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Sample records for organischen stoffgruppen pak

  1. PAK family kinases

    PubMed Central

    Zhao, Zhuo-shen; Manser, Ed

    2012-01-01

    The p21-activated kinases (PAKs) are a family of Ser/Thr protein kinases that are represented by six genes in humans (PAK 1–6), and are found in all eukaryotes sequenced to date. Genetic and knockdown experiments in frogs, fish and mice indicate group I PAKs are widely expressed, required for multiple tissue development, and particularly important for immune and nervous system function in the adult. The group II PAKs (human PAKs 4–6) are more enigmatic, but their restriction to metazoans and presence at cell-cell junctions suggests these kinases emerged to regulate junctional signaling. Studies of protozoa and fungal PAKs show that they regulate cell shape and polarity through phosphorylation of multiple cytoskeletal proteins, including microtubule binding proteins, myosins and septins. This chapter discusses what we know about the regulation of PAKs and their physiological role in different model organisms, based primarily on gene knockout studies. PMID:23162738

  2. The PAKs come of age

    PubMed Central

    2012-01-01

    Protein kinases are versatile signaling molecules that are involved in the regulation most physiological responses. The p21-activated kinases (PAKs) can be activated directly by the small GTPases Rac and Cdc42 and are among the best characterized downstream effectors of these Rho proteins. The structure, substrate specificity and functional role of PAKS are evolutionarily conserved from protozoa to mammals. Vertebrate PAKs are particularly important for cytoskeletal remodeling and focal adhesion assembly, thereby contributing to dynamic processes such as cell migration and synaptic plasticity. This issue of Cellular Logistics focuses on the PAK family of kinases, with ten reviews written by researchers currently working in the field. Here in this introductory overview we highlight some of the most interesting recent discoveries regarding PAK biochemistry and biology. The reviews in this issue cover a range of topics including the atomic structures of PAK1 and PAK4, their role in animals as assessed by knockout studies, and how PAKs are likely to contribute to cancer and neurodegenerative diseases. The promise remains that PAK inhibitors will emerge that validate current pre-clinical studies suggesting that blocking PAK activity will positively contribute to human health. PMID:23125949

  3. Role of PAK6 in Prostate Cancer

    DTIC Science & Technology

    2007-04-01

    threonine-proline-tyrosine, TPY ) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was also...for MKK6- mediated activation. PAK4 and PAK5 were similarly activated by MKK6, consistent with a conserved TPY motif in their activation domains. The...tyrosine, TPY ) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was also blocked by mutation of an

  4. Role of PAK6 in Prostate Cancer

    DTIC Science & Technology

    2005-10-01

    Alternatively, center of the activation loop within the catalytic domain. Crys- the TPY motif in the group 1 PAKs may be recognized by another tallographic...tyrosine, TPY ) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was also blocked by mutation of...activation. PAK4 and PAK5 were similarly activated by MKK6, consistent with a conserved TPY motif in their activation domains. The activation of PAK6 by

  5. A tale of two Paks.

    PubMed

    Arias-Romero, Luis E; Chernoff, Jonathan

    2008-02-01

    Paks (p21-activated kinases) are effectors for the small GTPases Cdc42 (cell division cycle 42) and Rac that play fundamental roles in a wide range of cellular processes, including cell morphology, motility, survival, gene transcription, apoptosis and hormone signalling. These enzymes are widely expressed in numerous tissues and are activated by extracellular signals through GTPase-dependent and -independent mechanisms. On the basis of structural and biochemical features, the Pak family members have been classified into two groups, comprising three members each. The two Pak groups have generally been considered as two halves of a single entity, but accumulating evidence indicates that this is not so. In this review, we discuss recent advances in our understanding of the structure, regulation and function of these kinases, emphasizing the many differences between these two groups of signalling proteins.

  6. Role of PAK6 in Prostate Cancer

    DTIC Science & Technology

    2006-10-01

    of tyrosine 566 in a consensus MKK6 site (threonine-proline-tyrosine, TPY ) in the activation loop of the PAK6 kinase domain prevented activation by... TPY motif in their activation domains. The activation of PAK6 by both p38 MAP kinase and MKK6 suggests that PAK6 plays a role in the cellular...site (threo- nine-proline-tyrosine, TPY ) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was

  7. Pyrolysis of the tetra pak

    SciTech Connect

    Korkmaz, Ahmet; Yanik, Jale Brebu, Mihai; Vasile, Cornelia

    2009-11-15

    This study deals with pyrolysis of tetra pak which is widely used as an aseptic beverage packaging material. Pyrolysis experiments were carried out under inert atmosphere in a batch reactor at different temperatures and by different pyrolysis modes (one- and two-step). The yields of char, liquid and gas were quantified. Pyrolysis liquids produced were collected as three separate phases; aqueous phase, tar and polyethylene wax. Characterization of wax and the determination of the total amount of phenols in aqueous phase were performed. Chemical compositions of gas and char products relevant to fuel applications were determined. Pure aluminum can be also recovered by pyrolysis.

  8. Pyrolysis of the tetra pak.

    PubMed

    Korkmaz, Ahmet; Yanik, Jale; Brebu, Mihai; Vasile, Cornelia

    2009-11-01

    This study deals with pyrolysis of tetra pak which is widely used as an aseptic beverage packaging material. Pyrolysis experiments were carried out under inert atmosphere in a batch reactor at different temperatures and by different pyrolysis modes (one- and two-step). The yields of char, liquid and gas were quantified. Pyrolysis liquids produced were collected as three separate phases; aqueous phase, tar and polyethylene wax. Characterization of wax and the determination of the total amount of phenols in aqueous phase were performed. Chemical compositions of gas and char products relevant to fuel applications were determined. Pure aluminum can be also recovered by pyrolysis.

  9. Structure, biochemistry, and biology of PAK kinases.

    PubMed

    Kumar, Rakesh; Sanawar, Rahul; Li, Xiaodong; Li, Feng

    2017-03-20

    PAKs, p21-activated kinases, play central roles and act as converging junctions for discrete signals elicited on the cell surface and for a number of intracellular signaling cascades. PAKs phosphorylate a vast number of substrates and act by remodeling cytoskeleton, employing scaffolding, and relocating to distinct subcellular compartments. PAKs affect wide range of processes that are crucial to the cell from regulation of cell motility, survival, redox, metabolism, cell cycle, proliferation, transformation, stress, inflammation, to gene expression. Understandably, their dysregulation disrupts cellular homeostasis and severely impacts key cell functions, and many of those are implicated in a number of human diseases including cancers, neurological disorders, and cardiac disorders. Here we provide an overview of the members of the PAK family and their current status. We give special emphasis to PAK1 and PAK4, the prototypes of groups I and II, for their profound roles in cancer, the nervous system, and the heart. We also highlight other family members. We provide our perspective on the current advancements, their growing importance as strategic therapeutic targets, and our vision on the future of PAKs.

  10. PAK4–6 in cancer and neuronal development

    PubMed Central

    2012-01-01

    PAKs 4, 5 and 6 are members of the group B family of p21-activated kinases. Among this group, PAK4 has been most extensively studied. While it has essential roles in embryonic development, in adults high levels of PAK4 are frequently associated with cancer. PAK4 is overexpressed in a variety of cancers, and the Pak4 gene is amplified in some cancers. PAK4 overexpression is sufficient to cause oncogenic transformation in cells and in mouse models. The tight connection between PAK4 and cancer make it a promising diagnostic tool as well as a potential drug target. The group B PAKs also have important developmental functions. PAK4 is important for many early developmental processes, while PAK5 and PAK6 play roles in learning and memory in mice. This chapter provides an overview of the roles of the group B PAKs in cancer as well as development, and includes a discussion of PAK mediated signaling pathways and cellular functions. PMID:23125951

  11. Pak2 regulates hematopoietic progenitor cell proliferation, survival and differentiation

    PubMed Central

    Zeng, Yi; Broxmeyer, Hal E.; Staser, Karl; Chitteti, Brahmananda Reddy; Park, Su-Jung; Hahn, Seongmin; Cooper, Scott; Sun, Zejin; Jiang, Li; Yang, XianLin; Yuan, Jin; Kosoff, Rachelle; Sandusky, George; Srour, Edward F.; Chernoff, Jonathan; Clapp, Wade

    2015-01-01

    p21-activated kinase 2 (Pak2), a serine/threonine kinase, has been previously shown to be essential for hematopoietic stem cell (HSC) engraftment. However, Pak2 modulation of long-term hematopoiesis and lineage commitment remain unreported. Utilizing a conditional Pak2 knock out (KO) mouse model, we found that disruption of Pak2 in HSCs induced profound leukopenia and a mild macrocytic anemia. Although loss of Pak2 in HSCs leads to less efficient short- and long-term competitive hematopoiesis than wild type (WT) cells, it does not affect HSC self-renewal per se. Pak2 disruption decreased the survival and proliferation of multi-cytokine stimulated immature progenitors. Loss of Pak2 skewed lineage differentiation toward granulocytopoiesis and monocytopoiesis in mice as evidenced by 1) a three to six-fold increase in the percentage of peripheral blood granulocytes and a significant increase in the percentage of granulocyte-monocyte progenitors (GMPs) in mice transplanted with Pak2-disrupted BM; 2) Pak2-disrupted BM and c-kit+ cells yielded higher numbers of more mature subsets of granulocyte-monocyte colonies and polymophonuclear neutrophils (PMNs), respectively, when cultured in the presence of granulocyte-macrophage colony stimulating factor (GM-CSF). Pak2 disruption resulted respectively in decreased and increased gene expression of transcription factors JunB and c-Myc, which may suggest underlying mechanisms by which Pak2 regulates granulocyte-monocyte lineage commitment. Furthermore, Pak2 disruption led to 1) higher percentage of CD4+CD8+ double positive T cells and lower percentages of CD4+CD8− or CD4−CD8+ single positive T cells in thymus and 2) decreased numbers of mature B cells and increased numbers of Pre-Pro B cells in BM, suggesting defects in lymphopoiesis. PMID:25586960

  12. Herbal therapeutics that block the oncogenic kinase PAK1: a practical approach towards PAK1-dependent diseases and longevity.

    PubMed

    Maruta, Hiroshi

    2014-05-01

    Over 35 years research on PAKs, RAC/CDC42(p21)-activated kinases, comes of age, and in particular PAK1 has been well known to be responsible for a variety of diseases such as cancer (mainly solid tumors), Alzheimer's disease, acquired immune deficiency syndrome and other viral/bacterial infections, inflammatory diseases (asthma and arthritis), diabetes (type 2), neurofibromatosis, tuberous sclerosis, epilepsy, depression, schizophrenia, learning disability, autism, etc. Although several distinct synthetic PAK1-blockers have been recently developed, no FDA-approved PAK1 blockers are available on the market as yet. Thus, patients suffering from these PAK1-dependent diseases have to rely on solely a variety of herbal therapeutics such as propolis and curcumin that block PAK1 without affecting normal cell growth. Furthermore, several recent studies revealed that some of these herbal therapeutics significantly extend the lifespan of nematodes (C. elegans) and fruit flies (Drosophila), and PAK1-deficient worm lives longer than the wild type. Here, I outline mainly pathological phenotypes of hyper-activated PAK1 and a list of herbal therapeutics that block PAK1, but cause no side (harmful) effect on healthy people or animals.

  13. PAKS--Arbeitsbericht Nr. 5. Juli 1970. [PAKS--Working Paper No. 5. July 1970.

    ERIC Educational Resources Information Center

    Stuttgart Univ. (Germany).

    This report, the fifth in a series of working papers issued by the Project on Applied Contrastive Linguistics (PAKS) at the University of Stuttgart, is dedicated to a consideration of error analysis in language learning, here seen as relevant not only for the teacher but for the text book writer and the curriculum planner as well. An introduction…

  14. Signalling to cancer cell invasion through PAK family kinases.

    PubMed

    Whale, Andrew; Hashim, Fariesha Nur; Fram, Sally; Jones, Gareth E; Wells, Claire M

    2011-01-01

    Cancer cell metastasis involves a series of changes in cell behaviour, driven by oncogenic transformation, that leads to local tissue invasion, migration through extracellular matrix, entry into the vascular or lymphatic system and colonisation of distant sites. It is well established that the Rho family GTPases Rho, Rac and Cdc42 orchestrate many of the processes required during metastasis. The Rho family GTPases regulate cellular behaviour through their interaction with downstream effector proteins. The p-21 activated kinases (PAKs), effector proteins for Rac and Cdc42, are known to be important regulators of cell migration and invasion. There are six mammalian PAKs which can be divided into two groups: group I PAKs (PAK1-3) and group II PAKs (PAK4-6). Although the two PAK groups are architecturally similar there are differences in their mode of regulation suggesting their cellular functions are likely to be different. This review will focus on the latest evidence relating to the role of PAK family kinases in the cell signalling pathways that drive cancer cell migration and invasion.

  15. PAK promotes morphological changes by acting upstream of Rac.

    PubMed Central

    Obermeier, A; Ahmed, S; Manser, E; Yen, S C; Hall, C; Lim, L

    1998-01-01

    The serine/threonine kinase p21-activated kinase (PAK) has been implicated as a downstream effector of the small GTPases Rac and Cdc42. While these GTPases evidently induce a variety of morphological changes, the role(s) of PAK remains elusive. Here we report that overexpression of betaPAK in PC12 cells induces a Rac phenotype, including cell spreading/membrane ruffling, and increased lamellipodia formation at growth cones and shafts of nerve growth factor-induced neurites. These effects are still observed in cells expressing kinase-negative or Rac/Cdc42 binding-deficient PAK mutants, indicating that kinase- and p21-binding domains are not involved. Furthermore, lamellipodia formation in all cell lines, including those expressing Rac binding-deficient PAK, is inhibited significantly by dominant-negative RacN17. Equal inhibition is achieved by blocking PAK interaction with the guanine nucleotide exchange factor PIX using a specific N-terminal PAK fragment. We conclude that PAK, via its N-terminal non-catalytic domain, acts upstream of Rac mediating lamellipodia formation through interaction with PIX. PMID:9687501

  16. Substrate and Inhibitor Specificity of the Type II p21-Activated Kinase, PAK6

    PubMed Central

    Gao, Jia; Ha, Byung Hak; Lou, Hua Jane; Morse, Elizabeth M.; Zhang, Rong; Calderwood, David A.; Turk, Benjamin E.; Boggon, Titus J.

    2013-01-01

    The p21-activated kinases (PAKs) are important effectors of Rho-family small GTPases. The PAK family consists of two groups, type I and type II, which have different modes of regulation and signaling. PAK6, a type II PAK, influences behavior and locomotor function in mice and has an ascribed role in androgen receptor signaling. Here we show that PAK6 has a peptide substrate specificity very similar to the other type II PAKs, PAK4 and PAK5 (PAK7). We find that PAK6 catalytic activity is inhibited by a peptide corresponding to its N-terminal pseudosubstrate. Introduction of a melanoma-associated mutation, P52L, into this peptide reduces pseudosubstrate autoinhibition of PAK6, and increases phosphorylation of its substrate PACSIN1 (Syndapin I) in cells. Finally we determine two co-crystal structures of PAK6 catalytic domain in complex with ATP-competitive inhibitors. We determined the 1.4 Å co-crystal structure of PAK6 with the type II PAK inhibitor PF-3758309, and the 1.95 Å co-crystal structure of PAK6 with sunitinib. These findings provide new insights into the structure-function relationships of PAK6 and may facilitate development of PAK6 targeted therapies. PMID:24204982

  17. PAK1 translocates into nucleus in response to prolactin but not to estrogen

    SciTech Connect

    Oladimeji, Peter Diakonova, Maria

    2016-04-22

    Tyrosyl phosphorylation of the p21-activated serine–threonine kinase 1 (PAK1) has an essential role in regulating PAK1 functions in breast cancer cells. We previously demonstrated that PAK1 serves as a common node for estrogen (E2)- and prolactin (PRL)-dependent pathways. We hypothesize herein that intracellular localization of PAK1 is affected by PRL and E2 treatments differently. We demonstrate by immunocytochemical analysis that PAK1 nuclear translocation is ligand-dependent: only PRL but not E2 stimulated PAK1 nuclear translocation. Tyrosyl phosphorylation of PAK1 is essential for this nuclear translocation because phospho-tyrosyl-deficient PAK1 Y3F mutant is retained in the cytoplasm in response to PRL. We confirmed these data by Western blot analysis of subcellular fractions. In 30 min of PRL treatment, only 48% of pTyr-PAK1 is retained in the cytoplasm of PAK1 WT clone while 52% re-distributes into the nucleus and pTyr-PAK1 shuttles back to the cytoplasm by 60 min of PRL treatment. In contrast, PAK1 Y3F is retained in the cytoplasm. E2 treatment causes nuclear translocation of neither PAK1 WT nor PAK1 Y3F. Finally, we show by an in vitro kinase assay that PRL but not E2 stimulates PAK1 kinase activity in the nuclear fraction. Thus, PAK1 nuclear translocation is ligand-dependent: PRL activates PAK1 and induces translocation of activated pTyr-PAK1 into nucleus while E2 activates pTyr-PAK1 only in the cytoplasm. - Highlights: • Prolactin but not estrogen causes translocation of PAK1 into nucleus. • Tyrosyl phosphorylation of PAK1 is required for nuclear localization. • Prolactin but not estrogen stimulates PAK1 kinase activity in nucleus.

  18. The Retinoblastoma Tumor Suppressor Transcriptionally Represses Pak1 in Osteoblasts

    PubMed Central

    Sosa-García, Bernadette; Vázquez-Rivera, Viviana; González-Flores, Jonathan N.; Engel, Brienne E.; Cress, W. Douglas; Santiago-Cardona, Pedro G.

    2015-01-01

    We previously characterized the retinoblastoma tumor suppressor protein (Rb) as a regulator of adherens junction assembly and cell-to-cell adhesion in osteoblasts. This is a novel function since Rb is predominantly known as a cell cycle repressor. Herein, we characterized the molecular mechanisms by which Rb performs this function, hypothesizing that Rb controls the activity of known regulators of adherens junction assembly. We found that Rb represses the expression of the p21-activated protein kinase (Pak1), an effector of the small Rho GTPase Rac1. Rac1 is a well-known regulator of adherens junction assembly whose increased activity in cancer is linked to perturbations of intercellular adhesion. Using nuclear run-on and luciferase reporter transcription assays, we found that Pak1 repression by Rb is transcriptional, without affecting Pak1 mRNA and protein stability. Pak1 promoter bioinformatics showed multiple E2F1 binding sites within 155 base pairs of the transcriptional start site, and a Pak1-promoter region containing these E2F sites is susceptible to transcriptional inhibition by Rb. Chromatin immunoprecipitations showed that an Rb-E2F complex binds to the region of the Pak1 promoter containing the E2F1 binding sites, suggesting that Pak1 is an E2F target and that the repressive effect of Rb on Pak1 involves blocking the trans-activating capacity of E2F. A bioinformatics analysis showed elevated Pak1 expression in several solid tumors relative to adjacent normal tissue, with both Pak1 and E2F increased relative to normal tissue in breast cancer, supporting a cancer etiology for Pak1 up-regulation. Therefore, we propose that by repressing Pak1 expression, Rb prevents Rac1 hyperactivity usually associated with cancer and related to cytoskeletal derangements that disrupt cell adhesion, consequently enhancing cancer cell migratory capacity. This de-regulation of cell adhesion due to Rb loss could be part of the molecular events associated with cancer progression

  19. Rac1-PAK2 pathway is essential for zebrafish heart regeneration

    SciTech Connect

    Peng, Xiangwen; He, Quanze; Li, Guobao; Ma, Jinmin; Zhong, Tao P.

    2016-04-15

    P-21 activated kinases, or PAKs, are serine–threonine kinases that play important roles in diverse heart functions include heart development, cardiovascular development and function in a range of models; however, the mechanisms by which PAKs mediate heart regeneration are unknown. Here, we demonstrate that PAK2 and PAK4 expression is induced in cardiomyocytes and vessels, respectively, following zebrafish heart injury. Inhibition of PAK2 and PAK4 using a specific small molecule inhibitor impedes cardiomyocyte proliferation/dedifferentiation and cardiovascular regeneration, respectively. Cdc42 is specifically expressed in the ventricle and may function upstream of PAK2 but not PAK4 under normal conditions and that cardiomyocyte proliferentation during heart regeneration relies on Rac1-mediated activation of Pak2. Our results indicate that PAKs play a key role in heart regeneration.

  20. Rac1-PAK2 pathway is essential for zebrafish heart regeneration.

    PubMed

    Peng, Xiangwen; He, Quanze; Li, Guobao; Ma, Jinmin; Zhong, Tao P

    2016-04-15

    P-21 activated kinases, or PAKs, are serine-threonine kinases that play important roles in diverse heart functions include heart development, cardiovascular development and function in a range of models; however, the mechanisms by which PAKs mediate heart regeneration are unknown. Here, we demonstrate that PAK2 and PAK4 expression is induced in cardiomyocytes and vessels, respectively, following zebrafish heart injury. Inhibition of PAK2 and PAK4 using a specific small molecule inhibitor impedes cardiomyocyte proliferation/dedifferentiation and cardiovascular regeneration, respectively. Cdc42 is specifically expressed in the ventricle and may function upstream of PAK2 but not PAK4 under normal conditions and that cardiomyocyte proliferentation during heart regeneration relies on Rac1-mediated activation of Pak2. Our results indicate that PAKs play a key role in heart regeneration. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. ARROW-PAK Macroencapsulation. Innovative Technology Summary Report

    SciTech Connect

    2002-04-01

    An ARROW-PAK is a high density polyethylene (HDPE) tube, about 21 feet long and 30 inches wide. Each ARROW-PAK can hold the equivalent of 21 55-gallon drums of mixed waste debris. Each tube is fused to HDPE endcaps using localized heating and high pressure contact. The sleeves and encaps form a tube for macroencapsulating mixed waste debris. The ARROW-PAK may achieve a mixed waste debris volume one-fourth that of the conventional macroencapsulation approach. The mixed waste debris is loaded into 55-gallon drums. Once filled a 'supercompactor' crushes the drums into 12-inch thick pucks. Three pucks can be loaded into a standard 85-gallon metal drum known as an 'overpack'. Seven overpacks fit into each ARROW-PAK.

  2. NMR binding and crystal structure reveal that intrinsically-unstructured regulatory domain auto-inhibits PAK4 by a mechanism different from that of PAK1.

    PubMed

    Wang, Wei; Lim, Liangzhong; Baskaran, Yohendran; Manser, Ed; Song, Jianxing

    2013-08-16

    Six human PAK members are classified into groups I (PAKs 1-3) and II (PAK4-6). Previously, only group I PAKs were thought to be auto-inhibited but very recently PAK4, the prototype of group II PAKs, has also been shown to be auto-inhibited by its N-terminal regulatory domain. However, the complete auto-inhibitory domain (AID) sequence remains undefined and the mechanism underlying its auto-inhibition is largely elusive. Here, the N-terminal regulatory domain of PAK4 sufficient for auto-inhibiting and binding Cdc42/Rac was characterized to be intrinsically unstructured, but nevertheless we identified the entire AID sequence by NMR. Strikingly, an AID peptide was derived by deleting the binding-unnecessary residues, which has a Kd of 320 nM to the PAK4 catalytic domain. Consequently, the PAK4 crystal structure complexed with the entire AID has been determined, which reveals that the complete kinase cleft is occupied by 20 AID residuescomposed of an N-terminal α-helix and a previously-identified pseudosubstrate motif, thus achieving auto-inhibition. Our study reveals that PAK4 is auto-inhibited by a novel mechanism which is completely different from that for PAK1, thus bearing critical implications for design of inhibitors specific for group II PAKs. Copyright © 2013 Elsevier Inc. All rights reserved.

  3. LAT-independent Erk activation via Bam32-PLC-γ1-Pak1 complexes: GTPase-independent Pak1 activation.

    PubMed

    Rouquette-Jazdanian, Alexandre K; Sommers, Connie L; Kortum, Robert L; Morrison, Deborah K; Samelson, Lawrence E

    2012-10-26

    In T cells, the adaptor Bam32 is coupled to Erk activation downstream of the TCR by an unknown mechanism. We characterized in Jurkat cells and primary T lymphocytes a pathway dependent on Bam32-PLC-γ1-Pak1 complexes, in which Pak1 kinase activates Raf-1 and Mek-1, both upstream of Erk. In the Bam32-PLC-γ1-Pak1 complex, catalytically inactive PLC-γ1 is used as a scaffold linking Bam32 to Pak1. PLC-γ1(C-SH2) directly binds S141 of Bam32, preventing LAT-mediated activation of Ras by PLC-γ1. The Bam32-PLC-γ1 interaction enhances the binding of the SH3 domain of the phospholipase with Pak1. The PLC-γ1(SH3)-Pak1 interaction activates Pak1 independently of the small GTPases Rac1/Cdc42, previously described as being the only activators of Pak1 in T cells. Direct binding of the SH3 domain of PLC-γ1 to Pak1 dissociates inactive Pak1 homodimers, a mechanism required for Pak1 activation. We have thus uncovered a LAT/Ras-independent, Bam32-nucleated pathway that activates Erk signaling in T cells.

