNaughton, M.; Ramsey, G.; Somers, B.; Turner, A.; Finn, J.

    1999-09-14

    Previous experience with emergency medical rescues in the presence of radiation or contamination indicates that the training provided to emergency responders is not always appropriate. A new course developed at Los Alamos includes specific procedures for emergency response in a variety of radiological conditions.

  9. 78 FR 39531 - Mine Rescue Teams

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-01

    ... July 1, 2013 Part V Department of Labor Mine Safety and Health Administration 30 CFR Part 49 Mine... and Regulations#0;#0; ] DEPARTMENT OF LABOR Mine Safety and Health Administration 30 CFR Part 49 Mine... Miner Act Requirements for Underground Coal Mine Operators and Mine Rescue Teams Type of mine...

  10. Rescue Fantasies in Homicide-Suicide

    ERIC Educational Resources Information Center

    Selkin, James

    1976-01-01

    Homicide-suicides (N=13), events in which an individual murders one or more persons and kills himself immediately afterward, were studied, The results are interpreted to suggest that persons who commit homicide-suicide are acting out a three-party rescue fantasy in an attempt to resolve unbearable stress. (Author)