  4. LAT-Independent Erk Activation via Bam32-PLC-γ1-Pak1 Complexes: GTPase-Independent Pak1 Activation

    PubMed Central

    Rouquette-Jazdanian, Alexandre K.; Sommers, Connie L.; Kortum, Robert L.; Morrison, Deborah K.; Samelson, Lawrence E.

    2012-01-01

    SUMMARY In T cells the adapter Bam32 is coupled to Erk activation downstream of the TCR by an unknown mechanism. We characterized in Jurkat cells and primary T lymphocytes, a pathway dependent on Bam32-PLC-γ1-Pak1 complexes, in which Pak1 kinase activates Raf-1 and Mek-1, both upstream of Erk. In the Bam32-PLC-γ1-Pak1 complex, catalytically inactive PLC-γ1 is used as a scaffold linking Bam32 to Pak1. PLC-γ1(C-SH2) directly binds S141 of Bam32, preventing LAT-mediated activation of Ras by PLC-γ1. The Bam32-PLC-γ1 interaction enhances the binding of the SH3 domain of the phospholipase with Pak1. The PLC-γ1(SH3)-Pak1 interaction activates Pak1 independently of the small GTPases Rac1/Cdc42, previously described as being the only activators of Pak1 in T cells. Direct binding of the SH3 domain of PLC-γ1 to Pak1 dissociates inactive Pak1 homodimers, a mechanism required for Pak1 activation. We have thus uncovered a LAT/Ras-independent, Bam32-nucleated pathway that activates Erk signaling in T cells. PMID:22981863

  5. Development of a Hydronic Rooftop Unit-HyPak-MA

    SciTech Connect

    Lee, Eric; Berman, Mark

    2009-11-14

    The majority of U.S. commercial floor space is cooled by rooftop HVAC units (RTUs). RTU popularity derives chiefly from their low initial cost and relative ease of service access without disturbing building occupants. Unfortunately, current RTUs are inherently inefficient due to a combination of characteristics that unnecessarily increase cooling loads and energy use. 36% percent of annual U.S. energy, and two-thirds of electricity, is consumed in and by buildings. Commercial buildings consume approximately 4.2 quads of energy each year at a cost of $230 billion per year, with HVAC equipment consuming 1.2 quads of electricity. More than half of all U.S. commercial floor space is cooled by packaged HVAC units, most of which are rooftop units (RTUs). Inefficient RTUs create an estimated 3.5% of U.S. CO{sub 2} emissions, thus contributing significantly to global warming5. Also, RTUs often fail to maintain adequate ventilation air and air filtration, reducing indoor air quality. This is the second HyPak project to be supported by DOE through NETL. The prior project, referred to as HyPak-1 in this report, had two rounds of prototype fabrication and testing as well as computer modeling and market research. The HyPak-1 prototypes demonstrated the high performance capabilities of the HyPak concept, but made it clear that further development was required to reduce heat exchanger cost and improve system reliability before HyPak commercialization can commence. The HyPak-1 prototypes were limited to about 25% ventilation air fraction, limiting performance and marketability. The current project is intended to develop a 'mixed-air' product that is capable of full 0-100% modulation in ventilation air fraction, hence it was referred to as HyPak-MA in the proposal. (For simplicity, the -MA has been dropped when referencing the current project.) The objective of the HyPak Project is to design, develop and test a hydronic RTU that provides a quantum improvement over conventional RTU

  6. PAK1 modulates a PPARγ/NF-κB cascade in intestinal inflammation

    PubMed Central

    Dammann, Kyle; Khare, Vineeta; Lang, Michaela; Claudel, Thierry; Harpain, Felix; Granofszky, Nicolas; Evstatiev, Rayko; Williams, Jonathan M.; Pritchard, D. Mark; Watson, Alastair; Gasche, Christoph

    2015-01-01

    P21-activated kinases (PAKs) are multifunctional effectors of Rho GTPases with both kinase and scaffolding activity. Here, we investigated the effects of inflammation on PAK1 signaling and its role in colitis-driven carcinogenesis. PAK1 and p-PAK1 (Thr423) were assessed by immunohistochemistry, immunofluorescence, and Western blot. C57BL6/J wildtype mice were treated with a single intraperitoneal TNFα injection. Small intestinal organoids from these mice and from PAK1-KO mice were cultured with TNFα. NF-κB and PPARγ were analyzed upon PAK1 overexpression and silencing for transcriptional/translational regulation. PAK1 expression and activation was increased on the luminal intestinal epithelial surface in inflammatory bowel disease and colitis-associated cancer. PAK1 was phosphorylated upon treatment with IFNγ, IL-1β, and TNFα. In vivo, mice administered with TNFα showed increased p-PAK1 in intestinal villi, which was associated with nuclear p65 and NF-κB activation. p65 nuclear translocation downstream of TNFα was strongly inhibited in PAK1-KO small intestinal organoids. PAK1 overexpression induced a PAK1–p65 interaction as visualized by co-immunoprecipitation, nuclear translocation, and increased NF-κB transactivation, all of which were impeded by kinase-dead PAK1. Moreover, PAK1 overexpression downregulated PPARγ and mesalamine recovered PPARγ through PAK1 inhibition. On the other hand PAK1 silencing inhibited NF-κB, which was recovered using BADGE, a PPARγ antagonist. Altogether these data demonstrate that PAK1 overexpression and activation in inflammation and colitis-associated cancer promote NF-κB activity via suppression of PPARγ in intestinal epithelial cells. PMID:26036343

  7. Autophosphorylation-dependent degradation of Pak1, triggered by the Rho-family GTPase, Chp

    PubMed Central

    Weisz Hubsman, Monika; Volinsky, Natalia; Manser, Edward; Yablonski, Deborah; Aronheim, Ami

    2007-01-01

    The Paks (p21-activated kinases) Pak1, Pak2 and Pak3 are among the most studied effectors of the Rho-family GTPases, Rac, Cdc42 (cell division cycle 42) and Chp (Cdc42 homologous protein). Pak kinases influence a variety of cellular functions, but the process of Pak down-regulation, following activation, is poorly understood. In the present study, we describe for the first time a negative-inhibitory loop generated by the small Rho-GTPases Cdc42 and Chp, resulting in Pak1 inhibition. Upon overexpression of Chp, we unexpectedly observed a T-cell migration phenotype consistent with Paks inhibition. In line with this observation, overexpression of either Chp or Cdc42 caused a marked reduction in the level of Pak1 protein in a number of different cell lines. Chp-induced degradation was accompanied by ubiquitination of Pak1, and was dependent on the proteasome. The susceptibility of Pak1 to Chp-induced degradation depended on its p21-binding domain, kinase activity and a number of Pak1 autophosphorylation sites, whereas the PIX- (Pak-interacting exchange factor) and Nck-binding sites were not required. Together, these results implicate Chp-induced kinase autophosphorylation in the degradation of Pak1. The N-terminal domain of Chp was found to be required for Chp-induced degradation, although not for Pak1 activation, suggesting that Chp provides a second function, distinct from kinase activation, to trigger Pak degradation. Collectively, our results demonstrate a novel mechanism of signal termination mediated by the Rho-family GTPases Chp and Cdc42, which results in ubiquitin-mediated degradation of one of their direct effectors, Pak1. PMID:17355222

  8. Autophosphorylation-dependent degradation of Pak1, triggered by the Rho-family GTPase, Chp.

    PubMed

    Weisz Hubsman, Monika; Volinsky, Natalia; Manser, Edward; Yablonski, Deborah; Aronheim, Ami

    2007-06-15

    The Paks (p21-activated kinases) Pak1, Pak2 and Pak3 are among the most studied effectors of the Rho-family GTPases, Rac, Cdc42 (cell division cycle 42) and Chp (Cdc42 homologous protein). Pak kinases influence a variety of cellular functions, but the process of Pak down-regulation, following activation, is poorly understood. In the present study, we describe for the first time a negative-inhibitory loop generated by the small Rho-GTPases Cdc42 and Chp, resulting in Pak1 inhibition. Upon overexpression of Chp, we unexpectedly observed a T-cell migration phenotype consistent with Paks inhibition. In line with this observation, overexpression of either Chp or Cdc42 caused a marked reduction in the level of Pak1 protein in a number of different cell lines. Chp-induced degradation was accompanied by ubiquitination of Pak1, and was dependent on the proteasome. The susceptibility of Pak1 to Chp-induced degradation depended on its p21-binding domain, kinase activity and a number of Pak1 autophosphorylation sites, whereas the PIX- (Pak-interacting exchange factor) and Nck-binding sites were not required. Together, these results implicate Chp-induced kinase autophosphorylation in the degradation of Pak1. The N-terminal domain of Chp was found to be required for Chp-induced degradation, although not for Pak1 activation, suggesting that Chp provides a second function, distinct from kinase activation, to trigger Pak degradation. Collectively, our results demonstrate a novel mechanism of signal termination mediated by the Rho-family GTPases Chp and Cdc42, which results in ubiquitin-mediated degradation of one of their direct effectors, Pak1.

  9. PAK1 modulates a PPARγ/NF-κB cascade in intestinal inflammation.

    PubMed

    Dammann, Kyle; Khare, Vineeta; Lang, Michaela; Claudel, Thierry; Harpain, Felix; Granofszky, Nicolas; Evstatiev, Rayko; Williams, Jonathan M; Pritchard, D Mark; Watson, Alastair; Gasche, Christoph

    2015-10-01

    P21-activated kinases (PAKs) are multifunctional effectors of Rho GTPases with both kinase and scaffolding activity. Here, we investigated the effects of inflammation on PAK1 signaling and its role in colitis-driven carcinogenesis. PAK1 and p-PAK1 (Thr423) were assessed by immunohistochemistry, immunofluorescence, and Western blot. C57BL6/J wildtype mice were treated with a single intraperitoneal TNFα injection. Small intestinal organoids from these mice and from PAK1-KO mice were cultured with TNFα. NF-κB and PPARγ were analyzed upon PAK1 overexpression and silencing for transcriptional/translational regulation. PAK1 expression and activation was increased on the luminal intestinal epithelial surface in inflammatory bowel disease and colitis-associated cancer. PAK1 was phosphorylated upon treatment with IFNγ, IL-1β, and TNFα. In vivo, mice administered with TNFα showed increased p-PAK1 in intestinal villi, which was associated with nuclear p65 and NF-κB activation. p65 nuclear translocation downstream of TNFα was strongly inhibited in PAK1-KO small intestinal organoids. PAK1 overexpression induced a PAK1-p65 interaction as visualized by co-immunoprecipitation, nuclear translocation, and increased NF-κB transactivation, all of which were impeded by kinase-dead PAK1. Moreover, PAK1 overexpression downregulated PPARγ and mesalamine recovered PPARγ through PAK1 inhibition. On the other hand PAK1 silencing inhibited NF-κB, which was recovered using BADGE, a PPARγ antagonist. Altogether these data demonstrate that PAK1 overexpression and activation in inflammation and colitis-associated cancer promote NF-κB activity via suppression of PPARγ in intestinal epithelial cells.

  10. Live donor kidney - PAK versus SPK: how to decide?

    PubMed

    Stites, Erik; Wiseman, Alexander C

    2017-08-01

    Patients with type 1 diabetes and end stage renal disease face a complex choice when considering the relative risks and benefits of kidney transplant alone with or without subsequent pancreas after kidney transplant (PAK) or simultaneous kidney pancreas transplant (SPK). SPK is considered the optimal treatment regarding long-term patient survival, but when also faced with the option of living donor kidney transplant with the potential for PAK later, the ideal option is less clear. This review summarizes the current literature regarding SPK, living donor kidney transplant alone, and PAK transplant outcomes and examines the relative risks of pre- and posttransplant variables that impact patient and graft survival to help inform this complex treatment decision.

  11. Pak1 and Pak2 are activated in recurrent respiratory papillomas, contributing to one pathway of Rac1-mediated COX-2 expression

    PubMed Central

    Wu, Rong; Abramson, Allan L.; Symons, Marc H.; Steinberg, Bettie M.

    2010-01-01

    Recurrent respiratory papillomas are pre-malignant tumors of the airway caused by human papillomaviruses (HPVs), primarily types 6 and 11. We had reported that respiratory papillomas overexpress the epidermal growth factor receptor (EGFR), the small GTPase Rac1 and cyclooxygenase-2 (COX-2), and have enhanced Nuclear Factor-κB (NFκB) activation with decreased levels of IκB-β but not IκB-α. We also showed that EGFR-activated Rac1 mediates expression of COX-2 through activation of p38 mitogen activated protein kinase. We have now asked whether the p21-activated kinases Pak 1 or Pak2 mediate activation of p38 by Rac1 in papilloma cells. Pak1 and Pak2 were constitutively activated in vivo in papilloma tissue compared to normal epithelium, and Rac1 siRNA reduced the level of both phospho-Pak1 and phospho-Pak2 in cultured papilloma cells. Reduction in Pak1 and Pak2 with siRNA decreased COX-2 expression in papilloma cells, increased levels of IκB-Iβ and reduced nuclear localization of NF-κB, but had no effect on p38 phosphorylation. Our studies suggest that Rac1→ Pak1/Pak2→ NFκB is a separate pathway that contributes to the expression of COX-2 in HPV-induced papillomas independently of the previously described Rac1→ p38 → COX-2 pathway. PMID:20131316

  12. Dock and Pak regulate olfactory axon pathfinding in Drosophila.

    PubMed

    Ang, Lay-Hong; Kim, Jenny; Stepensky, Vitaly; Hing, Huey

    2003-04-01

    The convergence of olfactory axons expressing particular odorant receptor (Or) genes on spatially invariant glomeruli in the brain is one of the most dramatic examples of precise axon targeting in developmental neurobiology. The cellular and molecular mechanisms by which olfactory axons pathfind to their targets are poorly understood. We report here that the SH2/SH3 adapter Dock and the serine/threonine kinase Pak are necessary for the precise guidance of olfactory axons. Using antibody localization, mosaic analyses and cell-type specific rescue, we observed that Dock and Pak are expressed in olfactory axons and function autonomously in olfactory neurons to regulate the precise wiring of the olfactory map. Detailed analyses of the mutant phenotypes in whole mutants and in small multicellular clones indicate that Dock and Pak do not control olfactory neuron (ON) differentiation, but specifically regulate multiple aspects of axon trajectories to guide them to their cognate glomeruli. Structure/function studies show that Dock and Pak form a signaling pathway that mediates the response of olfactory axons to guidance cues in the developing antennal lobe (AL). Our findings therefore identify a central signaling module that is used by ONs to project to their cognate glomeruli.

  13. Pak functions downstream of Dock to regulate photoreceptor axon guidance in Drosophila.

    PubMed

    Hing, H; Xiao, J; Harden, N; Lim, L; Zipursky, S L

    1999-06-25

    The SH2/SH3 adaptor protein Dock has been proposed to transduce signals from guidance receptors to the actin cytoskeleton in Drosophila photoreceptor (R cell) growth cones. Here, we demonstrate that Drosophila p21-activated kinase (Pak) is required in a Dock pathway regulating R cell axon guidance and targeting. Dock and Pak colocalize to R cell axons and growth cones, physically interact, and their loss-of-function phenotypes are indistinguishable. Normal patterns of R cell connectivity require Pak's kinase activity and binding sites for both Dock and Cdc42/Rac. A membrane-tethered form of Pak (Pak(myr) acts as a dominant gain-of-function protein. Retinal expression of Pak(myr) rescues the R cell connectivity phenotype in dock mutants. These data establish Pak as a critical regulator of axon guidance and a downstream effector of Dock in vivo.

  14. Lethality of PAK3 and SGK2 shRNAs to human papillomavirus positive cervical cancer cells is independent of PAK3 and SGK2 knockdown.

    PubMed

    Zhou, Nannan; Ding, Bo; Agler, Michele; Cockett, Mark; McPhee, Fiona

    2015-01-01

    The p21-activated kinase 3 (PAK3) and the serum and glucocorticoid-induced kinase 2 (SGK2) have been previously proposed as essential kinases for human papillomavirus positive (HPV+) cervical cancer cell survival. This was established using a shRNA knockdown approach. To validate PAK3 and SGK2 as potential targets for HPV+ cervical cancer therapy, the relationship between shRNA-induced phenotypes in HPV+ cervical cancer cells and PAK3 or SGK2 knockdown was carefully examined. We observed that the phenotypes of HPV+ cervical cancer cells induced by various PAK3 and SGK2 shRNAs could not be rescued by complement expression of respective cDNA constructs. A knockdown-deficient PAK3 shRNA with a single mismatch was sufficient to inhibit HeLa cell growth to a similar extent as wild-type PAK3 shRNA. The HPV+ cervical cancer cells were also susceptible to several non-human target shRNAs. The discrepancy between PAK3 and SGK2 shRNA-induced apoptosis and gene expression knockdown, as well as cell death stimulation, suggested that these shRNAs killed HeLa cells through different pathways that may not be target-specific. These data demonstrated that HPV+ cervical cancer cell death was not associated with RNAi-induced PAK3 and SGK2 knockdown but likely through off-target effects.

  15. Gene Expression Analysis of Pak Choi in Response to Vernalization

    PubMed Central

    Sun, Mengxia; Qi, Xianhui; Hou, Leiping; Xu, Xiaoyong; Zhu, Zhujun; Li, Meilan

    2015-01-01

    Pak choi is a seed vernalization-type plant whose vernalization mechanism is currently unclear. Therefore, it is critical to discover genes related to vernalization and research its functions during vernalization in pak choi. Here, the gene expression profiles in the shoot apex were analyzed after low temperature treatment using high-throughput RNA sequencing technology. The results showed that there are 1,664 and 1,192 differentially expressed genes (DEGs) in pak choi in cold treatment ending and before flower bud differentiation, respectively, including 42 genes that exhibited similar expression trend at both stages. Detailed annotation revealed that the proteins encoded by the DEGs are located in the extracellular region, cell junction and extracellular matrix. These proteins exhibit activity such as antioxidant activity and binding protein/transcription factor activity, and they are involved in signal transduction and the immune system/biological processes. Among the DEGs, Bra014527 was up-regulated in low temperature treatment ending, Bra024097 was up-regulated before flower bud differentiation and Bra035940 was down-regulated at both stages in low temperature-treated shoot apices. Homologues of these genes in A. thaliana, AT3G59790, AT4G30200 and AT5G61150, are involved in flowering and vernalization, suggesting that they take part in the vernalization process in pak choi. Further pathway enrichment analysis revealed that most genes were enriched in the tryptophan metabolism and glucosinolate biosynthesis pathways. However, the functions of tryptophan and glucosinolate in vernalization are not yet clear and require further analysis. PMID:26517271

  16. Trio combines with dock to regulate Pak activity during photoreceptor axon pathfinding in Drosophila.

    PubMed

    Newsome, T P; Schmidt, S; Dietzl, G; Keleman, K; Asling, B; Debant, A; Dickson, B J

    2000-04-28

    Correct pathfinding by Drosophila photoreceptor axons requires recruitment of p21-activated kinase (Pak) to the membrane by the SH2-SH3 adaptor Dock. Here, we identify the guanine nucleotide exchange factor (GEF) Trio as another essential component in photoreceptor axon guidance. Regulated exchange activity of one of the two Trio GEF domains is critical for accurate pathfinding. This GEF domain activates Rac, which in turn activates Pak. Mutations in trio result in projection defects similar to those observed in both Pak and dock mutants, and trio interacts genetically with Rac, Pak, and dock. These data define a signaling pathway from Trio to Rac to Pak that links guidance receptors to the growth cone cytoskeleton. We propose that distinct signals transduced via Trio and Dock act combinatorially to activate Pak in spatially restricted domains within the growth cone, thereby controlling the direction of axon extension.

  17. Pak6 protein kinase is a novel effector of an atypical Rho family GTPase Chp/RhoV.

    PubMed

    Shepelev, M V; Korobko, I V

    2012-01-01

    Chp/RhoV is an atypical Rho GTPase whose functions are far from being fully understood. To date several effector proteins of Chp have been identified, including p21-activated kinases Pak1, Pak2, and Pak4. Using a yeast two-hybrid system and co-immunoprecipitation, here we show that another p21-activated kinase, Pak6, is a novel Chp-binding protein. Interaction between Chp and Pak6 depends on the activation state of the GTPase, suggesting that Pak6 is an effector protein for Chp. Point mutations in the effector domain of Chp or in the CRIB motif of Pak6 significantly impair the interaction between Chp and Pak6 upon co-immunoprecipitation, suggesting that the binding interface involves the effector domain of Chp and the CRIB motif in Pak6. We found that Chp does not affect the phosphorylation status of the S560 residue in the catalytic domain of Pak6 when Chp and Pak6 are co-expressed in HEK293 cells. Therefore, similarly to Cdc42, Chp is not likely to activate Pak6. In NCI-H1299 cells, Chp co-localizes with Pak6 on vesicular structures in activation state-dependent manner. Taking the data together, we report here the identification of p21-activated kinase Pak6 as a novel effector of the atypical Rho GTPase Chp. Our data suggest further directions in elucidating biological functions of these proteins.

  18. Pak1 and PIX regulate contact inhibition during epithelial wound healing.

    PubMed

    Zegers, Mirjam M P; Forget, Marie-Annick; Chernoff, Jonathan; Mostov, Keith E; ter Beest, Martin B A; Hansen, Steen H

    2003-08-15

    Wound healing in epithelia requires coordinated cell migration and proliferation regulated by signaling mechanisms that are poorly understood. Here we show that epithelial cells expressing constitutively active or kinase-dead mutants of the Rac/Cdc42 effector Pak1 fail to undergo growth arrest upon wound closure. Strikingly, this phenotype is only observed when the Pak1 kinase mutants are expressed in cells possessing a free lateral surface, i.e. one that is not engaged in contact with neighboring cells. The Pak1 kinase mutants perturb contact inhibition by a mechanism that depends on the Pak-interacting Rac-GEF PIX. In control cells, endogenous activated Pak and PIX translocate from focal complexes to cell-cell contacts during wound closure. This process is abrogated in cells expressing Pak1 kinase mutants. In contrast, Pak1 mutants rendered defective in PIX binding do not impede translocation of activated Pak and PIX, and exhibit normal wound healing. Thus, recruitment of activated Pak and PIX to cell-cell contacts is pivotal to transduction of growth-inhibitory signals from neighboring cells in epithelial wound healing.

  19. PAK-PIX interactions regulate adhesion dynamics and membrane protrusion to control neurite outgrowth.

    PubMed

    Santiago-Medina, Miguel; Gregus, Kelly A; Gomez, Timothy M

    2013-03-01

    The roles of P21-activated kinase (PAK) in the regulation of axon outgrowth downstream of extracellular matrix (ECM) proteins are poorly understood. Here we show that PAK1-3 and PIX are expressed in the developing spinal cord and differentially localize to point contacts and filopodial tips within motile growth cones. Using a specific interfering peptide called PAK18, we found that axon outgrowth is robustly stimulated on laminin by partial inhibition of PAK-PIX interactions and PAK function, whereas complete inhibition of PAK function stalls axon outgrowth. Furthermore, modest inhibition of PAK-PIX stimulates the assembly and turnover of growth cone point contacts, whereas strong inhibition over-stabilizes adhesions. Point mutations within PAK confirm the importance of PIX binding. Together our data suggest that regulation of PAK-PIX interactions in growth cones controls neurite outgrowth by influencing the activity of several important mediators of actin filament polymerization and retrograde flow, as well as integrin-dependent adhesion to laminin.

  20. PAK4 interacts with p85 alpha: implications for pancreatic cancer cell migration

    PubMed Central

    King, Helen; Thillai, Kiruthikah; Whale, Andrew; Arumugam, Prabhu; Eldaly, Hesham; Kocher, Hemant M.; Wells, Claire M.

    2017-01-01

    It has been reported that p21-activated kinase 4 (PAK4) is amplified in pancreatic cancer tissue. PAK4 is a member of the PAK family of serine/threonine kinases, which act as effectors for several small GTPases, and has been specifically identified to function downstream of HGF-mediated c-Met activation in a PI3K dependent manner. However, the functionality of PAK4 in pancreatic cancer and the contribution made by HGF signalling to pancreatic cancer cell motility remain to be elucidated. We now find that elevated PAK4 expression is coincident with increased expression levels of c-Met and the p85α subunit of PI3K. Furthermore, we demonstrate that pancreatic cancer cells have a specific motility response to HGF both in 2D and 3D physiomimetic organotypic assays; which can be suppressed by inhibition of PI3K. Significantly, we report a specific interaction between PAK4 and p85α and find that PAK4 deficient cells exhibit a reduction in Akt phosphorylation downstream of HGF signalling. These results implicate a novel role for PAK4 within the PI3K pathway via interaction with p85α. Thus, PAK4 could be an essential player in PDAC progression representing an interesting therapeutic opportunity. PMID:28205613

  1. Optimization of a Dibenzodiazepine Hit to a Potent and Selective Allosteric PAK1 Inhibitor

    PubMed Central

    2015-01-01

    The discovery of inhibitors targeting novel allosteric kinase sites is very challenging. Such compounds, however, once identified could offer exquisite levels of selectivity across the kinome. Herein we report our structure-based optimization strategy of a dibenzodiazepine hit 1, discovered in a fragment-based screen, yielding highly potent and selective inhibitors of PAK1 such as 2 and 3. Compound 2 was cocrystallized with PAK1 to confirm binding to an allosteric site and to reveal novel key interactions. Compound 3 modulated PAK1 at the cellular level and due to its selectivity enabled valuable research to interrogate biological functions of the PAK1 kinase. PMID:26191365

  2. Development of 2, 4-diaminoquinazoline derivatives as potent PAK4 inhibitors by the core refinement strategy.

    PubMed

    Hao, Chenzhou; Huang, Wanxu; Li, Xiaodong; Guo, Jing; Chen, Meng; Yan, Zizheng; Wang, Kai; Jiang, Xiaolin; Song, Shuai; Wang, Jian; Zhao, Dongmei; Li, Feng; Cheng, Maosheng

    2017-05-05

    Upon analysis of the reported crystal structure of PAK4 inhibitor KY04031 (PAK4 IC50 = 0.790 μM) in the active site of PAK4, we investigated the possibility of changing the triazine core of KY04031 to a quinazoline. Using KY04031 as a starting compound, a library of 2, 4-diaminoquinazoline derivatives were designed and synthesized. These compounds were evaluated for PAK4 inhibition, leading to the identification of compound 9d (PAK4 IC50 = 0.033 μM). Compound 9d significantly induced the cell cycle in the G1/S phase and inhibited migration and invasion of A549 cells that over-express PAK4 via regulation of the PAK4-LIMK1 signalling pathway. A docking study of compound 9d was performed to elucidate its possible binding modes and to provide a structural basis for further structure-guided design of PAK4 inhibitors. Compound 9d may serve as a lead compound for anticancer drug discovery and as a valuable research probe for further biological investigation of PAK4. Copyright © 2017. Published by Elsevier Masson SAS.

  3. Dynamic recruitment of PAK1 to the immunological synapse is mediated by PIX independently of SLP-76 and Vav1.

    PubMed

    Phee, Hyewon; Abraham, Robert T; Weiss, Arthur

    2005-06-01

    T cell receptor engagement activates p21-activated kinase 1 (PAK1) through a LAT-SLP-76-Nck-Vav-Rac-dependent pathway. A second independent pathway involving a GIT-PIX-PAK1 trimolecular complex is also activated by T cell receptor ligation. Here we show a Vav-independent pathway exists that leads to PAK1 activation. In addition, PAK1, PIX and GIT1 were recruited to the T cell-antigen-presenting cell contact site independently of SLP-76 and Vav1. PAK1 recruitment to the T cell-antigen-presenting cell interface required interaction with PIX, which also led to optimal PLC-gamma1 activation and T cell receptor-dependent transcriptional responses. These data indicate that a pathway involving the GIT-PIX-PAK1 complex has a crucial function in PAK1 activation by recruiting PAK1 to the immunological synapse.

  4. Prognostic Importance and Therapeutic Implications of PAK1, a Drugable Protein Kinase, in Gastroesophageal Junction Adenocarcinoma

    PubMed Central

    Xie, Liangxi; Dong, Hongmei; Chen, Yuping; Liu, Qing; Wu, Xiao; Zhou, David; Tan, Dongfeng; Zhang, Hao

    2013-01-01

    Gastroesophageal junction (GEJ) adenocarcinoma is a lethal cancer with rising incidence, yet the molecular biomarkers that have strong prognostic impact and also hold great therapeutic promise remain elusive. We used a data mining approach and identified the p21 protein-activated kinase 1 (PAK1), an oncogene and drugable protein kinase, to be among the most promising targets for GEJ adenocarcinoma. Immunoblot analysis and data mining demonstrated that PAK1 protein and mRNA were upregulated in cancer tissues compared to the noncancerous tissues. Immunohistochemistry revealed PAK1 overexpression in 72.6% of primary GEJ adenocarcinomas (n = 113). A step-wise increase in PAK1 levels was noted from paired normal epithelium, to atypical hyperplasia and adenocarcinoma. PAK1 overexpression in tumor was associated with lymph node (LN) metastasis (P<0.001), advanced tumor stage (P<0.001), large tumor size (P = 0.006), residual surgical margin (P = 0.033), and unfavorable overall survival (P<0.001). Multivariate analysis showed PAK1 overexpression is an independent high-risk prognostic predictor (P<0.001). Collectively, PAK1 is overexpressed during tumorigenic progression and its upregulation correlates with malignant properties mainly relevant to invasion and metastasis. PAK1 expression could serve as a prognostic predictor that holds therapeutic promise for GEJ adenocarcinoma. PMID:24236193

  5. PAK4 suppresses PDZ-RhoGEF activity to drive invadopodia maturation in melanoma cells

    PubMed Central

    Nicholas, Nicole S.; Pipili, Aikaterini; Lesjak, Michaela S.; Ameer, Simon M.; Geh, Jenny L. C.; Healy, Ciaran; Ross, Alistair D. MacKenzie; Parsons, Maddy; Nestle, Frank O.; Lacy, Katie E.; Wells, Claire M.

    2016-01-01

    Cancer cells are thought to use actin rich invadopodia to facilitate matrix degradation. Formation and maturation of invadopodia requires the co-ordained activity of Rho-GTPases, however the molecular mechanisms that underlie the invadopodia lifecycle are not fully elucidated. Previous work has suggested a formation and disassembly role for Rho family effector p-21 activated kinase 1 (PAK1) however, related family member PAK4 has not been explored. Systematic analysis of isoform specific depletion using in vitro and in vivo invasion assays revealed there are differential invadopodia-associated functions. We consolidated a role for PAK1 in the invadopodia formation phase and identified PAK4 as a novel invadopodia protein that is required for successful maturation. Furthermore, we find that PAK4 (but not PAK1) mediates invadopodia maturation likely via inhibition of PDZ-RhoGEF. Our work points to an essential role for both PAKs during melanoma invasion but provides a significant advance in our understanding of differential PAK function. PMID:27765920

  6. ERK activation of p21 activated kinase-1 (Pak1) is critical for medulloblastoma cell migration

    PubMed Central

    Yuan, Liangping; Santi, Mariarita; Rushing, Elisabeth J.; Cornelison, Robert

    2010-01-01

    We previously identified that overexpression of the platelet-derived growth factor receptor (PDGFR) is associated with metastatic medulloblastoma (MB) and showed that PDGF treatment increases ERK activity and promotes MB cell migration. In this study, we investigated whether ERK regulates Rac1/Pak1 signaling and is critically linked to MB cell migration. Herein we demonstrate that PDGF-BB treatment of MB cells induces concomitant activation of PDGFRβ, MEK1/ERK, Rac1 and Pak1, but suppresses Rho activity, which together significantly promotes cell migration. Conversely, cells transfected with either PDGFRβ or Pak1 siRNA or treated with an inhibitor of Rac1 (NSC23766) or N-myristoyltransferase-1 (Tris-dipalladium) are unable to activate Rac1 or Pak1 in response to PDGF, and consequently, are unable to undergo PDGF-mediated cell migration. Furthermore, we also demonstrate that either chemical inhibition of MEK/ ERK (U0126) or stable downregulation of PDGFRβ by shRNA similarly results in the loss of PDGF-induced ERK phosphorylation and abolishes Rac1/Pak1 activation and cell migration in response to PDGF. However, specific depletion of Pak1 by siRNA has no effect on PDGF-induced ERK phosphorylation, indicating that in MB cells ERK signaling is Pak1-independent, but PDGF-induced migration is dependent on ERK-mediated activation of Pak1. Finally, using tissue microarrays, we detect phosphorylated Pak1 in 53% of medulloblastomas and show that immunopositivity is associated with unfavorable outcome. We conclude that Rac1/Pak1 signaling is critical to MB cell migration and is functionally dependent on PDGFRβ/ERK activity. PMID:20526801

  7. Pak2 is required for actin cytoskeleton remodeling, TCR signaling, and normal thymocyte development and maturation

    PubMed Central

    Phee, Hyewon; Au-Yeung, Byron B; Pryshchep, Olga; O'Hagan, Kyle Leonard; Fairbairn, Stephanie Grace; Radu, Maria; Kosoff, Rachelle; Mollenauer, Marianne; Cheng, Debra; Chernoff, Jonathan; Weiss, Arthur

    2014-01-01

    The molecular mechanisms that govern thymocyte development and maturation are incompletely understood. The P21-activated kinase 2 (Pak2) is an effector for the Rho family GTPases Rac and Cdc42 that regulate actin cytoskeletal remodeling, but its role in the immune system remains poorly understood. In this study, we show that T-cell specific deletion of Pak2 gene in mice resulted in severe T cell lymphopenia accompanied by marked defects in development, maturation, and egress of thymocytes. Pak2 was required for pre-TCR β-selection and positive selection. Surprisingly, Pak2 deficiency in CD4 single positive thymocytes prevented functional maturation and reduced expression of S1P1 and KLF2. Mechanistically, Pak2 is required for actin cytoskeletal remodeling triggered by TCR. Failure to induce proper actin cytoskeletal remodeling impaired PLCγ1 and Erk1/2 signaling in the absence of Pak2, uncovering the critical function of Pak2 as an essential regulator that governs the actin cytoskeleton-dependent signaling to ensure normal thymocyte development and maturation. DOI: http://dx.doi.org/10.7554/eLife.02270.001 PMID:24843022

  8. Resveratrol regulates neuronal glucose uptake and insulin sensitivity via P21-activated kinase 2 (PAK2).

    PubMed

    Varshney, Pallavi; Dey, Chinmoy Sankar

    2017-04-01

    We have recently reported P21-activated kinase 2 (PAK2), a serine/threonine kinase as a negative regulator of neuronal glucose uptake and insulin sensitivity. Resveratrol (RSV), a natural polyphenol with anti-oxidative, anti-inflammatory and anti-diabetic properties, regulates PAK2 activity in HepG2 and ESC-B5 cell apoptosis. However, regulation of PAK2 by RSV in neuronal insulin signaling pathway, if any, is still unknown. In the present study, RSV treatment significantly increased PAK2 activity under insulin-sensitive and insulin-resistant condition, along with a marked decrease in glucose uptake in differentiated N2A cells. Pretreatment with AMPK inhibitor, followed by RSV treatment resulted in reduction in PAK2 activity whereas glucose uptake showed an increase. However, pretreatment with Akt inhibitor and then RSV exposure significantly increased PAK2 activity, with a corresponding decrease in glucose uptake. RSV treatment increased AMPK activity and decreased Akt activity. In conclusion, RSV negatively regulates neuronal glucose uptake and insulin sensitivity via PAK2. Copyright © 2017. Published by Elsevier Inc.

  9. Outgrowth of Neurites from NIE-115 Neuroblastoma Cells Is Prevented on Repulsive Substrates through the Action of PAK

    PubMed Central

    Marler, Katharine J. M.; Kozma, Robert; Ahmed, Sohail; Dong, Jing-Ming; Hall, Christine; Lim, Louis

    2005-01-01

    In the central nervous system (CNS), damaged axons are inhibited from regeneration by glial scars, where secreted chondroitin sulfate proteoglycan (CSPG) and tenascin repulse outgrowth of neurites, the forerunners of axons and dendrites. During differentiation, these molecules are thought to form boundaries for guiding neurons to their correct targets. In neuroblastoma NIE-115 cells, outgrowth of neurites on laminin could be induced by serum starvation or inhibition of RhoA by Clostridium botulinum C3 toxin. The outgrowing neurites avoided crossing onto the repulsive substrate CSPG or tenascin. This avoidance response was partially overcome on expression of membrane-targeted and kinase-inactive forms of PAK. In these cells, the endogenous PAK isoforms colocalized with actin in distinctive sites, αPAK in the cell center as small clusters and along the neurite shaft and βPAK and γPAK in areas with membrane ruffles and filopodia, respectively. When isoform-specific N-terminal PAK sequences were introduced to interfere with PAK function, substantially more neurites crossed onto CSPG when cells contained a γPAK-derived peptide but not the corresponding αPAK- or βPAK-derived peptide. Thus, while neurite outgrowth can be promoted by RhoA inhibition, overcoming the accompanying repulsive guidance response will require modulation of PAK activity. These results have therapeutic implications for CNS repair processes. PMID:15923637

  10. Pak3 regulates apical-basal polarity in migrating border cells during Drosophila oogenesis.

    PubMed

    Felix, Martina; Chayengia, Mrinal; Ghosh, Ritabrata; Sharma, Aditi; Prasad, Mohit

    2015-11-01

    Group cell migration is a highly coordinated process that is involved in a number of physiological events such as morphogenesis, wound healing and tumor metastasis. Unlike single cells, collectively moving cells are physically attached to each other and retain some degree of apical-basal polarity during the migratory phase. Although much is known about direction sensing, how polarity is regulated in multicellular movement remains unclear. Here we report the role of the protein kinase Pak3 in maintaining apical-basal polarity in migrating border cell clusters during Drosophila oogenesis. Pak3 is enriched in border cells and downregulation of its function impedes border cell movement. Time-lapse imaging suggests that Pak3 affects protrusive behavior of the border cell cluster, specifically regulating the stability and directionality of protrusions. Pak3 functions downstream of guidance receptor signaling to regulate the level and distribution of F-actin in migrating border cells. We also provide evidence that Pak3 genetically interacts with the lateral polarity marker Scribble and that it regulates JNK signaling in the moving border cells. Since Pak3 depletion results in mislocalization of several apical-basal polarity markers and overexpression of Jra rescues the polarity of the Pak3-depleted cluster, we propose that Pak3 functions through JNK signaling to modulate apical-basal polarity of the migrating border cell cluster. We also observe loss of apical-basal polarity in Rac1-depleted border cell clusters, suggesting that guidance receptor signaling functions through Rac GTPase and Pak3 to regulate the overall polarity of the cluster and mediate efficient collective movement of the border cells to the oocyte boundary.

  11. Rho family-associated kinases PAK1 and rock.

    PubMed

    Maruta, Hiroshi; Nheu, Thao V; He, Hong; Hirokawa, Yumiko

    2003-01-01

    Rho family GTPases (Rho, Rac and CDC42) share around 30% sequence identity with RAS family GTPases, and are essential for RAS-induced malignant transformation, i.e., aberrant serum/anchorage-independent growth and actin cytoskeleton-linked morphological changes. Oncogenic RAS mutants such as v-Ha-RAS trigger cell cycle entry (G0-G1 transition) mainly by up-regulating cyclin D1, an activator of cyclin-dependent kinases (CDK), and down-regulating p27, a CDK inhibitor. Although both Rac and CDC42 are clearly activated by RAS, there is so far no evidence that RAS activates Rho. In this chapter, we will discuss the role of these Rho family GTPases and their effectors, in particular the Ser/Thr kinases PAK1 and Rock, in RAS-induced serum/anchorage-independent cell cycling, and discuss several potential therapeutics, peptides or chemical compounds, that could block this oncogenic cell cycle signalling pathway.

  12. Tuning PAK Activity to Rescue Abnormal Myelin Permeability in HNPP.

    PubMed

    Hu, Bo; Arpag, Sezgi; Zhang, Xuebao; Möbius, Wiebke; Werner, Hauke; Sosinsky, Gina; Ellisman, Mark; Zhang, Yang; Hamilton, Audra; Chernoff, Jonathan; Li, Jun

    2016-09-01

    Schwann cells in the peripheral nervous systems extend their membranes to wrap axons concentrically and form the insulating sheath, called myelin. The spaces between layers of myelin are sealed by myelin junctions. This tight insulation enables rapid conduction of electric impulses (action potentials) through axons. Demyelination (stripping off the insulating sheath) has been widely regarded as one of the most important mechanisms altering the action potential propagation in many neurological diseases. However, the effective nerve conduction is also thought to require a proper myelin seal through myelin junctions such as tight junctions and adherens junctions. In the present study, we have demonstrated the disruption of myelin junctions in a mouse model (Pmp22+/-) of hereditary neuropathy with liability to pressure palsies (HNPP) with heterozygous deletion of Pmp22 gene. We observed a robust increase of F-actin in Pmp22+/- nerve regions where myelin junctions were disrupted, leading to increased myelin permeability. These abnormalities were present long before segmental demyelination at the late phase of Pmp22+/- mice. Moreover, the increase of F-actin levels correlated with an enhanced activity of p21-activated kinase (PAK1), a molecule known to regulate actin polymerization. Pharmacological inhibition of PAK normalized levels of F-actin, and completely prevented the progression of the myelin junction disruption and nerve conduction failure in Pmp22+/- mice. Our findings explain how abnormal myelin permeability is caused in HNPP, leading to impaired action potential propagation in the absence of demyelination. We call it "functional demyelination", a novel mechanism upstream to the actual stripping of myelin that is relevant to many demyelinating diseases. This observation also provides a potential therapeutic approach for HNPP.

  13. Tuning PAK Activity to Rescue Abnormal Myelin Permeability in HNPP

    PubMed Central

    Hu, Bo; Zhang, Xuebao; Möbius, Wiebke; Werner, Hauke; Sosinsky, Gina; Ellisman, Mark; Zhang, Yang; Hamilton, Audra; Chernoff, Jonathan; Li, Jun

    2016-01-01

    Schwann cells in the peripheral nervous systems extend their membranes to wrap axons concentrically and form the insulating sheath, called myelin. The spaces between layers of myelin are sealed by myelin junctions. This tight insulation enables rapid conduction of electric impulses (action potentials) through axons. Demyelination (stripping off the insulating sheath) has been widely regarded as one of the most important mechanisms altering the action potential propagation in many neurological diseases. However, the effective nerve conduction is also thought to require a proper myelin seal through myelin junctions such as tight junctions and adherens junctions. In the present study, we have demonstrated the disruption of myelin junctions in a mouse model (Pmp22+/-) of hereditary neuropathy with liability to pressure palsies (HNPP) with heterozygous deletion of Pmp22 gene. We observed a robust increase of F-actin in Pmp22+/- nerve regions where myelin junctions were disrupted, leading to increased myelin permeability. These abnormalities were present long before segmental demyelination at the late phase of Pmp22+/- mice. Moreover, the increase of F-actin levels correlated with an enhanced activity of p21-activated kinase (PAK1), a molecule known to regulate actin polymerization. Pharmacological inhibition of PAK normalized levels of F-actin, and completely prevented the progression of the myelin junction disruption and nerve conduction failure in Pmp22+/- mice. Our findings explain how abnormal myelin permeability is caused in HNPP, leading to impaired action potential propagation in the absence of demyelination. We call it “functional demyelination”, a novel mechanism upstream to the actual stripping of myelin that is relevant to many demyelinating diseases. This observation also provides a potential therapeutic approach for HNPP. PMID:27583434

  14. p21-Activated kinase1 (Pak1) is a negative regulator of NADPH-oxidase 2 in ventricular myocytes.

    PubMed

    DeSantiago, Jaime; Bare, Dan J; Xiao, Lei; Ke, Yunbo; Solaro, R John; Banach, Kathrin

    2014-02-01

    Ischemic conditions reduce the activity of the p21-activated kinase (Pak1) resulting in increased arrhythmic activity. Triggered arrhythmic activity during ischemia is based on changes in cellular ionic balance and the cells Ca(2+) handling properties. In the current study we used isolated mouse ventricular myocytes (VMs) deficient for the expression of Pak1 (Pak1(-/-)) to determine the mechanism by which Pak1 influences the generation of arrhythmic activity during simulated ischemia. The Ca(2+) transient amplitude and kinetics did not significantly change in wild type (WT) and Pak1(-/-) VMs during 15 min of simulated ischemia. However, Pak1(-/-) VMs exhibited an exaggerated increase in [Ca(2+)]i, which resulted in spontaneous Ca(2+) release events and waves. The Ca(2+) overload in Pak1(-/-) VMs could be suppressed with a reverse mode blocker (KB-R7943) of the sodium calcium exchanger (NCX), a cytoplasmic scavenger of reactive oxygen species (ROS; TEMPOL) or a RAC1 inhibitor (NSC23766). Measurements of the cytoplasmic ROS levels revealed that decreased Pak1 activity in Pak1(-/-) VMs or VMs treated with the Pak1 inhibitor (IPA3) enhanced cellular ROS production. The Pak1 dependent increase in ROS was attenuated in VMs deficient for NADPH oxidase 2 (NOX2; p47(phox-/-)) or in VMs where NOX2 was inhibited (gp91ds-tat). Voltage clamp recordings showed increased NCX activity in Pak1(-/-) VMs that depended on enhanced NOX2 induced ROS production. The exaggerated Ca(2+) overload in Pak1(-/-) VMs could be mimicked by low concentrations of ouabain. Overall our data show that Pak1 is a critical negative regulator of NOX2 dependent ROS production and that a latent ROS dependent stimulation of NCX activity can predispose VMs to Ca(2+) overload under conditions where no significant changes in excitation-contraction coupling are yet evident. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. p21-activated kinase1 (Pak1) is a negative regulator of NADPH-oxidase 2 in ventricular myocytes

    PubMed Central

    DeSantiago, Jaime; Bare, Dan J; Xiao, Lei; Ke, Yunbo; Solaro, R. John; Banach, Kathrin

    2014-01-01

    Ischemic conditions reduce the activity of the p21-activated kinase (Pak1) resulting in increased arrhythmic activity. Triggered arrhythmic activity during ischemia is based on changes in cellular ionic balance and the cells Ca2+ handling properties. In the current study we used isolated mouse ventricular myocytes (VMs) deficient for the expression of Pak1 (Pak1-/-) to determine the mechanism by which Pak1 influences the generation of arrhythmic activity during simulated ischemia. The Ca2+ transient amplitude and kinetics did not significantly change in wild type (WT) and Pak1-/- VMs during 15 min of simulated ischemia. However, Pak1-/- VMs exhibited an exaggerated increase in [Ca2+]i, which resulted in spontaneous Ca2+ release events and waves. The Ca2+ overload in Pak1-/- VMs could be suppressed with a reverse mode blocker (KB-R7943) of the sodium calcium exchanger (NCX), a cytoplasmic scavenger of reactive oxygen species (ROS; TEMPOL) or a RAC1 inhibitor (NSC23766). Measurements of the cytoplasmic ROS levels revealed that decreased Pak1 activity in Pak1-/- VMs or VMs treated with the Pak1 inhibitor (IPA3) enhanced cellular ROS production. The Pak1 dependent increase in ROS was attenuated in VMs deficient for NADPH oxidase 2 (NOX2; p47phox-/-) or in VMs where NOX2 was inhibited (gp91ds-tat). Voltage clamp recordings showed increased NCX activity in Pak1-/- VMs that depended on enhanced NOX2 induced ROS production. The exaggerated Ca2+ overload in Pak1-/- VMs could be mimicked by low concentrations of ouabain. Overall our data show that Pak1 is a critical negative regulator of NOX2 dependent ROS production and that a latent ROS dependent stimulation of NCX activity can predispose VMs to Ca2+ overload under conditions where no significant changes in excitation-contraction coupling are yet evident. PMID:24380729

  16. Early Results from the HexPak and GradPak Variable-Scale Dual-Head IFUs on the WIYN 3.5-meter Telescope

    NASA Astrophysics Data System (ADS)

    Hooper, Eric; Bershady, Matthew A.; Eigenbrot, Arthur; Wood, Corey M.; Buckley, Scott; Smith, Michael; Corson, Charles; Wolf, Marsha J.; Zhu, Guanying Y.; Vang, Andrea; Gallagher, John S.; Sheinis, Andrew; Washburn Astronomical Laboratories

    2015-01-01

    The WIYN Observatory recently installed two new integral field units (IFUs) on its 3.5-meter telescope on Kitt Peak, Arizona. Each IFU is unique in that it contains different sized fibers in the same head to optimize the tradeoff between spatial resolution and surface brightness sensitivity for observations of galaxies. These instruments were designed and constructed (M. Bershady, PI) at the University of Wisconsin's Washburn Astronomical Laboratory. HexPak, with a central core of 1 arcsec fibers surrounded by a halo of 3 arcsec fibers, was designed for early type, face-on disk, and quasar host galaxies. GradPak, with a series of rows of fibers of increasing diameter from 2 arcsec to 6 arcsec (5 different diameters total), was designed for edge-on galaxies, where the small fibers lie along the midplane and larger fibers sample the progressively lower surface brightnesses above the plane. The instruments were installed alongside the existing SparsePak IFU in late 2013 and have been used in several observing runs since. The different fiber sizes present additional data reduction challenges, particularly regarding flux calibration and sky subtraction. Early results on studies of the stellar populations of galaxies are quite promising and demonstrate the advantages of fiber sizes tailored to the objects under study. HexPak and GradPak were built with funds from NSF award ATI-0804576.

  17. PAK6 targets to cell–cell adhesions through its N-terminus in a Cdc42-dependent manner to drive epithelial colony escape

    PubMed Central

    Morse, Elizabeth M.; Sun, Xiaowen; Olberding, Jordan R.; Ha, Byung Hak; Boggon, Titus J.; Calderwood, David A.

    2016-01-01

    ABSTRACT The six serine/threonine kinases in the p21-activated kinase (PAK) family are important regulators of cell adhesion, motility and survival. PAK6, which is overexpressed in prostate cancer, was recently reported to localize to cell–cell adhesions and to drive epithelial cell colony escape. Here we report that PAK6 targeting to cell–cell adhesions occurs through its N-terminus, requiring both its Cdc42/Rac interactive binding (CRIB) domain and an adjacent polybasic region for maximal targeting efficiency. We find PAK6 localization to cell–cell adhesions is Cdc42-dependent, as Cdc42 knockdown inhibits PAK6 targeting to cell–cell adhesions. We further find the ability of PAK6 to drive epithelial cell colony escape requires kinase activity and is disrupted by mutations that perturb PAK6 cell–cell adhesion targeting. Finally, we demonstrate that all type II PAKs (PAK4, PAK5 and PAK6) target to cell–cell adhesions, albeit to differing extents, but PAK1 (a type I PAK) does not. Notably, the ability of a PAK isoform to drive epithelial colony escape correlates with its targeting to cell–cell adhesions. We conclude that PAKs have a broader role in the regulation of cell–cell adhesions than previously appreciated. PMID:26598554

  18. Regulation of KRAS-PAK4 axis by microRNAs in cancer.

    PubMed

    Choudhry, Zeshan S; Tripathi, Vraj; Sutton, Mike; Bao, Bin; Mohammad, Ramzi M; Azmi, Asfar S

    2014-01-01

    MicroRNAs (miRNAs), often aberrantly expressed in cancer, have been implicated in the regulation of a number of critical cell survival pathways including the genes in the Kras signaling. Kras mutations are observed in more than half of cancers and its inhibition has been the focus of intense research for the past 30 years. However, Kras itself has proven to be non-druggable due in part to the absence of binding pockets for small molecule drugs. These hurdles resulted in researchers shifting their focus on targeting proteins downstream to Kras pathways. P21 activated kinase 4 (PAK4) belongs to the family of serine/threonine kinases comprising of 6 isoforms (PAK 1-6) and is considered as a key effector of Rho family of GTPases downstream of RAS. PAK4 controls critical processes such as cellular motility, proliferation and survival. Recently a number of small molecule PAK4 antagonists have been investigated in preclinical and clinical setting; albeit without any success. Emerging evidence shows that PAK is tightly regulated by a number of miRNAs that are also recognized to promote hyper-activation of oncogenic Kras signaling. Therefore, the understanding of the role of miRNAs in the regulation of PAK4 is critical to the development of therapies against this important player in the Kras pathway. Through this review, we bring forward mechanistic insights on PAK4 regulation by aberrantly expressed miRNAs in cancer and its implications on Kras signaling. We anticipate that enhanced knowledge of the miRNA-PAK4 interaction network will allow the development of successful therapies targeting this critical protein to ultimately rein in Kras.

  19. Relationship Between Pak-Mediated Cell Death and Stress-Activated Kinase Signaling in Breast Cancer

    DTIC Science & Technology

    2000-02-01

    part of the cell death execution machinery. Here we show that a correlation exists in breast cancer cells between caspase- dependent cleavage of the...inhibits its activity might allow us to specifically inhibit signaling pathways downstream of Pak and evaluate how the cell death process is affected. In...a biochemical approach screening for substrates and possible mediators of cell death signaling components via Pak kinases we identified a guanine

  20. Interaction Between a Novel p21 Activated Kinase (PAK6) and Androgen Receptor in Prostate Cancer

    DTIC Science & Technology

    2005-02-01

    cancer. The cell cycle signaling regulated by the mitogen activated protein/extracellular-signal-regulated kinase (MAPK/ERK) have been linked to tumor...biological roles of PAK6 in prostate cancer cells , and to examine the expression of PAK6 in prostate tissues. We anticipate that by completing the...of prostate cells (Balk, 2002; Gelmann, 2002). Androgen ablation is an effective treatment for the majority of advanced prostate cancer patients

  1. A RAC/CDC-42-independent GIT/PIX/PAK signaling pathway mediates cell migration in C. elegans.

    PubMed

    Lucanic, Mark; Cheng, Hwai-Jong

    2008-11-01

    P21 activated kinase (PAK), PAK interacting exchange factor (PIX), and G protein coupled receptor kinase interactor (GIT) compose a highly conserved signaling module controlling cell migrations, immune system signaling, and the formation of the mammalian nervous system. Traditionally, this signaling module is thought to facilitate the function of RAC and CDC-42 GTPases by allowing for the recruitment of a GTPase effector (PAK), a GTPase activator (PIX), and a scaffolding protein (GIT) as a regulated signaling unit to specific subcellular locations. Instead, we report here that this signaling module functions independently of RAC/CDC-42 GTPases in vivo to control the cell shape and migration of the distal tip cells (DTCs) during morphogenesis of the Caenorhabditis elegans gonad. In addition, this RAC/CDC-42-independent PAK pathway functions in parallel to a classical GTPase/PAK pathway to control the guidance aspect of DTC migration. Among the C. elegans PAKs, only PAK-1 functions in the GIT/PIX/PAK pathway independently of RAC/CDC42 GTPases, while both PAK-1 and MAX-2 are redundantly utilized in the GTPase/PAK pathway. Both RAC/CDC42-dependent and -independent PAK pathways function with the integrin receptors, suggesting that signaling through integrins can control the morphology, movement, and guidance of DTC through discrete pathways. Collectively, our results define a new signaling capacity for the GIT/PIX/PAK module that is likely to be conserved in vertebrates and demonstrate that PAK family members, which are redundantly utilized as GTPase effectors, can act non-redundantly in pathways independent of these GTPases.

  2. p21-activated kinase (Pak) regulates NADPH oxidase activation in human neutrophils

    PubMed Central

    Martyn, Kendra D.; Kim, Moon-Ju; Quinn, Mark T.; Dinauer, Mary C.; Knaus, Ulla G.

    2005-01-01

    The phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase plays an instrumental role in host defense and contributes to microbicial killing by releasing highly reactive oxygen species. This multicomponent enzyme is composed of membrane and cytosolic components that assemble in the plasma membrane or phagolysosome. While the guanosine S′-triphosphatase (GTPase) Rac2 has been shown to be a critical regulator of NADPH oxidase activity and assembly, the role of its effector, p21-activated kinase (Pak), in oxidase function has not been well defined. Using HIV-1 Tat-mediated protein transduction of Pak inhibitory domain, we show here that Pak activity is indeed required for efficient superoxide generation in intact neutrophils. Furthermore, we show that Pak translocates to the plasma membrane upon N-formyl-methionyl-leucyl-phenylalanine (fMLF) stimulation and colocalizes with translocated p47phox and with p22phox, a subunit of flavocytochrome b558. Although activated Pak phosphorylated several essential serine residues in the C-terminus of p47phox, direct binding to p47phox was not observed. In contrast, active Pak bound directly to p22phox, suggesting flavocytochrome b was the oxidase-associated membrane target of this kinase and this association may facilitate further phosphorylation of p47phox in the assembling NADPH oxidase complex. PMID:16099876

  3. Reduced PAK1 activity sensitizes FA/BRCA-proficient breast cancer cells to PARP inhibition.

    PubMed

    Villamar Cruz, Olga; Prudnikova, Tatiana Y; Araiza-Olivera, Daniela; Perez-Plasencia, Carlos; Johnson, Neil; Bernhardy, Andrea J; Slifker, Michael; Renner, Catherine; Chernoff, Jonathan; Arias-Romero, Luis E

    2016-11-22

    Cells that are deficient in homologous recombination, such as those that have mutations in any of the Fanconi Anemia (FA)/BRCA genes, are hypersensitive to inhibition of poly(ADP-ribose) polymerase (PARP). However, FA/BRCA-deficient tumors represent a small fraction of breast cancers, which might restrict the therapeutic utility of PARP inhibitor monotherapy. The gene encoding the serine-threonine protein kinase p21-activated kinase 1 (PAK1) is amplified and/or overexpressed in several human cancer types including 25-30% of breast tumors. This enzyme controls many cellular processes by phosphorylating both cytoplasmic and nuclear substrates. Here, we show that depletion or pharmacological inhibition of PAK1 down-regulated the expression of genes involved in the FA/BRCA pathway and compromised the ability of cells to repair DNA by Homologous Recombination (HR), promoting apoptosis and reducing colony formation. Combined inhibition of PAK1 and PARP in PAK1 overexpressing breast cancer cells had a synergistic effect, enhancing apoptosis, suppressing colony formation, and delaying tumor growth in a xenograft setting. Because reduced PAK1 activity impaired FA/BRCA function, inhibition of this kinase in PAK1 amplified and/or overexpressing breast cancer cells represents a plausible strategy for expanding the utility of PARP inhibitors to FA/BRCA-proficient cancers.

  4. PAK1 negatively regulates the activity of the Rho exchange factor NET1.

    PubMed

    Alberts, Arthur S; Qin, Huajun; Carr, Heather S; Frost, Jeffrey A

    2005-04-01

    Rho family small G-protein activity is controlled by guanine nucleotide exchange factors that stimulate the release of GDP, thus allowing GTP binding. Once activated, Rho proteins control cell signaling through interactions with downstream effector proteins, leading to changes in cytoskeletal organization and gene expression. The ability of Rho family members to modulate the activity of other Rho proteins is also intrinsic to these processes. In this work we show that the Rac/Cdc42hs-regulated protein kinase PAK1 down-regulates the activity of the RhoA-specific guanine nucleotide exchange factor NET1. Specifically, PAK1 phosphorylates NET1 on three sites in vitro: serines 152, 153, and 538. Replacement of serines 152 and 153 with glutamate residues down-regulates the activity of NET1 as an exchange factor in vitro and its ability to stimulate actin stress fiber formation in cells. Using a phospho-specific antibody that recognizes NET1 phosphorylated on serine 152, we show that PAK1 phosphorylates NET1 on this site in cells and that Rac1 stimulates serine 152 phosphorylation in a PAK1-dependent manner. Furthermore, coexpression of constitutively active PAK1 inhibits the ability of NET1 to stimulate actin polymerization only when serines 152 and 153 are present. These data provide a novel mechanism for the control of RhoA activity by Rac1 through the PAK-dependent phosphorylation of NET1 to reduce its activity as a guanine nucleotide exchange factor.

  5. Reduced PAK1 activity sensitizes FA/BRCA-proficient breast cancer cells to PARP inhibition

    PubMed Central

    Araiza-Olivera, Daniela; Perez-Plasencia, Carlos; Johnson, Neil; Bernhardy, Andrea J.; Slifker, Michael; Renner, Catherine; Chernoff, Jonathan; Arias, Luis E.

    2016-01-01

    Cells that are deficient in homologous recombination, such as those that have mutations in any of the Fanconi Anemia (FA)/BRCA genes, are hypersensitive to inhibition of poly(ADP-ribose) polymerase (PARP). However, FA/BRCA-deficient tumors represent a small fraction of breast cancers, which might restrict the therapeutic utility of PARP inhibitor monotherapy. The gene encoding the serine-threonine protein kinase p21-activated kinase 1 (PAK1) is amplified and/or overexpressed in several human cancer types including 25-30% of breast tumors. This enzyme controls many cellular processes by phosphorylating both cytoplasmic and nuclear substrates. Here, we show that depletion or pharmacological inhibition of PAK1 down-regulated the expression of genes involved in the FA/BRCA pathway and compromised the ability of cells to repair DNA by Homologous Recombination (HR), promoting apoptosis and reducing colony formation. Combined inhibition of PAK1 and PARP in PAK1 overexpressing breast cancer cells had a synergistic effect, enhancing apoptosis, suppressing colony formation, and delaying tumor growth in a xenograft setting. Because reduced PAK1 activity impaired FA/BRCA function, inhibition of this kinase in PAK1 amplified and/or overexpressing breast cancer cells represents a plausible strategy for expanding the utility of PARP inhibitors to FA/BRCA-proficient cancers. PMID:27740936

  6. PAKs supplement improves immune status and body composition but not muscle strength in resistance trained individuals

    PubMed Central

    2010-01-01

    Mixed formula supplements are very popular among recreational and professional weightlifters. They are usually known as PAKs and they are supposed to have a synergistic effect of their different nutrients. The purpose of this study was to determine the effects of chronic (4 weeks) PAKS supplementation in combination with strength training on body composition, immune status and performance measures in recreationally trained individuals with or without PAKs supplementation. Methods: Twelve male subjects (Placebo n = 6 and PAKs supplement n = 6) were recruited for this study. The body composition, one maximum strength repetition tests and immune status were assessed before and after 4 week supplementation. Our data showed that, 4 week PAK supplementation associated with strength exercise not was effective in change strength than compared with placebo group. However, we observed that, PAK supplement was able to improve immune status and reduced body composition when compared with placebo group. These results indicate that, a mixed formula supplement is able to improve immune status and body composition but not maximum strength in recreational strength trained subjects in a 4 weeks period. PMID:21059194

  7. Group I Paks as therapeutic targets in NF2-deficient meningioma

    PubMed Central

    Duron, Sergio G.; Campbell, David A.; Ong, Christy C.; Hoeflich, Klaus P.; Chang, Long-Sheng; Welling, D. Bradley; Yang, Zeng-jie; Chernoff, Jonathan

    2015-01-01

    Neurofibromatosis type 2 (NF2) is an autosomal dominant disorder characterized by the development of multiple tumors in the central nervous system, most notably schwannomas and meningiomas. Mutational inactivation of NF2 is found in 40–60% of sporadic meningiomas, but the molecular mechanisms underlying malignant changes of meningioma cells remain unclear. Because group I p21-activated kinases (Paks) bind to and are inhibited by the NF2-encoded protein Merlin, we assessed the signaling and anti-tumor effects of three group-I specific Pak inhibitors - Frax597, 716 and 1036 - in NF2−/− meningiomas in vitro and in an orthotopic mouse model. We found that these Pak inhibitors suppressed the proliferation and motility of both benign (Ben-Men1) and malignant (KT21-MG1) meningiomas cells. In addition, we found a strong reduction in phosphorylation of Mek and S6, and decreased cyclin D1 expression in both cell lines after treatment with Pak inhibitors. Using intracranial xenografts of luciferase-expressing KT21-MG1 cells, we found that treated mice showed significant tumor suppression for all three Pak inhibitors. Similar effects were observed in Ben-Men1 cells. Tumors dissected from treated animals exhibited an increase in apoptosis without notable change in proliferation. Collectively, these results suggest that Pak inhibitors might be useful agents in treating NF2-deficient meningiomas. PMID:25596744

  8. PAK is regulated by PI3K, PIX, CDC42, and PP2Calpha and mediates focal adhesion turnover in the hyperosmotic stress-induced p38 pathway.

    PubMed

    Chan, Perry M; Lim, Louis; Manser, Edward

    2008-09-05

    Fractionation of brain extracts and functional biochemical assays identified PP2Calpha, a serine/threonine phosphatase, as the major biochemical activity inhibiting PAK1. PP2Calpha dephosphorylated PAK1 and p38, both of which were activated upon hyperosmotic shock with the same kinetics. In comparison to growth factors, hyperosmolality was a more potent activator of PAK1. Therefore we characterize the PAK signaling pathway in the hyperosmotic shock response. Endogenous PAKs were recruited to the p38 kinase complex in a phosphorylation-dependent manner. Overexpression of a PAK inhibitory peptide or dominant negative Cdc42 revealed that p38 activation was dependent on PAK and Cdc42 activities. PAK mutants deficient in binding to Cdc42 or PAK-interacting exchange factor were not activated. Using a panel of kinase inhibitors, we identified PI3K acting upstream of PAK, which correlated with PAK repression by pTEN overexpression. RNA interference knockdown of PAK expression reduced stress-induced p38 activation and conversely, PP2Calpha knockdown increased its activation. Hyperosmotic stress-induced PAK translocation away from focal adhesions to the perinuclear compartment and resulted in disassembly of focal adhesions, which are hallmarks of PAK activation. Inhibition of PAK by overexpression of PP2Calpha or the kinase inhibitory domain prevented sorbitol-induced focal adhesion dissolution. Inhibition of MAPK pathways showed that MEK-ERK signaling but not p38 is required for full PAK activation and focal adhesion turnover. We conclude that 1) PAK plays a required role in hyperosmotic signaling through the PI3K/pTEN/Cdc42/PP2Calpha/p38 pathway, and 2) PAK and PP2Calpha modulate the effects of this pathway on focal adhesion dynamics.

  9. Comparison of the SidePak personal monitor with the Aerosol Particle Sizer (APS).

    PubMed

    Sánchez Jiménez, Araceli; van Tongeren, Martie; Galea, Karen S; Steinsvåg, Kjersti; MacCalman, Laura; Cherrie, John W

    2011-06-01

    The aim of this study was to compare the performance of the TSI Aerodynamic Particle Sizer (APS) and the TSI portable photometer SidePak to measure airborne oil mist particulate matter (PM) with aerodynamic diameters below 10 μm, 2.5 μm and 1 μm (PM(10), PM(2.5) and PM(1)). Three SidePaks each fitted with either a PM(10), PM(2.5) or a PM(1) impactor and an APS were run side by side in a controlled chamber. Oil mist from two different mineral oils and two different drilling fluid systems commonly used in offshore drilling technologies were generated using a nebulizer. Compared to the APS, the SidePaks overestimated the concentration of PM(10) and PM(2.5) by one order of magnitude and PM(1) concentrations by two orders of magnitude after exposure to oil mist for 3.3-6.5 min at concentrations ranging from 0.003 to 18.1 mg m(-3) for PM(10), 0.002 to 3.96 mg m(-3) for PM(2.5) and 0.001 to 0.418 mg m(-3) for PM(1) (as measured by the APS). In a second experiment a SidePak monitor previously exposed to oil mist overestimated PM(10) concentrations by 27% compared to measurements from another SidePak never exposed to oil mist. This could be a result of condensation of oil mist droplets in the optical system of the SidePak. The SidePak is a very useful instrument for personal monitoring in occupational hygiene due to its light weight and quiet pump. However, it may not be suitable for the measurement of particle concentrations from oil mist.

  10. New realism in north Korean propaganda: The death of Pak Chong-ch`ol. Final report

    SciTech Connect

    Shaw, W.

    1987-04-01

    North Korea`s Voice of National Salvation (VNS) broadcasts to South Korea in January and February gave considerable attention to the 13 January 1987 death under police torture of Seoul National University student Pak Chong-ch`ol. VNS commentary emphasized the routine nature of police torture in South Korea and sharply criticized the South Korean Government for blocking attendance at memorial services on 7 February and 3 March. Many VNS comments on the Pak case closely paralleled reactions of the South Korean press and parliamentary opposition, demonstrating close North Korean attention to opinion trends in the South. In a new mood of realism, P`yongyang also used the Pak case to urge South Korean radical students to drop their `avant-gardism`, including excessively leftist slogans and violent tactics that alienate the general populace. P` yongyang clearly wants students to take advantage of issues that have wide public appeal and to broaden resistance to the Chon Tu-hwan Government to include social groups beyond the student population. In P`yongyang`s view, such issues include the Pak case and the signature campaign for constitutional revision in early 1986. If South Korea`s radical students take P`yonyyang`s advice, and if the Republic of Korea Government fails to curb and punish abuses like the Pak killing, this shift in tactics could have significant influence during the coming spring demonstration season.

  11. Inhibition of Nischarin Expression Promotes Neurite Outgrowth through Regulation of PAK Activity

    PubMed Central

    Ding, Yuemin; Li, Yuying; Lu, Lingchao; Zhang, Ruyi; Zeng, Linghui; Wang, Linlin; Zhang, Xiong

    2015-01-01

    Nischarin is a cytoplasmic protein expressed in various organs that plays an inhibitory role in cell migration and invasion and the carcinogenesis of breast cancer cells. We previously reported that Nischarin is highly expressed in neuronal cell lines and is differentially expressed in the brain tissue of adult rats. However, the physiological function of Nischarin in neural cells remains unknown. Here, we show that Nischarin is expressed in rat primary cortical neurons but not in astrocytes. Nischarin is localized around the nucleus and dendrites. Using shRNA to knockdown the expression of endogenous Nischarin significantly increases the percentage of neurite-bearing cells, remarkably increases neurite length, and accelerates neurite extension in neuronal cells. Silencing Nischarin expression also promotes dendrite elongation in rat cortical neurons where Nischarin interacts with p21-activated kinase 1/2 (PAK1/2) and negatively regulates phosphorylation of both PAK1 and PAK2. The stimulation of neurite growth observed in cells with decreased levels of Nischarin is partially abolished by IPA3-mediated inhibition of PAK1 activity. Our findings indicate that endogenous Nischarin inhibits neurite outgrowth by blocking PAK1 activation in neurons. PMID:26670864

  12. A GIT1/PIX/Rac/PAK signaling module regulates spine morphogenesis and synapse formation through MLC.

    PubMed

    Zhang, Huaye; Webb, Donna J; Asmussen, Hannelore; Niu, Shuang; Horwitz, Alan F

    2005-03-30

    Three of seven recently identified genes mutated in nonsyndromic mental retardation are involved in Rho family signaling. Two of the gene products, alpha-p-21-activated kinase (PAK) interacting exchange factor (alphaPIX) and PAK3, form a complex with the synaptic adaptor protein G-protein-coupled receptor kinase-interacting protein 1 (GIT1). Using an RNA interference approach, we show that GIT1 is critical for spine and synapse formation. We also show that Rac is locally activated in dendritic spines using fluorescence resonance energy transfer. This local activation of Rac is regulated by PIX, a Rac guanine nucleotide exchange factor. PAK1 and PAK3 serve as downstream effectors of Rac in regulating spine and synapse formation. Active PAK promotes the formation of spines and dendritic protrusions, which correlates with an increase in the number of excitatory synapses. These effects are dependent on the kinase activity of PAK, and PAK functions through phosphorylating myosin II regulatory light chain (MLC). Activated MLC causes an increase in dendritic spine and synapse formation, whereas inhibiting myosin ATPase activity results in decreased spine and synapse formation. Finally, both activated PAK and activated MLC can rescue the defects of GIT1 knockdown, suggesting that PAK and MLC are downstream of GIT1 in regulating spine and synapse formation. Our results point to a signaling complex, consisting of GIT1, PIX, Rac, and PAK, that plays an essential role in the regulation of dendritic spine and synapse formation and provides a potential mechanism by which alphaPIX and PAK3 mutations affect cognitive functions in mental retardation.

  13. PAK Inactivation Impairs Social Recognition in 3xTg-AD Mice without Increasing Brain Deposition of Tau and Aβ

    PubMed Central

    Arsenault, Dany; Dal-Pan, Alexandre; Tremblay, Cyntia; Bennett, David A.; Guitton, Matthieu J.; De Koninck, Yves; Tonegawa, Susumu

    2013-01-01

    Defects in p21-activated kinase (PAK) are suspected to play a role in cognitive symptoms of Alzheimer's disease (AD). Dysfunction in PAK leads to cofilin activation, drebrin displacement from its actin-binding site, actin depolymerization/severing, and, ultimately, defects in spine dynamics and cognitive impairment in mice. To determine the role of PAK in AD, we first quantified PAK by immunoblotting in homogenates from the parietal neocortex of subjects with a clinical diagnosis of no cognitive impairment (n = 12), mild cognitive impairment (n = 12), or AD (n = 12). A loss of total PAK, detected in the cortex of AD patients (−39% versus controls), was correlated with cognitive impairment (r2 = 0.148, p = 0.027) and deposition of total and phosphorylated tau (r2 = 0.235 and r2 = 0.206, respectively), but not with Aβ42 (r2 = 0.056). Accordingly, we found a decrease of total PAK in the cortex of 12- and 20-month-old 3xTg-AD mice, an animal model of AD-like Aβ and tau neuropathologies. To determine whether PAK dysfunction aggravates AD phenotype, 3xTg-AD mice were crossed with dominant-negative PAK mice. PAK inactivation led to obliteration of social recognition in old 3xTg-AD mice, which was associated with a decrease in cortical drebrin (−25%), but without enhancement of Aβ/tau pathology or any clear electrophysiological signature. Overall, our data suggest that PAK decrease is a consequence of AD neuropathology and that therapeutic activation of PAK may exert symptomatic benefits on high brain function. PMID:23804095

  14. Remarkable reductions of PAKs in the brain tissues of scrapie-infected rodent possibly linked closely with neuron loss.

    PubMed

    Meng, Ge; Tian, Chan; Wang, Hui; Xu, Yin; Zhang, Bao-Yun; Shi, Qi; Gao, Chen; Chen, Cao; Fan, Xue-Yu; Wang, Jing; Xiao, Kang; Ren, Ke; Xue, Ming-Ming; Dong, Xiao-Ping

    2014-10-01

    Prion diseases are irreversible progressive neurodegenerative diseases characterized in the brain by PrP(Sc) deposits, neuronal degeneration, gliosis and by cognitive, behavioral and physical impairments, leading to severe incapacity and inevitable death. Proteins of the p21-activated kinase (PAK) family are noted for roles in gene transcription, cytoskeletal dynamics, cell cycle progression and survival signaling. In the present study, we aimed to identify the potential roles of PAKs during prion infection, utilizing the brains of scrapie agent-infected hamsters. Western blots and immunohistochemical assays showed that brain levels of PAK3 and PAK1, as well as their upstream activator Rac/cdc42 and downstream substrate Raf1, were remarkably reduced at terminal stage. Double-stained immunofluorescent assay demonstrated that PAK3 was expressed mainly in neurons. Dynamic analyses of the brain samples collected at the different time points during the incubation period illustrated successive decreases of PAK3, PAK1 and Raf1, especially phosphor Raf1, which correlated well with neuron loss. Rac/cdc42 in the brain tissues increased at early stage and reached to the top at mid-late stage, but diminished at final stage. Unlike the alteration of PAKs in vivo, PAK3 and PAK1, as well as Rac/cdc42 and Raf1 in the prion-infected cell line SMB-S15 remained unchanged compared with those of its normal cell line SMB-PS. Our data here indicate that the functions of PAKs and their associated signaling pathways are seriously affected in the brains of prion disease, which appear to associate closely with the extensive neuron loss.

  15. Viekira Pak Induced Fatal Lactic Acidosis: A Case Report of an Unusual Side Effect

    PubMed Central

    2016-01-01

    Viekira Pak is a new direct-acting antiviral agent that has an excellent efficacy in treating patients with chronic HCV. FDA released a safety warning that Viekira Pak can cause serious liver injury mostly in patients with underlying advanced liver disease. We report the first case of fatal lactic acidosis presenting 3 days after initiating therapy with Viekira Pak. Although it is very hard to precisely determine the cause of lactic acidosis, our case highlights an unusual side effect that ensued after starting the medication. Given the complexity of drug-drug interactions that can happen with the new direct-acting antiviral agents and the paucity of data regarding coadministration and methods of monitoring, a thorough review should be pursued prior to initiating these medications. PMID:28044114

  16. Glucosinolates from pak choi and broccoli induce enzymes and inhibit inflammation and colon cancer differently.

    PubMed

    Lippmann, Doris; Lehmann, Carsten; Florian, Simone; Barknowitz, Gitte; Haack, Michael; Mewis, Inga; Wiesner, Melanie; Schreiner, Monika; Glatt, Hansruedi; Brigelius-Flohé, Regina; Kipp, Anna P

    2014-06-01

    High consumption of Brassica vegetables is considered to prevent especially colon carcinogenesis. The content and pattern of glucosinolates (GSLs) can highly vary among different Brassica vegetables and may, thus, affect the outcome of Brassica intervention studies. Therefore, we aimed to feed mice with diets containing plant materials of the Brassica vegetables broccoli and pak choi. Further enrichment of the diets by adding GSL extracts allowed us to analyze the impact of different amounts (GSL-poor versus GSL-rich) and different patterns (broccoli versus pak choi) of GSLs on inflammation and tumor development in a model of inflammation-triggered colon carcinogenesis (AOM/DSS model). Serum albumin adducts were analyzed to confirm the up-take and bioactivation of GSLs after feeding the Brassica diets for four weeks. In agreement with their high glucoraphanin content, broccoli diets induced the formation of sulforaphane-lysine adducts. Levels of 1-methoxyindolyl-3-methyl-histidine adducts derived from neoglucobrassicin were the highest in the GSL-rich pak choi group. In the colon, the GSL-rich broccoli and the GSL-rich pak choi diet up-regulated the expression of different sets of typical Nrf2 target genes like Nqo1, Gstm1, Srxn1, and GPx2. GSL-rich pak choi induced the AhR target gene Cyp1a1 but did not affect Ugt1a1 expression. Both colitis and tumor number were drastically reduced after feeding the GSL-rich pak choi diet while the other three diets had no effect. GSLs can act anti-inflammatory and anti-carcinogenic but both effects depend on the specific amount and pattern of GSLs within a vegetable. Thus, a high Brassica consumption cannot be generally considered to be cancer-preventive.

  17. Fission yeast pak1+ encodes a protein kinase that interacts with Cdc42p and is involved in the control of cell polarity and mating.

    PubMed Central

    Ottilie, S; Miller, P J; Johnson, D I; Creasy, C L; Sells, M A; Bagrodia, S; Forsburg, S L; Chernoff, J

    1995-01-01

    A STE20/p65pak homolog was isolated from fission yeast by PCR. The pak1+ gene encodes a 72 kDa protein containing a putative p21-binding domain near its amino-terminus and a serine/threonine kinase domain near its carboxyl-terminus. The Pak1 protein autophosphorylates on serine residues and preferentially binds to activated Cdc42p both in vitro and in vivo. This binding is mediated through the p21 binding domain on Pak1p and the effector domain on Cdc42p. Overexpression of an inactive mutant form of pak1 gives rise to cells with markedly abnormal shape with mislocalized actin staining. Pak1 overexpression does not, however, suppress lethality associated with cdc42-null cells or the morphologic defeat caused by overexpression of mutant cdc42 alleles. Gene disruption of pak1+ establishes that, like cdc42+, pak1+ function is required for cell viability. In budding yeast, pak1+ expression restores mating function to STE20-null cells and, in fission yeast, overexpression of an inactive form of Pak inhibits mating. These results indicate that the Pak1 protein is likely to be an effector for Cdc42p or a related GTPase, and suggest that Pak1p is involved in the maintenance of cell polarity and in mating. Images PMID:8846783

  18. Cadherins and Pak1 control contact inhibition of proliferation by Pak1-betaPIX-GIT complex-dependent regulation of cell-matrix signaling.

    PubMed

    Liu, Fengming; Jia, Liwei; Thompson-Baine, Ann-Marie; Puglise, Jason M; Ter Beest, Martin B A; Zegers, Mirjam M P

    2010-04-01

    It is crucial for organ homeostasis that epithelia have effective mechanisms to restrict motility and cell proliferation in order to maintain tissue architecture. On the other hand, epithelial cells need to rapidly and transiently acquire a more mesenchymal phenotype, with high levels of cell motility and proliferation, in order to repair epithelia upon injury. Cross talk between cell-cell and cell-matrix signaling is crucial for regulating these transitions. The Pak1-betaPIX-GIT complex is an effector complex downstream of the small GTPase Rac1. We previously showed that translocation of this complex from cell-matrix to cell-cell adhesion sites was required for the establishment of contact inhibition of proliferation. In this study, we provide evidence that this translocation depends on cadherin function. Cadherins do not recruit the complex by direct interaction. Rather, we found that inhibition of the normal function of cadherin or Pak1 leads to defects in focal adhesion turnover and to increased signaling by phosphatidylinositol 3-kinase. We propose that cadherins are involved in regulation of contact inhibition by controlling the function of the Pak1-betaPIX-GIT complex at focal contacts.

  19. US EPA, Pesticide Product Label, CUSTOM-PAK HI-CIDE ...

    EPA Pesticide Factsheets

    2011-04-21

    ... I • I • • • • • I • • • I . • Manufactured by: CUSTOM-PAK, INC. 14800 Miles Avenue • Cleveland. ()hio 44128 I , • , • • I • • , • • I • . , II • • • • • ...

  20. Pak1 regulates the orientation of apical polarization and lumen formation by distinct pathways.

    PubMed

    deLeon, Orlando; Puglise, Jason M; Liu, Fengming; Smits, Jos; ter Beest, Martin B; Zegers, Mirjam M

    2012-01-01

    The development of the basic architecture of branching tubules enclosing a central lumen that characterizes most epithelial organs crucially depends on the apico-basolateral polarization of epithelial cells. Signals from the extracellular matrix control the orientation of the apical surface, so that it faces the lumen interior, opposite to cell-matrix adhesion sites. This orientation of the apical surface is thought to be intrinsically linked to the formation of single lumens. We previously demonstrated in three-dimensional cyst cultures of Madin-Darby canine kidney (MDCK) cells that signaling by β1 integrins regulates the orientation of the apical surface, via a mechanism that depends on the activity of the small GTPase Rac1. Here, we investigated whether the Rac1 effector Pak1 is a downstream effector in this pathway. Expression of constitutive active Pak1 phenocopies the effect of β1 integrin inhibition in that it misorients the apical surface and induces a multilumen phenotype. The misorientation of apical surfaces depends on the interaction of active Pak1 with PIX proteins and is linked to defects in basement membrane assembly. In contrast, the multilumen phenotype was independent of PIX and the basement membrane. Therefore, Pak1 likely regulates apical polarization and lumen formation by two distinct pathways.

  1. Pak1 Regulates the Orientation of Apical Polarization and Lumen Formation by Distinct Pathways

    PubMed Central

    Smits, Jos; ter Beest, Martin B.; Zegers, Mirjam M.

    2012-01-01

    The development of the basic architecture of branching tubules enclosing a central lumen that characterizes most epithelial organs crucially depends on the apico-basolateral polarization of epithelial cells. Signals from the extracellular matrix control the orientation of the apical surface, so that it faces the lumen interior, opposite to cell-matrix adhesion sites. This orientation of the apical surface is thought to be intrinsically linked to the formation of single lumens. We previously demonstrated in three-dimensional cyst cultures of Madin-Darby canine kidney (MDCK) cells that signaling by β1 integrins regulates the orientation of the apical surface, via a mechanism that depends on the activity of the small GTPase Rac1. Here, we investigated whether the Rac1 effector Pak1 is a downstream effector in this pathway. Expression of constitutive active Pak1 phenocopies the effect of β1 integrin inhibition in that it misorients the apical surface and induces a multilumen phenotype. The misorientation of apical surfaces depends on the interaction of active Pak1 with PIX proteins and is linked to defects in basement membrane assembly. In contrast, the multilumen phenotype was independent of PIX and the basement membrane. Therefore, Pak1 likely regulates apical polarization and lumen formation by two distinct pathways. PMID:22815903

  2. Induction of vascular permeability: beta PIX and GIT1 scaffold the activation of extracellular signal-regulated kinase by PAK.

    PubMed

    Stockton, Rebecca; Reutershan, Jörg; Scott, David; Sanders, John; Ley, Klaus; Schwartz, Martin Alexander

    2007-06-01

    Increased permeability of blood vessels is an important component of inflammation, but in some circumstances it contributes to tissue injury and organ failure. Previous work showed that p21-activated kinase (PAK) is a critical regulator of endothelial cell-cell junctions through effects on myosin light chain phosphorylation and cell contractility. We now show that blocking PAK function inhibits fluid leak in a mouse model of acute lung injury. In cultured endothelial cells, induction of myosin light chain phosphorylation by PAK is mediated by mitogen-activated protein kinase kinase and extracellular signal-regulated kinase (Erk). Erk in lipopolysaccharide (LPS)-treated mouse lung is activated in a PAK-dependent manner in several cell types, most prominently vascular endothelium. Activation of Erk requires the integrity of the complex between PAK, PIX, and GIT1. Several means of disrupting this complex inhibit stimulation of vascular permeability in vitro. A cell-permeant peptide that blocks binding of PAK to PIX inhibits LPS-induced fluid leak in the mouse lung injury model. We conclude that the PAK-PIX-GIT1 complex is critical for Erk-dependent myosin phosphorylation and vascular permeability.

  3. Directional sensing requires G beta gamma-mediated PAK1 and PIX alpha-dependent activation of Cdc42.

    PubMed

    Li, Zhong; Hannigan, Michael; Mo, Zhicheng; Liu, Bo; Lu, Wei; Wu, Yue; Smrcka, Alan V; Wu, Guanqing; Li, Lin; Liu, Mingyao; Huang, Chi-Kuang; Wu, Dianqing

    2003-07-25

    Efficient chemotaxis requires directional sensing and cell polarization. We describe a signaling mechanism involving G beta gamma, PAK-associated guanine nucleotide exchange factor (PIX alpha), Cdc42, and p21-activated kinase (PAK) 1. This pathway is utilized by chemoattractants to regulate directional sensing and directional migration of myeloid cells. Our results suggest that G beta gamma binds PAK1 and, via PAK-associated PIX alpha, activates Cdc42, which in turn activates PAK1. Thus, in this pathway, PAK1 is not only an effector for Cdc42, but it also functions as a scaffold protein required for Cdc42 activation. This G beta gamma-PAK1/PIX alpha/Cdc42 pathway is essential for the localization of F-actin formation to the leading edge, the exclusion of PTEN from the leading edge, directional sensing, and the persistent directional migration of chemotactic leukocytes. Although ligand-induced production of PIP(3) is not required for activation of this pathway, PIP(3) appears to localize the activation of Cdc42 by the pathway.

  4. Decontamination of the Shaft no.1 and cleaning container of 2. block NPP Paks

    SciTech Connect

    Bolcha, Jan; Mala, Zuzana; Tilky, Peter

    2007-07-01

    Available in abstract form only. Full text of publication follows: Meanwhile cleaning fuel assemblies on Paks NPP Unit 2. in 2003 year, the fuel assemblies were damaged, followed by contamination of cleaning container and operating shaft No. 1., in which was the container. As a part of the task - to restore operation NPP Paks, Unit 2, VUJE and.. realized decontamination of the wall of shaft prior to withdrawal of the defected fuel, decontamination of cleaning tank and in consequence decontamination of full shaft No. 1. Solution rest at finished conceptual decontamination proposal, fabrication of special purpose furnished, necessary documentation according to national legislative exigency. Real facilities on decontamination were examined on the stand and on shaft No. 1 in real conditions. This paper describes access method decontaminating procedure, applied facilities assigned on decontamination and present achievement results from decontamination shaft No. 1 realized in August 2006 and February 2007, respectively. Decontamination procedures were chosen on the base of experiments realized in laboratories VUJE and in Paks NPP. Laboratory experiments were realized on the sample of tube used for measurement of neutron flow, from NPP Paks, located in the shaft No.1 in time of event (INES-3). In NPP Paks were realized experiments on cover of cleaning container, which was in time of event situated on cleaning container. To compare decontaminated factors, the chemical and electrochemical procedures for decontamination were tested, and most effective practices were selected. Equipment ROS-740 can be used for the top part of the shaft decontamination. It allows high-pressure admission, rinse and chemical decontamination. Manipulator MAOS-170 is assigned for high-pressure admission of central part of the shaft. (authors)

  5. The p21-activated kinase PAK is negatively regulated by POPX1 and POPX2, a pair of serine/threonine phosphatases of the PP2C family.

    PubMed

    Koh, Cheng-Gee; Tan, E-Jean; Manser, Edward; Lim, Louis

    2002-02-19

    The Rho GTPases are involved in many signaling pathways and cellular functions, including the organization of the actin cytoskeleton, regulation of transcription, cell motility, and cell division. The p21 (Cdc42/Rac)-activated kinase PAK mediates a number of biological effects downstream of these Rho GTPases (reviewed by [1]). The phosphorylation state of mammalian PAK is highly regulated: upon binding of GTPases, PAK is potently activated by autophosphorylation at multiple sites, although the mechanisms of PAK downregulation are not known. We now report two PP2C-like serine/threonine phosphatases (POPX1 and POPX2) that efficiently inactivate PAK. POPX1 was isolated as a binding partner for the PAK interacting guanine nucleotide exchange factor PIX. The dephosphorylating activity of POPX correlates with an ability to block the in vivo effects of active PAK. Consonant with these effects on PAK, POPX can also inhibit actin stress fiber breakdown and morphological changes driven by active Cdc42(V12). The association of the POPX phosphatases with PAK complexes may allow PAK to cycle rapidly between active and inactive states; it represents a unique regulatory component of the signaling pathways of the PAK kinase family.

  6. Invasive Fusobacterium nucleatum activates beta-catenin signaling in colorectal cancer via a TLR4/P-PAK1 cascade.

    PubMed

    Chen, Yongyu; Peng, Yan; Yu, Jiahui; Chen, Ting; Wu, Yaxin; Shi, Lei; Li, Qing; Wu, Jiao; Fu, Xiangsheng

    2017-05-09

    The underlying mechanism of Fusobacterium nucleatum (Fn) in the carcinogenesis of colorectal cancer (CRC) is poorly understood. Here, we examined Fn abundance in CRC tissues, as well as β-catenin, TLR4 and PAK1 protein abundance in Fn positive and Fn negative CRCs. Furthermore, we isolated a strain of Fn (F01) from a CRC tissue and examined whether Fn (F01) infection of colon cancer cells activated β-catenin signaling via the TLR4/P-PAK1/P-β-catenin S675 cascade. Invasive Fn was abundant in 62.2% of CRC tissues. TLR4, PAK1 and nuclear β-catenin proteins were more abundant within Fn-positive over Fn-negative CRCs (P < 0.05). Fn and its lipopolysaccharide induced a significant increase in TLR4/P-PAK1/P-β-catenin S675/C-myc/CyclinD1 protein abundance, as well as in the nuclear translocation of β-catenin. Furthermore, inhibition of TLR4 or PAK1 prior to challenge with Fn significantly decreased protein abundance of P-β-catenin S675, C-myc and Cyclin D1, as well as nuclear β-catenin accumulation. Inhibition of TLR4 significantly decreased P-PAK1 protein abundance, and for the first time, we observed an interaction between TLR4 and P-PAK1 using immunoprecipitation. Our data suggest that invasive Fn activates β-catenin signaling via a TLR4/P-PAK1/P-β-catenin S675 cascade in CRC. Furthermore, TLR4 and PAK1 could be potential pharmaceutical targets for the treatment of Fn-related CRCs.

  7. p21‐Activated kinase (Pak) regulates airway smooth muscle contraction by regulating paxillin complexes that mediate actin polymerization

    PubMed Central

    Zhang, Wenwu; Huang, Youliang

    2016-01-01

    Key points In airway smooth muscle, tension development caused by a contractile stimulus requires phosphorylation of the 20 kDa myosin light chain (MLC), which activates crossbridge cycling and the polymerization of a pool of submembraneous actin.The p21‐activated kinases (Paks) can regulate the contractility of smooth muscle and non‐muscle cells, and there is evidence that this occurs through the regulation of MLC phosphorylation.We show that Pak has no effect on MLC phosphorylation during the contraction of airway smooth muscle, and that it regulates contraction by mediating actin polymerization.We find that Pak phosphorylates the adhesion junction protein, paxillin, on Ser273, which promotes the formation of a signalling complex that activates the small GTPase, cdc42, and the actin polymerization catalyst, neuronal Wiskott–Aldrich syndrome protein (N‐WASP).These studies demonstrate a novel role for Pak in regulating the contractility of smooth muscle by regulating actin polymerization. Abstract The p21‐activated kinases (Pak) can regulate contractility in smooth muscle and other cell and tissue types, but the mechanisms by which Paks regulate cell contractility are unclear. In airway smooth muscle, stimulus‐induced contraction requires phosphorylation of the 20 kDa light chain of myosin, which activates crossbridge cycling, as well as the polymerization of a small pool of actin. The role of Pak in airway smooth muscle contraction was evaluated by inhibiting acetylcholine (ACh)‐induced Pak activation through the expression of a kinase inactive mutant, Pak1 K299R, or by treating tissues with the Pak inhibitor, IPA3. Pak inhibition suppressed actin polymerization and contraction in response to ACh, but it did not affect myosin light chain phosphorylation. Pak activation induced paxillin phosphorylation on Ser273; the paxillin mutant, paxillin S273A, inhibited paxillin Ser273 phosphorylation and inhibited actin polymerization and contraction

  8. A PAK1-PIX-PKL complex is activated by the T-cell receptor independent of Nck, Slp-76 and LAT.

    PubMed

    Ku, G M; Yablonski, D; Manser, E; Lim, L; Weiss, A

    2001-02-01

    Given the importance of the Rho GTPase family member Rac1 and the Rac1/Cdc42 effector PAK1 in T-cell activation, we investigated the requirements for their activation by the T-cell receptor (TCR). Rac1 and PAK1 activation required the tyrosine kinases ZAP-70 and Syk, but not the cytoplasmic adaptor Slp-76. Surprisingly, PAK1 was activated in the absence of the transmembrane adaptor LAT while Rac1 was not. However, efficient PAK1 activation required its binding sites for Rho GTPases and for PIX, a guanine nucleotide exchange factor for Rho GTPases. The overexpression of ssPIX that either cannot bind PAK1 or lacks GEF function blocked PAK1 activation. These data suggest that a PAK1-PIX complex is recruited to appropriate sites for activation and that PIX is required for Rho family GTPase activation upstream of PAK1. Furthermore, we detected a stable trimolecular complex of PAK1, PIX and the paxillin kinase linker p95PKL. Taken together, these data show that PAK1 contained in this trimolecular complex is activated by a novel LAT- and Slp-76-independent pathway following TCR stimulation.

  9. PAK6 increase chemoresistance and is a prognostic marker for stage II and III colon cancer patients undergoing 5-FU based chemotherapy

    PubMed Central

    Yan, Dongwang; Cui, Feifei; Wang, Xiaoliang; Yu, Fudong; Xue, Yingming; Feng, Xiaodong; Wang, Jingtao; Wang, Xiao; Jiang, Tao; Zhang, Meng; Zhao, Senlin; Yu, Yang; Tang, Huamei; Peng, Zhihai

    2015-01-01

    p21-Activated kinase 6 (PAK6) has been implicated in radiotherapy and docetaxel resistance. We have further evaluated PAK6 as a predictor of 5-fluorouracil (5-FU) treatment response in colon cancer. Here we report that in colon cancer PAK6 promotes tumor progression and chemoresistance both in vitro and in vivo. In the clinical analysis, PAK6 was overexpressed in 104 of 147 (70.75%) stage II and III patients who received 5-FU based chemotherapy after surgery. Multivariate Cox regression analysis indicated that PAK6 was an independent prognostic factor for overall survival (P < 0.001) and disease-free survival (P < 0.001). Colon cancer cell lines showed increased PAK6 expression upon 5-FU treatment. In PAK6-knockdown cells treated with 5-FU, cell viability and phosphorylation of BAD decreased, and the number of apoptotic cells, levels of cleaved caspase 3 and PARP increased compared to control cells. The opposite was observed in PAK6 overexpressing cells. Short hairpin RNA knockdown of PAK6 blocked cells in G2-M phase. Furthermore, Animal experiments results in vivo are consistent with outcomes in vitro. This study demonstrates that PAK6 is an independent prognostic factor for adjuvant 5-FU-based chemotherapy in patients with stage II and stage III colon cancer. PMID:25426562

  10. miR-129 suppresses tumor cell growth and invasion by targeting PAK5 in hepatocellular carcinoma

    SciTech Connect

    Zhai, Jian; Qu, Shuping; Li, Xiaowei; Zhong, Jiaming; Chen, Xiaoxia; Qu, Zengqiang; Wu, Dong

    2015-08-14

    Emerging evidence suggests that microRNAs (miRNAs) play important roles in regulating HCC development and progression; however, the mechanisms by which their specific functions and mechanisms remained to be further explored. miR-129 has been reported in gastric cancers, lung cancer and colon cancer. In this study, we disclosed a new tumor suppresser function of miR-129 in HCC. We also found the downregulation of miR-129 occurred in nearly 3/4 of the tumors examined (56/76) compared with adjacent nontumorous tissues, which was more importantly, correlated to the advanced stage and vascular invasion. We then demonstrated that miR-129 overexpression attenuated HCC cells proliferation and invasion, inducing apoptosis in vitro. Moreover, we used miR-129 antagonist and found that anti-miR-129 promoted HCC cells malignant phenotypes. Mechanistically, our further investigations revealed that miR-129 suppressed cell proliferation and invasion by targeting the 3’-untranslated region of PAK5, as well as miR-129 silencing up-regulated PAK5 expression. Moreover, miR-129 expression was inversely correlated with PAK5 expression in 76 cases of HCC samples. RNA interference of PAK5 attenuated anti-miR-129 mediated cell proliferation and invasion in HCC cells. Taken together, these results demonstrated that miR-129 suppressed tumorigenesis and progression by directly targeting PAK5, defining miR-129 as a potential treatment target for HCC. - Highlights: • Decreased of miR-129 is found in HCC and associated with advanced stage and metastasis. • miR-129 suppresses proliferation and invasion of HCC cells. • miR-129 directly targets the 3′ UTR of PAK5 and diminishes PAK5 expression. • PAK5 is involved in miR-129 mediated suppression functions.

  11. Coupling of PAK-interacting exchange factor PIX to GIT1 promotes focal complex disassembly.

    PubMed

    Zhao, Z S; Manser, E; Loo, T H; Lim, L

    2000-09-01

    The p21-activated kinase PAK is targeted to focal complexes (FCs) through interactions with the SH3 domains of the PAK-interacting exchange factor PIX and Nck. PIX is a Rac GTP exchange factor that also binds the G-protein-coupled receptor kinase-interacting protein known as GIT1. Overexpression of GIT1 in fibroblasts or epithelial cells causes a loss of paxillin from FCs and stimulates cell motility. This is due to the direct interaction of a C-terminal 125-residue domain of GIT1 with paxillin, under the regulation of PIX. In its activated state, GIT1 can promote FC disassembly independent of actin-myosin contractile events. Additionally, GIT directly couples to a key component of FCs, focal adhesion kinase (FAK), via a conserved Spa2 homology domain. We propose that GIT1 and FAK cooperate to promote motility both by directly regulating focal complex dynamics and by the activation of Rac.

  12. A novel orally bioavailable compound KPT-9274 inhibits PAK4, and blocks triple negative breast cancer tumor growth

    PubMed Central

    Rane, Chetan; Senapedis, William; Baloglu, Erkan; Landesman, Yosef; Crochiere, Marsha; Das-Gupta, Soumyasri; Minden, Audrey

    2017-01-01

    Breast cancer is a heterogeneous disease consisting of several subtypes. Among these subtypes, triple negative breast cancer is particularly difficult to treat. This is due to a lack of understanding of the mechanisms behind the disease, and consequently a lack of druggable targets. PAK4 plays critical roles in cell survival, proliferation, and morphology. PAK4 protein levels are high in breast cancer cells and breast tumors, and the gene is often amplified in basal like breast cancers, which are frequently triple negative. PAK4 is also overexpressed in other types of cancer, making it a promising drug target. However, its inhibition is complicated by the fact that PAK4 has both kinase-dependent and -independent functions. Here we investigate a new clinical compound KPT-9274, which has been shown to inhibit PAK4 and NAMPT. We find that KPT-9274 (and its analog, KPT-8752) can reduce the steady state level of PAK4 protein in triple negative breast cancer cells. These compounds also block the growth of the breast cancer cells in vitro, and stimulate apoptosis. Most importantly, oral administration of KPT-9274 reduces tumorigenesis in mouse models of human triple negative breast cancer. Our results indicate that KPT-9274 is a novel therapeutic option for triple negative breast cancer therapy. PMID:28198380

  13. Effect of Okinawa Propolis on PAK1 Activity, Caenorhabditis elegans Longevity, Melanogenesis, and Growth of Cancer Cells.

    PubMed

    Taira, Nozomi; Nguyen, Binh Cao Quan; Be Tu, Pham Thi; Tawata, Shinkichi

    2016-07-13

    Propolis from different areas has been reported to inhibit oncogenic/aging kinase PAK1, which is responsible for a variety of conditions, including cancer, longevity, and melanogenesis. Here, a crude extract of Okinawa propolis (OP) was tested against PAK1 activity, Caenorhabditis elegans (C. elegans) longevity, melanogenesis, and growth of cancer cells. We found that OP blocks PAK1 and exhibits anticancer activity in the A549 cell (human lung cancer cell) line with IC50 values of 6 μg/mL and 12 μg/mL, respectively. Most interestingly, OP (1 μg/mL) significantly reduces reproduction and prolongs the lifespan of C. elegans by activating the HSP-16.2 gene, as shown in the PAK1-deficient strain. Furthermore, OP inhibits melanogenesis in a melanoma cell line (B16F10) by downregulating intracellular tyrosinase activity with an IC50 of 30 μg/mL. Our results suggest that OP demonstrated a life span extending effect, C. elegans, anticancer, and antimelanogenic effects via PAK1 inactivation; therefore, this can be a potent natural medicinal supplement against PAK1-dependent diseases.

  14. The dynamic growth exhibition and accumulation of cadmium of Pak choi (Brassica campestris L. ssp. chinensis) grown in contaminated soils.

    PubMed

    Lai, Hung-Yu; Chen, Bo-Ching

    2013-10-25

    The accumulation of heavy metals, especially cadmium (Cd), in leafy vegetables was compared with other vegetables. Pak choi (Brassica campestris L. ssp. chinensis) is a leafy vegetable consumed in Taiwan and its safety for consumption after growing in contaminated soils is a public concern. A pot experiment (50 days) was conducted to understand the dynamic accumulation of Cd by pak choi grown in artificially contaminated soils. The edible parts of pak choi were sampled and analyzed every 2-3 days. The dry weight (DW) of pak choi was an exponential function of leaf length, leaf width, and chlorophyll content. The accumulation of Cd increased when the soil Cd concentration was raised, but was kept at a constant level during different growth stages. Pak choi had a high bioconcentration factor (BCF = ratio of the concentration in the edible parts to that in the soils), at values of 3.5-4.0. The consumption of pak choi grown in soils contaminated at levels used in this study would result in the ingestion of impermissible amounts of Cd and could possibly have harmful effects on health.

  15. Bistability in the Rac1, PAK, and RhoA Signaling Network Drives Actin Cytoskeleton Dynamics and Cell Motility Switches

    PubMed Central

    Byrne, Kate M.; Monsefi, Naser; Dawson, John C.; Degasperi, Andrea; Bukowski-Wills, Jimi-Carlo; Volinsky, Natalia; Dobrzyński, Maciej; Birtwistle, Marc R.; Tsyganov, Mikhail A.; Kiyatkin, Anatoly; Kida, Katarzyna; Finch, Andrew J.; Carragher, Neil O.; Kolch, Walter; Nguyen, Lan K.; von Kriegsheim, Alex; Kholodenko, Boris N.

    2016-01-01

    Summary Dynamic interactions between RhoA and Rac1, members of the Rho small GTPase family, play a vital role in the control of cell migration. Using predictive mathematical modeling, mass spectrometry-based quantitation of network components, and experimental validation in MDA-MB-231 mesenchymal breast cancer cells, we show that a network containing Rac1, RhoA, and PAK family kinases can produce bistable, switch-like responses to a graded PAK inhibition. Using a small chemical inhibitor of PAK, we demonstrate that cellular RhoA and Rac1 activation levels respond in a history-dependent, bistable manner to PAK inhibition. Consequently, we show that downstream signaling, actin dynamics, and cell migration also behave in a bistable fashion, displaying switches and hysteresis in response to PAK inhibition. Our results demonstrate that PAK is a critical component in the Rac1-RhoA inhibitory crosstalk that governs bistable GTPase activity, cell morphology, and cell migration switches. PMID:27136688

  16. Dbo/Henji Modulates Synaptic dPAK to Gate Glutamate Receptor Abundance and Postsynaptic Response

    PubMed Central

    Wang, Manyu; Chen, Pei-Yi; Wang, Chien-Hsiang; Lai, Tzu-Ting; Tsai, Pei-I; Cheng, Ying-Ju; Kao, Hsiu-Hua; Chien, Cheng-Ting

    2016-01-01

    In response to environmental and physiological changes, the synapse manifests plasticity while simultaneously maintains homeostasis. Here, we analyzed mutant synapses of henji, also known as dbo, at the Drosophila neuromuscular junction (NMJ). In henji mutants, NMJ growth is defective with appearance of satellite boutons. Transmission electron microscopy analysis indicates that the synaptic membrane region is expanded. The postsynaptic density (PSD) houses glutamate receptors GluRIIA and GluRIIB, which have distinct transmission properties. In henji mutants, GluRIIA abundance is upregulated but that of GluRIIB is not. Electrophysiological results also support a GluR compositional shift towards a higher IIA/IIB ratio at henji NMJs. Strikingly, dPAK, a positive regulator for GluRIIA synaptic localization, accumulates at the henji PSD. Reducing the dpak gene dosage suppresses satellite boutons and GluRIIA accumulation at henji NMJs. In addition, dPAK associated with Henji through the Kelch repeats which is the domain essential for Henji localization and function at postsynapses. We propose that Henji acts at postsynapses to restrict both presynaptic bouton growth and postsynaptic GluRIIA abundance by modulating dPAK. PMID:27736876

  17. Subgroup II PAK-mediated phosphorylation regulates Ran activity during mitosis

    PubMed Central

    Bompard, Guillaume; Rabeharivelo, Gabriel; Frank, Marie; Cau, Julien; Delsert, Claude

    2010-01-01

    Ran is an essential GTPase that controls nucleocytoplasmic transport, mitosis, and nuclear envelope formation. These functions are regulated by interaction of Ran with different partners, and by formation of a Ran-GTP gradient emanating from chromatin. Here, we identify a novel level of Ran regulation. We show that Ran is a substrate for p21-activated kinase 4 (PAK4) and that its phosphorylation on serine-135 increases during mitosis. The endogenous phosphorylated Ran and active PAK4 dynamically associate with different components of the microtubule spindle during mitotic progression. A GDP-bound Ran phosphomimetic mutant cannot undergo RCC1-mediated GDP/GTP exchange and cannot induce microtubule asters in mitotic Xenopus egg extracts. Conversely, phosphorylation of GTP-bound Ran facilitates aster nucleation. Finally, phosphorylation of Ran on serine-135 impedes its binding to RCC1 and RanGAP1. Our study suggests that PAK4-mediated phosphorylation of GDP- or GTP-bound Ran regulates the assembly of Ran-dependent complexes on the mitotic spindle. PMID:20805321

  18. A Pak- and Pix-dependent branch of the SDF-1alpha signalling pathway mediates T cell chemotaxis across restrictive barriers.

    PubMed

    Volinsky, Natalia; Gantman, Anna; Yablonski, Deborah

    2006-07-01

    Pak (p21-activated kinase) serine/threonine kinases have been shown to mediate directional sensing of chemokine gradients. We hypothesized that Pak may also mediate chemokine-induced shape changes, to facilitate leucocyte chemotaxis through restrictive barriers, such as the extracellular matrix. A potent inhibitor, Pak(i), was characterized and used to probe the role of Pak-family kinases in SDF-1alpha (stromal-cell derived factor-1alpha/CXCL12)-induced chemotaxis in a T cell model. Pak(i) potently inhibited SDF-1alpha-induced Pak activation by a bivalent mechanism, as indicated by its complete inactivation upon point mutation of two binding sites, but partial inactivation upon mutation of either site alone. Importantly, Pak(i) was not toxic to cells over the time frame of our experiments, since it did not substantially affect cell surface expression of CXCR4 (CXC chemokine receptor 4) or integrins, cell cycle progression, or a number of ligand-induced responses. Pak(i) produced dose-dependent inhibition of SDF-1alpha-induced migration through rigid filters bearing small pores; but unexpectedly, did not substantially affect the magnitude or kinetics of chemotaxis through filters bearing larger pores. SDF-1alpha-induced Pak activation was partly dependent on PIX (Pak-interactive exchange factor); correspondingly, an allele of beta-PIX that cannot bind Pak inhibited SDF-1alpha-induced chemotaxis through small, but not large pores. By contrast, other key players in chemotaxis: G(i), PI3K (phosphoinositide 3-kinase), and the Rho-family G-proteins, Rac and Cdc42 (cell division cycle 42), were required for SDF-1alpha-induced migration regardless of the barrier pore-size. These studies have revealed a distinct branch of the SDF-1alpha signalling pathway, in which the Rac/Cdc42 effector, Pak, and its partner, PIX, specifically regulate the cellular events required for chemokine-induced migration through restrictive barriers.

  19. Tyrosyl Phosphorylated Serine-Threonine Kinase PAK1 is a Novel Regulator of Prolactin-Dependent Breast Cancer Cell Motility and Invasion

    PubMed Central

    Hammer, Alan

    2015-01-01

    Despite efforts to discover the cellular pathways regulating breast cancer metastasis, little is known as to how prolactin (PRL) cooperates with extracellular environment and cytoskeletal proteins to regulate breast cancer cell motility and invasion. We implicated serine-threonine kinase p21-activated kinase 1 (PAK1) as a novel target for PRL-activated Janus-kinase 2 (JAK2). JAK2-dependent PAK1 tyrosyl phosphorylation plays a critical role in regulation of both PAK1 kinase activity and scaffolding properties of PAK1. Tyrosyl phosphorylated PAK1 facilitates PRL-dependent motility via at least two mechanisms: formation of paxillin/GIT1/βPIX/pTyr-PAK1 complexes resulting in increased adhesion turnover and phosphorylation of actin-binding protein filamin A. Increased adhesion turnover is the basis for cell migration and phosphorylated filamin A stimulates the kinase activity of PAK1 and increases actin-regulating activity to facilitate cell motility. Tyrosyl phosphorylated PAK1 also stimulates invasion of breast cancer cells in response to PRL and three-dimensional (3D) collagen IV via transcription and secretion of MMP-1 and MMP-3 in a MAPK-dependent manner. These data illustrate the complex interaction between PRL and the cell microenvironment in breast cancer cells and suggest a pivotal role for PRL/PAK1 signaling in breast cancer metastasis. PMID:25472536

  20. PAK4, a novel effector for Cdc42Hs, is implicated in the reorganization of the actin cytoskeleton and in the formation of filopodia.

    PubMed Central

    Abo, A; Qu, J; Cammarano, M S; Dan, C; Fritsch, A; Baud, V; Belisle, B; Minden, A

    1998-01-01

    The GTPases Rac and Cdc42Hs control diverse cellular functions. In addition to being mediators of intracellular signaling cascades, they have important roles in cell morphogenesis and mitogenesis. We have identified a novel PAK-related kinase, PAK4, as a new effector molecule for Cdc42Hs. PAK4 interacts only with the activated form of Cdc42Hs through its GTPase-binding domain (GBD). Co-expression of PAK4 and the constitutively active Cdc42HsV12 causes the redistribution of PAK4 to the brefeldin A-sensitive compartment of the Golgi membrane and the subsequent induction of filopodia and actin polymerization. Importantly, the reorganization of the actin cytoskeleton is dependent on PAK4 kinase activity and on its interaction with Cdc42Hs. Thus, unlike other members of the PAK family, PAK4 provides a novel link between Cdc42Hs and the actin cytoskeleton. The cellular locations of PAK4 and Cdc42Hs suggest a role for the Golgi in cell morphogenesis. PMID:9822598

  1. PAK5 mediates cell: cell adhesion integrity via interaction with E-cadherin in bladder cancer cells.

    PubMed

    Ismail, Ahmad Fahim; Oskay Halacli, Sevil; Babteen, Nouf; De Piano, Mario; Martin, Tracey A; Jiang, Wen G; Khan, Muhammad Shamim; Dasgupta, Prokar; Wells, Claire M

    2017-03-24

    Urothelial bladder cancer is a major cause of morbidity and mortality worldwide, causing an estimated 150 000 deaths per year. Whilst non-muscle-invasive bladder tumours can be effectively treated, with high survival rates, many tumours recur, and some will progress to muscle-invasive disease with a much poorer long-term prognosis. Thus, there is a pressing need to understand the molecular transitions occurring within the progression of bladder cancer to an invasive disease. Tumour invasion is often associated with a down-regulation of E-cadherin expression concomitant with a suppression of cell:cell junctions, and decreased levels of E-cadherin expression have been reported in higher grade urothelial bladder tumours. We find that expression of E-cadherin in a panel of bladder cancer cell lines correlated with the presence of cell:cell junctions and the level of PAK5 expression. Interestingly, exogenous PAK5 has recently been described to be associated with cell:cell junctions and we now find that endogenous PAK5 is localised to cell junctions and interacts with an E-cadherin complex. Moreover, depletion of PAK5 expression significantly reduced junctional integrity. These data suggest a role for PAK5 in maintaining junctional stability and we find that, in both our own patient samples and a commercially available dataset, PAK5mRNA levels are reduced in human bladder cancer compared with normal controls. Taken together, the present study proposes that PAK5 expression levels could be used as a novel prognostic marker for bladder cancer progression. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

  2. A mutation in mouse Pak1ip1 causes orofacial clefting while human PAK1IP1 maps to 6p24 translocation breaking points associated with orofacial clefting.

    PubMed

    Ross, Adam P; Mansilla, M Adela; Choe, Youngshik; Helminski, Simon; Sturm, Richard; Maute, Roy L; May, Scott R; Hozyasz, Kamil K; Wójcicki, Piotr; Mostowska, Adrianna; Davidson, Beth; Adamopoulos, Iannis E; Pleasure, Samuel J; Murray, Jeffrey C; Zarbalis, Konstantinos S

    2013-01-01

    Orofacial clefts are among the most common birth defects and result in an improper formation of the mouth or the roof of the mouth. Monosomy of the distal aspect of human chromosome 6p has been recognized as causative in congenital malformations affecting the brain and cranial skeleton including orofacial clefts. Among the genes located in this region is PAK1IP1, which encodes a nucleolar factor involved in ribosomal stress response. Here, we report the identification of a novel mouse line that carries a point mutation in the Pak1ip1 gene. Homozygous mutants show severe developmental defects of the brain and craniofacial skeleton, including a median orofacial cleft. We recovered this line of mice in a forward genetic screen and named the allele manta-ray (mray). Our findings prompted us to examine human cases of orofacial clefting for mutations in the PAK1IP1 gene or association with the locus. No deleterious variants in the PAK1IP1 gene coding region were recognized, however, we identified a borderline association effect for SNP rs494723 suggesting a possible role for the PAK1IP1 gene in human orofacial clefting.

  3. A Mutation in Mouse Pak1ip1 Causes Orofacial Clefting while Human PAK1IP1 Maps to 6p24 Translocation Breaking Points Associated with Orofacial Clefting

    PubMed Central

    Helminski, Simon; Sturm, Richard; Maute, Roy L.; May, Scott R.; Hozyasz, Kamil K.; Wójcicki, Piotr; Mostowska, Adrianna; Davidson, Beth; Adamopoulos, Iannis E.; Pleasure, Samuel J.; Murray, Jeffrey C.; Zarbalis, Konstantinos S.

    2013-01-01

    Orofacial clefts are among the most common birth defects and result in an improper formation of the mouth or the roof of the mouth. Monosomy of the distal aspect of human chromosome 6p has been recognized as causative in congenital malformations affecting the brain and cranial skeleton including orofacial clefts. Among the genes located in this region is PAK1IP1, which encodes a nucleolar factor involved in ribosomal stress response. Here, we report the identification of a novel mouse line that carries a point mutation in the Pak1ip1 gene. Homozygous mutants show severe developmental defects of the brain and craniofacial skeleton, including a median orofacial cleft. We recovered this line of mice in a forward genetic screen and named the allele manta-ray (mray). Our findings prompted us to examine human cases of orofacial clefting for mutations in the PAK1IP1 gene or association with the locus. No deleterious variants in the PAK1IP1 gene coding region were recognized, however, we identified a borderline association effect for SNP rs494723 suggesting a possible role for the PAK1IP1 gene in human orofacial clefting. PMID:23935987

  4. Paxillin phosphorylation at Ser273 localizes a GIT1-PIX-PAK complex and regulates adhesion and protrusion dynamics.

    PubMed

    Nayal, Anjana; Webb, Donna J; Brown, Claire M; Schaefer, Erik M; Vicente-Manzanares, Miguel; Horwitz, Alan Rick

    2006-05-22

    Continuous adhesion formation and disassembly (adhesion turnover) in the protrusions of migrating cells is regulated by unclear mechanisms. We show that p21-activated kinase (PAK)-induced phosphorylation of serine 273 in paxillin is a critical regulator of this turnover. Paxillin-S273 phosphorylation dramatically increases migration, protrusion, and adhesion turnover by increasing paxillin-GIT1 binding and promoting the localization of a GIT1-PIX-PAK signaling module near the leading edge. Mutants that interfere with the formation of this ternary module abrogate the effects of paxillin-S273 phosphorylation. PAK-dependent paxillin-S273 phosphorylation functions in a positive-feedback loop, as active PAK, active Rac, and myosin II activity are all downstream effectors of this turnover pathway. Finally, our studies led us to identify in highly motile cells a class of small adhesions that reside near the leading edge, turnover in 20-30 s, and resemble those seen with paxillin-S273 phosphorylation. These adhesions appear to be regulated by the GIT1-PIX-PAK module near the leading edge.

  5. Smad3 Couples Pak1 With the Antihypertrophic Pathway Through the E3 Ubiquitin Ligase, Fbxo32.

    PubMed

    Tsui, Hoyee; Zi, Min; Wang, Shunyao; Chowdhury, Sanjoy K; Prehar, Sukhpal; Liang, Qiangrong; Cartwright, Elizabeth J; Lei, Ming; Liu, Wei; Wang, Xin

    2015-12-01

    Pathological cardiac hypertrophy is regarded as a critical intermediate step toward the development of heart failure. Many signal transduction cascades are demonstrated to dictate the induction and progression of pathological hypertrophy; however, our understanding in regulatory mechanisms responsible for the suppression of hypertrophy remains limited. In this study, we showed that exacerbated hypertrophy induced by pressure overload in cardiac-deleted Pak1 mice was attributable to a failure to upregulate the antihypertrophic E3 ligase, Fbxo32, responsible for targeting proteins for the ubiquitin-degradation pathway. Under pressure overload, cardiac overexpression of constitutively active Pak1 mice manifested strong resilience against pathological hypertrophic remodeling. Mechanistic studies demonstrated that subsequent to Pak1 activation, the binding of Smad3 on a critical singular AGAC(-286)-binding site on the FBXO32 promoter was crucial for its transcriptional regulation. Pharmacological upregulation of Fbxo32 by Berberine ameliorated hypertrophic remodeling and improved cardiac performance in cardiac-deficient Pak1 mice under pressure overload. Our findings discover Smad3 and Fbxo32 as novel downstream components of the Pak1-dependent signaling pathway for the suppression of hypertrophy. This discovery opens a new venue for opportunities to identify novel targets for the management of cardiac hypertrophy.

  6. PakB binds to the SH3 domain of Dictyostelium Abp1 and regulates its effects on cell polarity and early development.

    PubMed

    Yang, Yidai; de la Roche, Marc; Crawley, Scott W; Li, Zhihao; Furmaniak-Kazmierczak, Emilia; Côté, Graham P

    2013-07-01

    Dictyostelium p21-activated kinase B (PakB) phosphorylates and activates class I myosins. PakB colocalizes with myosin I to actin-rich regions of the cell, including macropinocytic and phagocytic cups and the leading edge of migrating cells. Here we show that residues 1-180 mediate the cellular localization of PakB. Yeast two-hybrid and pull-down experiments identify two proline-rich motifs in PakB-1-180 that directly interact with the SH3 domain of Dictyostelium actin-binding protein 1 (dAbp1). dAbp1 colocalizes with PakB to actin-rich regions in the cell. The loss of dAbp1 does not affect the cellular distribution of PakB, whereas the loss of PakB causes dAbp1 to adopt a diffuse cytosolic distribution. Cosedimentation studies show that the N-terminal region of PakB (residues 1-70) binds directly to actin filaments, whereas dAbp1 exhibits only a low affinity for filamentous actin. PakB-1-180 significantly enhances the binding of dAbp1 to actin filaments. When overexpressed in PakB-null cells, dAbp1 completely blocks early development at the aggregation stage, prevents cell polarization, and significantly reduces chemotaxis rates. The inhibitory effects are abrogated by the introduction of a function-blocking mutation into the dAbp1 SH3 domain. We conclude that PakB plays a critical role in regulating the cellular functions of dAbp1, which are mediated largely by its SH3 domain.

  7. Combination of immunoprecipitation (IP)-ATP_Glo kinase assay and melanogenesis for the assessment of potent and safe PAK1-blockers in cell culture.

    PubMed

    Nguyen, Binh Cao Quan; Be Tu, Pham Thi; Tawata, Shinkichi; Maruta, Hiroshi

    2015-08-01

    Cucurbitacin I (CBI) is a triterpene from a bitter melon called Goya grown in Okinawa, Japan, and directly inhibits both the Tyr-kinase JAK2 and the G protein RAC, leading to the inactivation of PAK1 (RAC/CDC42-activated kinase 1). Bio 30, a propolis produced in New Zealand, contains CAPE (caffeic acid phenethyl ester) as the major anti-cancer ingredient which directly down-regulates RAC, leading to the inactivation of PAK1. Since PAK1 is essential for the growth of RAS cancer cells such as A549 cell line which carry an oncogenic K-RAS mutant, and the melanogenesis in skin cells, here using these PAK1-blockers as model compounds, we introduce a new approach to the quick assessment of PAK1-blockers in cell culture. First, combining the immuno-precipitation (IP) of PAK1 from cell lysate and the in vitro ATP_Glo kinase assay kit (called "Macaroni-Western" assay), we confirmed that both CBI and Bio 30 inactivate PAK1 in A549 lung cancer cells in 24 h, and inhibit their PAK1-dependent growth in 72 h. Furthermore, we verified that CBI inhibits the PAK1/PAK4-dependent melanogenesis in melanoma cells by far more than 50%, while Bio 30 inhibits the melanogenesis only by 50%, with only a merginal effect on their growth per se. Since the "Macaroni-Western" kinase assay and melanogenesis are both rather simple and quick, the combination of these two cell culture assays would be highly useful for selecting both "potent" (highly cell-permeable) and "safe" (non-toxic) natural or synthetic PAK1-blockers.

  8. Involvement of alpha-PAK-interacting exchange factor in the PAK1-c-Jun NH(2)-terminal kinase 1 activation and apoptosis induced by benzo[a]pyrene.

    PubMed

    Yoshii, S; Tanaka, M; Otsuki, Y; Fujiyama, T; Kataoka, H; Arai, H; Hanai, H; Sugimura, H

    2001-10-01

    Benzo[a]pyrene [B(a)P], a potent procarcinogen found in combustion products such as diesel exhaust and cigarette smoke, has been recently shown to activate the c-Jun NH(2)-terminal kinase 1 (JNK1) and induce caspase-3-mediated apoptosis in Hepa1c1c7 cells. However, the molecules of the signaling pathway that control the mitogen-activated protein kinase cascades induced by B(a)P and the interaction between those and apoptosis by B(a)P have not been well defined. We report here that B(a)P promoted Cdc42/Rac1, p21-activated kinase 1 (PAK1), and JNK1 activities in 293T and HeLa cells. Moreover, alpha-PAK-interacting exchange factor (alpha PIX) mRNA and its protein expression were upregulated by B(a)P. While overexpression of an active mutant of alpha PIX (DeltaCH) facilitated B(a)P-induced activation of Cdc42/Rac1, PAK1, and JNK1, overexpression of mutated alphaPIX (L383R, L384S), which lacks guanine nucleotide exchange factor activity, SH3 domain-deleted alphaPIX (Delta SH3), which lacks the ability to bind PAK, kinase-negative PAK1 (K299R), and kinase-negative SEK1 (K220A, K224L) inhibited B(a)P-triggered JNK1 activation. Interestingly, overexpression of alphaPIX (Delta CH) and a catalytically active mutant PAK1 (T423E) accelerated B(a)P-induced apoptosis in HeLa cells, whereas alphaPIX (Delta SH3), PAK1 (K299R), and SEK 1 (K220A, K224L) inhibited B(a)P-initiated apoptosis. Finally, a preferential caspase inhibitor, Z-Asp-CH2-DCB, strongly blocked the alphaPIX (Delta CH)-enhanced apoptosis in cells treated with B(a)P but did not block PAK1/JNK1 activation. Taken together, these results indicate that alphaPIX plays a crucial role in B(a)P-induced apoptosis through activation of the JNK1 pathway kinases.

  9. Cadmium accumulation in pak choi (Brassica chinensis L.) and estimated dietary intake in the suburb of Hangzhou city, China.

    PubMed

    Yan, S; Ling, Q; Bao, Z; Chen, Z; Yan, S; Dong, Z; Zhang, B; Deng, B

    2009-01-01

    The total content of cadmium and its distribution between leaves and roots were investigated for pak choi (Brassica chinensis L.), which was grown in a contaminated area of Hangzhou city, China. Results showed that cadmium concentrations in 90% of samples exceeded the Chinese tolerance limit for food. Although roots of pak choi had a higher concentration of cadmium than leaves, leaf tissue (the edible part) accumulated over 80% (on average) of the whole plant cadmium burden due to the greater biomass in leaf tissue. Since pak choi is the staple vegetable in the study area and of great relevance to human health, the dietary intake of cadmium and calculated risk indexes to local residents through the food chain were determined.

  10. Structural analysis of the SH3 domain of beta-PIX and its interaction with alpha-p21 activated kinase (PAK).

    PubMed

    Mott, Helen R; Nietlispach, Daniel; Evetts, Katrina A; Owen, Darerca

    2005-08-23

    The PAK Ser/Thr kinases are important downstream effectors of the Rho family GTPases Cdc42 and Rac, partly mediating the role of these G proteins in cell proliferation and cytoskeletal rearrangements. As well as small G proteins, PAK interacts with the Cdc42/Rac exchange factor beta-PIX via the PIX SH3 domain and a nontypical Pro-rich region in PAK. This interaction is thought to affect the localization of PAK, as well as increased GTP/GDP exchange of Rac and Cdc42. We have determined the structure of the PIX-SH3/PAK peptide complex and shown that it differs from typical Src-like SH3/peptide complexes. The peptide makes contacts through the Pro-rich sequence in a similar way to standard SH3/peptide complexes, even though the Pro residue positions are not conserved. In addition, there are interactions with a Pro and Lys in the PAK, which are C-terminal to the conserved Arg found in all SH3-binding sequences. These contact a fourth binding pocket on the SH3 domain. We have measured the affinity of PIX-SH3 for the PAK peptide and found that it is of intermediate affinity. When PAK is activated, Ser-199 in the PIX-binding site is phosphorylated. This phosphorylation is sufficient to reduce the affinity for PIX 6-fold.

  11. Novel p21-Activated Kinase 4 (PAK4) Allosteric Modulators Overcome Drug Resistance and Stemness in Pancreatic Ductal Adenocarcinoma.

    PubMed

    Aboukameel, Amro; Muqbil, Irfana; Senapedis, William; Baloglu, Erkan; Landesman, Yosef; Shacham, Sharon; Kauffman, Michael; Philip, Philip A; Mohammad, Ramzi M; Azmi, Asfar S

    2017-01-01

    The p21-activated kinase 4 (PAK4) is a key downstream effector of the Rho family GTPases and is found to be overexpressed in pancreatic ductal adenocarcinoma (PDAC) cells but not in normal human pancreatic ductal epithelia (HPDE). Gene copy number amplification studies in PDAC patient cohorts confirmed PAK4 amplification making it an attractive therapeutic target in PDAC. We investigated the antitumor activity of novel PAK4 allosteric modulators (PAM) on a panel of PDAC cell lines and chemotherapy-resistant flow-sorted PDAC cancer stem cells (CSC). The toxicity and efficacy of PAMs were evaluated in multiple subcutaneous mouse models of PDAC. PAMs (KPT-7523, KPT-7189, KPT-8752, KPT-9307, and KPT-9274) show antiproliferative activity in vitro against different PDAC cell lines while sparing normal HPDE. Cell growth inhibition was concurrent with apoptosis induction and suppression of colony formation in PDAC. PAMs inhibited proliferation and antiapoptotic signals downstream of PAK4. Co-immunoprecipitation experiments showed disruption of PAK4 complexes containing vimentin. PAMs disrupted CSC spheroid formation through suppression of PAK4. Moreover, PAMs synergize with gemcitabine and oxaliplatin in vitro KPT-9274, currently in a phase I clinical trial (clinicaltrials.gov; NCT02702492), possesses desirable pharmacokinetic properties and is well tolerated in mice with the absence of any signs of toxicity when 200 mg/kg daily is administered either intravenously or orally. KPT-9274 as a single agent showed remarkable antitumor activity in subcutaneous xenograft models of PDAC cell lines and CSCs. These proof-of-concept studies demonstrated the antiproliferative effects of novel PAMs in PDAC and warrant further clinical investigations. Mol Cancer Ther; 16(1); 76-87. ©2016 AACR.

  12. MiR-145 regulates PAK4 via the MAPK pathway and exhibits an antitumor effect in human colon cells

    SciTech Connect

    Wang, Zhigang; Zhang, Xiaoping; Yang, Zhili; Du, Hangxiang; Wu, Zhenqian; Gong, Jianfeng; Yan, Jun; Zheng, Qi

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer MiR-145 targets a putative binding site in the 3 Prime UTR of PAK4. Black-Right-Pointing-Pointer MiR-145 played an important role in inhibiting cell growth by directly targeting PAK4. Black-Right-Pointing-Pointer MiR-145 may function as tumor suppressors. -- Abstract: MicroRNAs (miRNAs) are regulators of numerous cellular events; accumulating evidence indicates that miRNAs play a key role in a wide range of biological functions, such as cellular proliferation, differentiation, and apoptosis in cancer. Down-regulated expression of miR-145 has been reported in colon cancer tissues and cell lines. The molecular mechanisms underlying miR-145 and the regulation of colon carcinogenesis remain unclear. In this study, we investigated the levels of miR-145 in human colon cancer cells using qRT-PCR and found markedly decreased levels compared to normal epithelial cells. We identified PAK4 as a novel target of miR-145 using informatics screening. Additionally, we demonstrated that miR-145 targets a putative binding site in the 3 Prime UTR of PAK4 and that its abundance is inversely associated with miR-145 expression in colon cancer cells; we confirmed this relationship using the luciferase reporter assay. Furthermore, restoration of miR-145 by mimics in SW620 cells significantly attenuated cell growth in vitro, in accordance with the inhibitory effects induced by siRNA mediated knockdown of PAK4. Taken together, these findings demonstrate that miR-145 downregulates P-ERK expression by targeting PAK4 and leads to inhibition of tumor growth.

  13. Crystal Structure of the SH3 Domain of beta PIX in Complex with a High Affinity Peptide from PAK2

    SciTech Connect

    Hoelz,A.; Janz, J.; Lawrie, S.; Corwin, B.; Lee, A.; Sakmar, T.

    2006-01-01

    The p21-activated kinases (PAKs) are important effector proteins of the small GTPases Cdc42 and Rac and control cytoskeletal rearrangements and cell proliferation. The direct interaction of PAKs with guanine nucleotide exchange factors from the PIX/Cool family, which is responsible for the localization of PAK kinases to focal complexes in the cell, is mediated by a 24-residue peptide segment in PAKs and an N-terminal src homology 3 (SH3) domain in PIX/Cool. The SH3-binding segment of PAK contains the atypical consensus-binding motif PxxxPR, which is required for unusually high affinity binding. In order to understand the structural basis for the high affinity and specificity of the PIX-PAK interaction, we solved crystal structures for the N-terminal SH3 domain of {beta}PIX and for the complex of the atypical binding segment of PAK2 with the N-terminal SH3 domain of {beta}PIX at 0.92 Angstroms and 1.3 Angstroms resolution, respectively. The asymmetric unit of the crystal contains two SH3 domains and two peptide ligands. The bound peptide adopts a conformation that allows for intimate contacts with three grooves on the surface of the SH3 domain that lie between the n-Src and RT-loops. Most notably, the arginine residue of the PxxxPR motif forms a salt-bridge and is tightly coordinated by a number of residues in the SH3 domain. This arginine-specific interaction appears to be the key determinant for the high affinity binding of PAK peptides. Furthermore, C-terminal residues of the peptide engage in additional interactions with the surface of the RT-loop, which significantly increases binding specificity. Compared to a recent NMR structure of a similar complex, our crystal structure reveals an alternate binding mode. Finally, we compare our crystal structure with the recently published {beta}PIX/Cbl-b complex structure, and suggest the existence of a molecular switch.

  14. [Rapid isolation of 4-hydroxyphenylethanol from the bovine brain using Water Sep-Pak Plus cartridges with lipophilic sorbent (Florisil)].

    PubMed

    Panova, N G; Veselovskaia, N V; Medvedev, A E

    1996-01-01

    4-Hydroxypherlylethanol, MAO-A Inhibitory component of endogenous MAO Inhibitory activity, tribulin, was isolated from a small quantity of bovine brain using Maters Sep-Pak Florisil cartridges. Procedure included isolation of crude tribulin fraction, which was passed through the cartridge. Preliminary data revealed that this substance is not absorbed on Florisil. The employment of this type of cartridges essentially accelerates isolation of 4-hydroxyphenyethanol from brain samples for quantitative determination. Pilot experiments also revealed that Sep-Pak Folrisil cartridges could be used for the rapid isolation of isatin, another endogenous MAO (B) inhibitor, from brain tissue.

  15. PAK-dependent STAT5 serine phosphorylation is required for BCR-ABL-induced leukemogenesis

    PubMed Central

    Berger, A; Hoelbl-Kovacic, A; Bourgeais, J; Hoefling, L; Warsch, W; Grundschober, E; Uras, I Z; Menzl, I; Putz, E M; Hoermann, G; Schuster, C; Fajmann, S; Leitner, E; Kubicek, S; Moriggl, R; Gouilleux, F; Sexl, V

    2014-01-01

    The transcription factor STAT5 (signal transducer and activator of transcription 5) is frequently activated in hematological malignancies and represents an essential signaling node downstream of the BCR-ABL oncogene. STAT5 can be phosphorylated at three positions, on a tyrosine and on the two serines S725 and S779. We have investigated the importance of STAT5 serine phosphorylation for BCR-ABL-induced leukemogenesis. In cultured bone marrow cells, expression of a STAT5 mutant lacking the S725 and S779 phosphorylation sites (STAT5SASA) prohibits transformation and induces apoptosis. Accordingly, STAT5SASA BCR-ABL+ cells display a strongly reduced leukemic potential in vivo, predominantly caused by loss of S779 phosphorylation that prevents the nuclear translocation of STAT5. Three distinct lines of evidence indicate that S779 is phosphorylated by group I p21-activated kinase (PAK). We show further that PAK-dependent serine phosphorylation of STAT5 is unaffected by BCR-ABL tyrosine kinase inhibitor treatment. Interfering with STAT5 phosphorylation could thus be a novel therapeutic approach to target BCR-ABL-induced malignancies. PMID:24263804

  16. PAK1-cofilin phosphorylation mediates human lung adenocarcinoma cells migration induced by apelin-13.

    PubMed

    Lv, Deguan; Li, Lanfang; Lu, Qixuan; Li, Yao; Xie, Feng; Li, Hening; Cao, Jiangang; Liu, Meiqing; Wu, Di; He, Lu; Chen, Linxi

    2016-05-01

    Adipocytokines apelin peptide, the ligand of APJ (putative receptor related to the angiotensin receptor AT1), plays key roles in the pathogenesis and deterioration of cancer. In lung cancer, apelin elevating microvessel densities has been reported. Our previous research has characterized that apelin-13 promoted lung adenocarcinoma cell proliferation. However, the effect of apelin on metastasis in lung adenocarcinoma and the underlying mechanisms remain unclear. This study shows that apelin-13 induced human adenocarcinoma cell migration via the APJ receptor. Apelin-13 phosphorylated PAK1 and cofilin increase the migration of lung adenocarcinoma cells. Moreover, the results verify that over-expression of apelin and APJ contributed to reducing the effect of doxorubicin and razoxane on inhibiting lung adenocarcinoma cells metastasis. Hypoxia activated APJ expression and apelin release in lung adenocarcinoma cells. The results demonstrate a PAK1-cofilin phosphorylation mechanism to mediate lung adenocarcinoma cells migration promoted by apelin-13. This discovery further suggests that APJ and its downstream signalling is a potential target for anti-metastatic therapies in lung adenocarcinoma patients. © 2016 John Wiley & Sons Australia, Ltd.

  17. PAK proteins and YAP-1 signalling downstream of integrin beta-1 in myofibroblasts promote liver fibrosis

    PubMed Central

    Martin, Katherine; Pritchett, James; Llewellyn, Jessica; Mullan, Aoibheann F.; Athwal, Varinder S.; Dobie, Ross; Harvey, Emma; Zeef, Leo; Farrow, Stuart; Streuli, Charles; Henderson, Neil C.; Friedman, Scott L.; Hanley, Neil A.; Piper Hanley, Karen

    2016-01-01

    Fibrosis due to extracellular matrix (ECM) secretion from myofibroblasts complicates many chronic liver diseases causing scarring and organ failure. Integrin-dependent interaction with scar ECM promotes pro-fibrotic features. However, the pathological intracellular mechanism in liver myofibroblasts is not completely understood, and further insight could enable therapeutic efforts to reverse fibrosis. Here, we show that integrin beta-1, capable of binding integrin alpha-11, regulates the pro-fibrotic phenotype of myofibroblasts. Integrin beta-1 expression is upregulated in pro-fibrotic myofibroblasts in vivo and is required in vitro for production of fibrotic ECM components, myofibroblast proliferation, migration and contraction. Serine/threonine-protein kinase proteins, also known as P21-activated kinase (PAK), and the mechanosensitive factor, Yes-associated protein 1 (YAP-1) are core mediators of pro-fibrotic integrin beta-1 signalling, with YAP-1 capable of perpetuating integrin beta-1 expression. Pharmacological inhibition of either pathway in vivo attenuates liver fibrosis. PAK protein inhibition, in particular, markedly inactivates the pro-fibrotic myofibroblast phenotype, limits scarring from different hepatic insults and represents a new tractable therapeutic target for treating liver fibrosis. PMID:27535340

  18. Ivermectin Induces Cytostatic Autophagy by Blocking the PAK1/Akt Axis in Breast Cancer.

    PubMed

    Dou, Qianhui; Chen, Hai-Ning; Wang, Kui; Yuan, Kefei; Lei, Yunlong; Li, Kai; Lan, Jiang; Chen, Yan; Huang, Zhao; Xie, Na; Zhang, Lu; Xiang, Rong; Nice, Edouard C; Wei, Yuquan; Huang, Canhua

    2016-08-01

    Breast cancer is the most common cancer among women worldwide, yet successful treatment remains a clinical challenge. Ivermectin, a broad-spectrum antiparasitic drug, has recently been characterized as a potential anticancer agent due to observed antitumor effects. However, the molecular mechanisms involved remain poorly understood. Here, we report a role for ivermectin in breast cancer suppression by activating cytostatic autophagy both in vitro and in vivo Mechanistically, ivermectin-induced autophagy in breast cancer cells is associated with decreased P21-activated kinase 1 (PAK1) expression via the ubiquitination-mediated degradation pathway. The inhibition of PAK1 decreases the phosphorylation level of Akt, resulting in the blockade of the Akt/mTOR signaling pathway. In breast cancer xenografts, the ivermectin-induced cytostatic autophagy leads to suppression of tumor growth. Together, our results provide a molecular basis for the use of ivermectin to inhibit the proliferation of breast cancer cells and indicate that ivermectin is a potential option for the treatment of breast cancer. Cancer Res; 76(15); 4457-69. ©2016 AACR. ©2016 American Association for Cancer Research.

  19. Monitoring system with automatic sampling units in the surroundings Paks NPP

    NASA Astrophysics Data System (ADS)

    Svingor, É.; Molnár, M.; Palcsu, L.; Veres, M.; Pintér, T.; Köves, L.

    2006-01-01

    Continuous monitoring of the uncontrolled tritium emission from Paks Nuclear Power Plant (NPP) has been carried out since starting the NPP operation. In order to control the release of fission and corrosion products as well as 14C automatic samplers were designed and installed into 20 observation wells in the vicinity of the reactor units. The automatic samplers contain two columns filled with anion and cation exchanger resins and are equipped with filters for trapping the chelating agents. The samplers are placed two meters below the groundwater level. Water pumps fed by accumulator ensure the water flowing through the ion-exchange columns. After a two-month working period the trapped ions are eluted from the resins. The activity of the gamma-emitters, 14C, 90Sr and transuranium elements are measured in the dried elute, the tritium is measured in the collected water phase. The experiences of a five-year monitoring work in the surroundings of Paks NPP are given here.

  20. Pak1 regulates branching morphogenesis in 3D MDCK cell culture by a PIX and beta1-integrin-dependent mechanism.

    PubMed

    Hunter, Michael P; Zegers, Mirjam M

    2010-07-01

    Branching morphogenesis is a fundamental process in the development of the kidney. This process gives rise to a network of ducts, which form the collecting system. Defective branching can lead to a multitude of kidney disorders including agenesis and reduced nephron number. The formation of branching tubules involves changes in cell shape, cell motility, and reorganization of the cytoskeleton. However, the exact intracellular mechanisms involved are far from understood. We have used the three-dimensional (3D) Madin-Darby canine kidney (MDCK) cell culture system to study how p21-activated kinase 1 (Pak1), which is an important regulator of the cytoskeleton, modulates branching. Our data reveal that Pak1 plays a crucial role in regulating branching morphogenesis. Expression of a dominant-negative Pak1 mutant (DN-Pak1) in MDCK cysts resulted in the spontaneous formation of extensions and branching tubules. Cellular contractility and levels of phosphorylated myosin light chain (pMLC) were increased in DN-Pak1 cells in collagen. Expression of a DN-Pak1 mutant that does not bind to PIX (DN-Pak1-DeltaPIX) failed to form extensions in collagen and did not have increased contractility. This shows that the DN-Pak1 mutant requires PIX binding to generate extensions and increased contractility in 3D culture. Furthermore, a beta1-integrin function-blocking antibody (AIIB2) inhibited the formation of branches and blocked the increased contractility in DN-Pak1 cysts. Taken together, our work shows that DN-Pak1-induced branching morphogenesis requires PIX binding and beta1-integrin signaling.

  1. Effects of selenite and selenate application on growth and shoot selenium accumulation of pak choi (Brassica chinensis L.) during successive planting conditions.

    PubMed

    Li, Jun; Liang, Dongli; Qin, Siyue; Feng, Puyang; Wu, Xiongping

    2015-07-01

    Selenate and selenite are two main kinds of inorganic selenium (Se) sources in soil, but these substances can pose threats to the environment. Phytoextraction is an emerging technology to remove Se from polluted soils by using a hyper-accumulator. In this study, a pot experiment was conducted to investigate Se phytoextraction potential of pak choi (Brassica chinensis L.) and to determine the effects of Se on growth and Se accumulation of pak choi under successive planting conditions (four crops). Results showed that Se concentration in pak choi shoots significantly increased as selenate and selenite rates increased. Se concentration increased in successive crops on soil treated with selenite; by contrast, Se concentration decreased in crops on soil treated with selenate. Se concentrations of pak choi on soil treated with selenate were higher than those on soil treated with selenite. The maximum Se accumulations amount in crops on selenite- and selenate-treated soil were 7818 and 8828 μg · pot(-1), respectively. High bioconcentration factor (BCF) values indicated that pak choi could accumulate more Se from Se-contaminated soil. The Se phytoextraction efficiency of pak choi increased under successive planting conditions in selenite and selenate treatments; the maximum Se phytoextraction efficiencies of four successive crops of pak choi on selenite- and selenate-treated soil were 4.91 and 31.90 %, respectively. These differences between selenate and selenite treatments were attributed to the differences in Se forms in soil. Total and available Se contents in soil decreased significantly during repeated planting crops on soil treated with selenate; conversely, total and available Se contents decreased slightly in crops on soil treated with selenite. These results suggested that pak choi could highly tolerate and accumulate Se. Thus, pak choi may remove Se from contaminated soil; indeed, pak choi can be used in the phytoextraction of Se in polluted soil.

  2. Pak1 regulates branching morphogenesis in 3D MDCK cell culture by a PIX and β1-integrin-dependent mechanism

    PubMed Central

    Hunter, Michael P.

    2010-01-01

    Branching morphogenesis is a fundamental process in the development of the kidney. This process gives rise to a network of ducts, which form the collecting system. Defective branching can lead to a multitude of kidney disorders including agenesis and reduced nephron number. The formation of branching tubules involves changes in cell shape, cell motility, and reorganization of the cytoskeleton. However, the exact intracellular mechanisms involved are far from understood. We have used the three-dimensional (3D) Madin-Darby canine kidney (MDCK) cell culture system to study how p21-activated kinase 1 (Pak1), which is an important regulator of the cytoskeleton, modulates branching. Our data reveal that Pak1 plays a crucial role in regulating branching morphogenesis. Expression of a dominant-negative Pak1 mutant (DN-Pak1) in MDCK cysts resulted in the spontaneous formation of extensions and branching tubules. Cellular contractility and levels of phosphorylated myosin light chain (pMLC) were increased in DN-Pak1 cells in collagen. Expression of a DN-Pak1 mutant that does not bind to PIX (DN-Pak1-ΔPIX) failed to form extensions in collagen and did not have increased contractility. This shows that the DN-Pak1 mutant requires PIX binding to generate extensions and increased contractility in 3D culture. Furthermore, a β1-integrin function-blocking antibody (AIIB2) inhibited the formation of branches and blocked the increased contractility in DN-Pak1 cysts. Taken together, our work shows that DN-Pak1-induced branching morphogenesis requires PIX binding and β1-integrin signaling. PMID:20457839

  3. Downregulation of microRNA-23a suppresses prostate cancer metastasis by targeting the PAK6-LIMK1 signaling pathway

    PubMed Central

    Li, Xiaojuan; Cai, Yi; Ye, Zhiqiang; Li, Shigeng; Li, Jun; Huang, Huaiqiu; Peng, Shubin; Wang, Jun; Tao, Yiran; Huang, Hongxing; Wen, Xinglai; Mo, Jianfeng; Deng, Zhupeng; Wang, Jian; Zhang, Yangfan; Gao, Xin; Wen, Xingqiao

    2015-01-01

    Here we found that levels of miR-23a were decreased in prostate cancer cell lines and tumor tissues. These low levels were associated with poor patients' prognosis. MiR-23a inhibited migration and invasion of prostate cancer in vivo and in orthotopic prostate cancer mice model. MiR-23a decreased levels of p21-activated kinase 6 (PAK6). Expression of miR-23a inhibited phosphorylation of LIM kinase 1 (LIMK1) and cofilin, in turn suppressing formation of stress fibers and actin filaments, which was required for cell motility and invasion. PAK6 bound to LIMK1 and activated it via phosphorylation at Thr-508. Also, PAK6 and LIMK1 were colocalized in the cytoplasma. Thus, miR-23a regulated cytoskeleton by affecting LIMK1 and cofilin. In summary, we have identified the miR-23a-PAK6-LIMK1 pathway of prostate cancer metastasis. Potential therapeutic approach by targeting miR-23 is suggested. PMID:25714010

  4. Cdk1 phosphorylates the Rac activator Tiam1 to activate centrosomal Pak and promote mitotic spindle formation

    PubMed Central

    Whalley, Helen J.; Porter, Andrew P.; Diamantopoulou, Zoi; White, Gavin R. M.; Castañeda-Saucedo, Eduardo; Malliri, Angeliki

    2015-01-01

    Centrosome separation is critical for bipolar spindle formation and the accurate segregation of chromosomes during mammalian cell mitosis. Kinesin-5 (Eg5) is a microtubule motor essential for centrosome separation, and Tiam1 and its substrate Rac antagonize Eg5-dependent centrosome separation in early mitosis promoting efficient chromosome congression. Here we identify S1466 of Tiam1 as a novel Cdk1 site whose phosphorylation is required for the mitotic function of Tiam1. We find that this phosphorylation of Tiam1 is required for the activation of group I p21-activated kinases (Paks) on centrosomes in prophase. Further, we show that both Pak1 and Pak2 counteract centrosome separation in a kinase-dependent manner and demonstrate that they act downstream of Tiam1. We also show that depletion of Pak1/2 allows cells to escape monopolar arrest by Eg5 inhibition, highlighting the potential importance of this signalling pathway for the development of Eg5 inhibitors as cancer therapeutics. PMID:26078008

  5. Cdk1 phosphorylates the Rac activator Tiam1 to activate centrosomal Pak and promote mitotic spindle formation.

    PubMed

    Whalley, Helen J; Porter, Andrew P; Diamantopoulou, Zoi; White, Gavin R M; Castañeda-Saucedo, Eduardo; Malliri, Angeliki

    2015-06-16

    Centrosome separation is critical for bipolar spindle formation and the accurate segregation of chromosomes during mammalian cell mitosis. Kinesin-5 (Eg5) is a microtubule motor essential for centrosome separation, and Tiam1 and its substrate Rac antagonize Eg5-dependent centrosome separation in early mitosis promoting efficient chromosome congression. Here we identify S1466 of Tiam1 as a novel Cdk1 site whose phosphorylation is required for the mitotic function of Tiam1. We find that this phosphorylation of Tiam1 is required for the activation of group I p21-activated kinases (Paks) on centrosomes in prophase. Further, we show that both Pak1 and Pak2 counteract centrosome separation in a kinase-dependent manner and demonstrate that they act downstream of Tiam1. We also show that depletion of Pak1/2 allows cells to escape monopolar arrest by Eg5 inhibition, highlighting the potential importance of this signalling pathway for the development of Eg5 inhibitors as cancer therapeutics.

  6. Degradation Behavior and Accelerated Weathering of Composite Boards Produced from Waste Tetra Pak® Packaging Materials

    Treesearch

    Nural Yilgor; Coskun Kose; Evren Terzi; Aysel Kanturk Figen; Rebecca Ibach; S. Nami Kartal; Sabriye Piskin

    2014-01-01

    Manufacturing panels from Tetra Pak® (TP) packaging material might be an alternative to conventional wood-based panels. This study evaluated some chemical and physical properties as well as biological, weathering, and fire performance of panels with and without zinc borate (ZnB) by using shredded TP packaging cartons. Such packaging material, a worldwide well-known...

  7. IPA-3 Inhibits the Growth of Liver Cancer Cells By Suppressing PAK1 and NF-κB Activation

    PubMed Central

    Wong, Leo Lap-Yan; Lam, Ian Pak-Yan; Wong, Tracy Yuk-Nar; Lai, Wai-Lung; Liu, Heong-Fai; Yeung, Lam-Lung; Ching, Yick-Pang

    2013-01-01

    Hepatocellular carcinoma (HCC) is one of the major malignancies worldwide and is associated with poor prognosis due to the high incidences of metastasis and tumor recurrence. Our previous study showed that overexpression of p21-activated protein kinase 1 (PAK1) is frequently observed in HCC and is associated with a more aggressive tumor behavior, suggesting that PAK1 is a potential therapeutic target in HCC. In the current study, an allosteric small molecule PAK1 inhibitor, IPA-3, was evaluated for the potential in suppressing hepatocarcinogenesis. Consistent with other reports, inhibition of PAK1 activity was observed in several human HCC cell lines treated with various dosages of IPA-3. Using cell proliferation, colony formation and BrdU incorporation assays, we demonstrated that IPA-3 treatment significantly inhibited the growth of HCC cells. The mechanisms through which IPA-3 treatment suppresses HCC cell growth are enhancement of apoptosis and blockage of activation of NF-κB. Furthermore, our data suggested that IPA-3 not only inhibits the HCC cell growth, but also suppresses the metastatic potential of HCC cells. Nude mouse xenograft assay demonstrated that IPA-3 treatment significantly reduced the tumor growth rate and decreased tumor volume, indicating that IPA-3 can suppress the in vivo tumor growth of HCC cells. Taken together, our demonstration of the potential preclinical efficacy of IPA-3 in HCC provides the rationale for cancer therapy. PMID:23894351

  8. Abnormalities of the Duo/Rac-1/PAK1 Pathway Drive Myosin Light Chain Phosphorylation in Frontal Cortex in Schizophrenia

    PubMed Central

    Rubio, María D.; Haroutunian, Vahram; Meador-Woodruff, James H.

    2012-01-01

    BACKGROUND Recent studies on GTPases have suggested that reduced Duo and Cdc42 transcript expression is involved in dendritic spine loss in schizophrenia. In murine models, Duo and Cdc42 phosphorylate PAK1, which modifies the activity of regulatory myosin light chain (MLC) and cofilin by altering their phosphorylation. Therefore, we hypothesized that in schizophrenia abnormal Duo and Cdc42 expression result in changes in MLC and/or cofilin phosphorylation, which may alter actin cytoskeleton dynamics underlying dendritic spine maintenance. METHODS We performed Western blot protein expression analysis in postmortem brains from patients diagnosed with schizophrenia and a comparison group. We focused our studies in the anterior cingulate cortex (ACC) (n=33 comparison group; n=36 schizophrenia) and dorsolateral prefrontal cortex (DLPFC) (n=29 comparison group; n=35 schizophrenia). RESULTS In both ACC and DLPFC, we found a reduction of Duo expression and PAK1 phosphorylation in schizophrenia. Cdc42 protein expression was decreased in ACC, but not in DLPFC. In ACC, we observed decreased PAK1 phosphorylation and increased MLC (pMLC) phosphorylation, while in DLPFC pMLC remained unchanged. DISCUSSION These data suggest a novel mechanism that may underlie dendritic spine loss in schizophrenia. The increase in pMLC seen in ACC may be associated with dendritic spine shrinkage. The lack of an effect on pMLC in DLPFC suggests that in schizophrenia PAK1 downstream pathways are differentially affected in these cortical areas. PMID:22458949

  9. Accelerated Evolution of PAK3- and PIM1-like Kinase Gene Families in the Zebra Finch, Taeniopygia guttata

    PubMed Central

    Kong, Lesheng; Lovell, Peter V.; Heger, Andreas; Mello, Claudio V.; Ponting, Chris P.

    2010-01-01

    Genes encoding protein kinases tend to evolve slowly over evolutionary time, and only rarely do they appear as recent duplications in sequenced vertebrate genomes. Consequently, it was a surprise to find two families of kinase genes that have greatly and recently expanded in the zebra finch (Taeniopygia guttata) lineage. In contrast to other amniotic genomes (including chicken) that harbor only single copies of p21-activated serine/threonine kinase 3 (PAK3) and proviral integration site 1 (PIM1) genes, the zebra finch genome appeared at first to additionally contain 67 PAK3-like (PAK3L) and 51 PIM1-like (PIM1L) protein kinase genes. An exhaustive analysis of these gene models, however, revealed most to be incomplete, owing to the absence of terminal exons. After reprediction, 31 PAK3L genes and 10 PIM1L genes remain, and all but three are predicted, from the retention of functional sites and open reading frames, to be enzymatically active. PAK3L, but not PIM1L, gene sequences show evidence of recurrent episodes of positive selection, concentrated within structures spatially adjacent to N- and C-terminal protein regions that have been discarded from zebra finch PAK3L genes. At least seven zebra finch PAK3L genes were observed to be expressed in testis, whereas two sequences were found transcribed in the brain, one broadly including the song nuclei and the other in the ventricular zone and in cells resembling Bergmann's glia in the cerebellar Purkinje cell layer. Two PIM1L sequences were also observed to be expressed with broad distributions in the zebra finch brain, one in both the ventricular zone and the cerebellum and apparently associated with glial cells and the other showing neuronal cell expression and marked enrichment in midbrain/thalamic nuclei. These expression patterns do not correlate with zebra finch-specific features such as vocal learning. Nevertheless, our results show how ancient and conserved intracellular signaling molecules can be co

  10. Signal therapy of breast cancers by the HDAC inhibitor FK228 that blocks the activation of PAK1 and abrogates the tamoxifen-resistance.

    PubMed

    Hirokawa, Yumiko; Arnold, Melissa; Nakajima, Hidenori; Zalcberg, John; Maruta, Hiroshi

    2005-09-01

    PAK1, a Rac/CDC42-dependent Ser/Thr kinase, is required for both neurofibromatosis (NF) and RAS transformation in vivo. FK228, a histone deacetylase (HDAC) inhibitor, activates a very specific set of genes such as the tumor suppressor WAF1, an inhibitor of cyclin-dependent kinases (CDKs), and suppresses the growth of these tumors. In addition, this drug downregulates cyclin D1, which is upregulated by RAS through PAK1, in breast cancers. In this study, we demonstrate that FK228 at 0.1-1 nM significantly reduces the kinase activity of PAK1 in these cells, without affecting the protein level of PAK1. Interestingly, estrogen receptor (ER) and PAK1 mutually activate each other in breast cancers. Here we provide an evidence suggesting that breast cancers require PAK1 for their estrogen-dependent growth. Moreover, the treatment with FK228 strongly inhibits the estrogen-dependent growth of human breast cancers (both tamoxifen-sensitive and resistant cell lines) in vivo, suggesting that FK228 and other anti-PAK1 drugs would be useful for the treatment of breast cancers which become resistant to currently used estrogen antagonists such as tamoxifen.

  11. p-21 activated kinase 4 (PAK4) maintains stem cell-like phenotypes in pancreatic cancer cells through activation of STAT3 signaling

    PubMed Central

    Tyagi, Nikhil; Marimuthu, Saravanakumar; Bhardwaj, Arun; Deshmukh, Sachin K.; Srivastava, Sanjeev K.; Singh, Ajay P.; McClellan, Steven; Carter, James E.; Singh, Seema

    2015-01-01

    Pancreatic cancer (PC) remains a highly lethal malignancy due to its unusual chemoresistance and high aggressiveness. A subpopulation of pancreatic tumor cells, known as cancer stem cells (CSCs), is considered responsible not only for tumor-maintenance, but also for its widespread metastasis and therapeutic failure. Here we investigated the role of p-21 activated kinase 4 (PAK4) in driving PC stemness properties. Our data demonstrate that triple-positive (CD24+/CD44+/EpCAM+) subpopulation of pancreatic CSCs exhibits greater level of PAK4 as compared to triple-negative (CD24−/CD44−/EpCAM−) cells. Moreover, PAK4 silencing in PC cells leads to diminished fraction of CD24, CD44, and EpCAM positive cells. Furthermore, we show that PAK4-silenced PC cells exhibit decreased sphere-forming ability and increased chemo-sensitivity to gemcitabine toxicity. PAK4 expression is also associated with enhanced levels of stemness-associated transcription factors (Oct4/Nanog/Sox2 and KLF4). Furthermore, our data show decreased nuclear accumulation and transcriptional activity of STAT3 in PAK4-silenced PC cells and restitution of its activity leads to restoration of stem cell phenotypes. Together, our findings deliver first experimental evidence for the involvement of PAK4 in PC stemness and support its clinical utility as a novel therapeutic target in PC. PMID:26546043

  12. PREX1 Protein Function Is Negatively Regulated Downstream of Receptor Tyrosine Kinase Activation by p21-activated Kinases (PAKs).

    PubMed

    Barrows, Douglas; He, John Z; Parsons, Ramon

    2016-09-16

    Downstream of receptor tyrosine kinase and G protein-coupled receptor (GPCR) stimulation, the phosphatidylinositol 3,4,5-trisphosphate (PIP3)-dependent Rac exchange factor (PREX) family of guanine nucleotide exchange factors (GEFs) activates Rho GTPases, leading to important roles for PREX proteins in numerous cellular processes and diseases, including cancer. PREX1 and PREX2 GEF activity is activated by the second messengers PIP3 and Gβγ, and further regulation of PREX GEF activity occurs by phosphorylation. Stimulation of receptor tyrosine kinases by neuregulin and insulin-like growth factor 1 (IGF1) leads to the phosphorylation of PREX1; however, the kinases that phosphorylate PREX1 downstream of these ligands are not known. We recently reported that the p21-activated kinases (PAKs), which are activated by GTP-bound Ras-related C3 botulinum toxin substrate 1 (Rac1), mediate the phosphorylation of PREX2 after insulin receptor activation. Here we show that certain phosphorylation events on PREX1 after insulin, neuregulin, and IGF1 treatment are PAK-dependent and lead to a reduction in PREX1 binding to PIP3 Like PREX2, PAK-mediated phosphorylation also negatively regulates PREX1 GEF activity. Furthermore, the onset of PREX1 phosphorylation was delayed compared with the phosphorylation of AKT, supporting a model of negative feedback downstream of PREX1 activation. We also found that the phosphorylation of PREX1 after isoproterenol and prostaglandin E2-mediated GPCR activation is partially PAK-dependent and likely also involves protein kinase A, which is known to reduce PREX1 function. Our data point to multiple mechanisms of PREX1 negative regulation by PAKs within receptor tyrosine kinase and GPCR-stimulated signaling pathways that have important roles in diseases such as diabetes and cancer.

  13. Postsynaptic density scaffold SAP102 regulates cortical synapse development through EphB and PAK signaling pathway

    PubMed Central

    Murata, Yasunobu; Constantine-Paton, Martha

    2013-01-01

    Membrane associated guanylate kinases (MAGUKs), including SAP102, PSD-95, PSD-93 and SAP97, are scaffolding proteins for ionotropic glutamate receptors at excitatory synapses. MAGUKs play critical roles in synaptic plasticity; however, details of signaling roles for each MAGUK remain largely unknown. Here we report that SAP102 regulates cortical synapse development through the EphB and PAK signaling pathways. Using lentivirus-delivered shRNAs, we found that SAP102 and PSD-95, but not PSD-93, are necessary for excitatory synapse formation and synaptic AMPA receptor localization in developing mouse cortical neurons. SAP102 knockdown (KD) increased numbers of elongated dendritic filopodia, which is often observed in mouse models and human patients with mental retardation. Further analysis revealed that SAP102 co-immunoprecipitated the receptor tyrosine kinase EphB2 and RacGEF Kalirin-7 in neonatal cortex, and SAP102 KD reduced surface expression and dendritic localization of EphB. Moreover, SAP102 KD prevented reorganization of actin filaments, synapse formation and synaptic AMPAR trafficking in response to EphB activation triggered by its ligand ephrinB. Lastly, p21-activated kinases (PAKs) were down-regulated in SAP102 KD neurons. These results demonstrate that SAP102 has unique roles in cortical synapse development by mediating EphB and its downstream PAK signaling pathway. Both SAP102 and PAKs are associated with X-linked mental retardation in humans; thus, synapse formation mediated by EphB/SAP102/PAK signaling in the early postnatal brain may be crucial for cognitive development. PMID:23486974

  14. A novel interaction of PAK4 with PPARγ to regulate Nox1 and radiation-induced epithelial-to-mesenchymal transition in glioma.

    PubMed

    Kesanakurti, D; Maddirela, D; Banasavadi-Siddegowda, Y K; Lai, T-H; Qamri, Z; Jacob, N K; Sampath, D; Mohanam, S; Kaur, B; Puduvalli, V K

    2017-09-14

    Tumor recurrence in glioblastoma (GBM) is, in part, attributed to increased epithelial-to-mesenchymal transition (EMT) and enhanced tumor cell dissemination in adjacent brain parenchyma after ionizing radiation (IR). EMT is associated with aggressive behavior, increased stem-like characteristics and treatment resistance in malignancies; however, the underlying signaling mechanisms that regulate EMT are poorly understood. We identified grade-dependent p21-activated kinases 4 (PAK4) upregulation in gliomas and further determined its role in mesenchymal transition and radioresistance. IR treatment significantly elevated expression and nuclear localization of PAK4 in correlation with induction of reactive oxygen species (ROS) and mesenchymal transition in GBM cells. Stable PAK4 overexpression promoted mesenchymal transition by elevating EMT marker expression in these cells. Of note, transcription factor-DNA-binding arrays and chromatin immunoprecipitation experiments identified the formation of a novel nuclear PAK4/PPARγ complex which was recruited to the promoter of Nox1, a peroxisome proliferator-activated receptor gamma (PPARγ) target gene. In addition, IR further elevated PAK4/PPARγ complex co-recruitment to Nox1 promoter, and increased Nox1 expression and ROS levels associated with mesenchymal transition in these cells. Conversely, specific PAK4 downregulation decreased PPARγ-mediated Nox1 expression and suppressed EMT in IR-treated cells. In vivo orthotopic tumor experiments showed inhibition of growth and suppression of IR-induced PPARγ and Nox1 expression by PAK4 downregulation in tumors. Our results provide the first evidence of a novel role for PAK4 in IR-induced EMT and suggest potential therapeutic efficacy of targeting PAK4 to overcome radioresistance in gliomas.

  15. The Pakistan National Emergency Department Surveillance Study (Pak-NEDS): Introducing a pilot surveillance

    PubMed Central

    2015-01-01

    Background Evidence-based decision making is essential for appropriate prioritization and service provision by healthcare systems. Despite higher demands, data needs for this practice are not met in many cases in low- and middle-income countries because of underdeveloped sources, among other reasons. Emergency departments (EDs) provide an important channel for such information because of their strategic position within healthcare systems. This paper describes the design and pilot test of a national ED based surveillance system suitable for the Pakistani context. Methods The Pakistan National Emergency Department Surveillance Study (Pak-NEDS) was pilot tested in the emergency departments of seven major tertiary healthcare centres across the country. The Aga Khan University, Karachi, served as the coordinating centre. Key stakeholders and experts from all study institutes were involved in outlining data needs, development of the study questionnaire, and identification of appropriate surveillance mechanisms such as methods for data collection, monitoring, and quality assurance procedures. The surveillance system was operational between November 2010 and March 2011. Active surveillance was done 24 hours a day by data collectors hired and trained specifically for the study. All patients presenting to the study EDs were eligible participants. Over 270,000 cases were registered in the surveillance system over a period of four months. Coverage levels in the final month ranged from 91-100% and were highest in centres with the least volume of patients. Overall the coverage for the four months was 79% and crude operational costs were less than $0.20 per patient. Conclusions Pak-NEDS is the first multi-centre ED based surveillance system successfully piloted in a sample of major EDs having some of the highest patient volumes in Pakistan. Despite the challenges identified, our pilot shows that the system is flexible and scalable, and could potentially be adapted for many other

  16. PAK1 interacts with beta-catenin and is required for the regulation of the beta-catenin signalling pathway by gastrins.

    PubMed

    He, Hong; Shulkes, Arthur; Baldwin, Graham S

    2008-10-01

    Beta-catenin regulates cell-cell adhesion by binding to E-cadherin at the cell membrane and, when translocated into the nucleus, mediates signalling by activation of transcription factors such as TCF4. Mutations of the components of the Wnt/beta-catenin pathway are found in many gastrointestinal cancers. Gastrins, including amidated (Gamide) and glycine-extended (Ggly) gastrin(17), stimulate the proliferation of gastrointestinal cancer cells. Gastrins also regulate beta-catenin signalling through multiple pathways which seem to converge on p21-activated kinase 1 (PAK1). In this study, we have investigated the role of PAK1 in the regulation of beta-catenin signalling by gastrins. Here we report for the first time that PAK1 associated with beta-catenin. Both Gamide and Ggly stimulated the phosphorylation and activation of beta-catenin in a PAK1-dependent manner. A kinase-inactive mutant PAK1(K299A) blocked the gastrin-stimulated dissociation of beta-catenin from E-cadherin, translocation of beta-catenin from the cell membrane to the nucleus, and association of beta-catenin with the transcription factor TCF4. The PAK1(K299A) mutant also inhibited the stimulation of the expression of c-myc and cyclin D1, and of cell proliferation and migration, by gastrins. The results indicate that gastrins regulate beta-catenin signalling through a PAK1-dependent pathway. PAK1 seems to be the point of convergence of multiple signalling pathways activated by gastrins.

  17. Clinical evaluation of the HeartPak. A new pneumatic portable driver for use with the HeartMate Implantable Pneumatic Left Ventricular Assist System.

    PubMed

    Tamez, D; Myers, T J; Inman, R W; Miller, K A; Frazier, O H

    1997-01-01

    The HeartPak Portable Pneumatic Driver was designed for use with the HeartMate Implantable Pneumatic Left Ventricular Assist Device (IP-LVAS) (Thermo Cardiosystems, Inc., Woburn, MA). The HeartPak measures 48 x 23 x 15 cm, weighs 9.3 kg with batteries, and can be carried by a handle, by a shoulder strap, or on a trolley. Four 12 V batteries provide power for as long as 8 hr. To test the HeartPak in the hospital environment, seven men were studied who were bridge-to-transplant patients (mean age, 59.8 +/- 8.87 years) undergoing HeartMate IP-LVAS therapy. They were supported by the HeartPak for 429 days with a cycle count of 57,826,560. To normalize the mean pump flow rate, we used the body surface area to obtain a pump flow index in each case. The mean flow rate was 2.65 +/- 0.57 L/min/m2 for the HeartPak vs. 2.64 +/- 0.45 L/min/m2 for the HeartMate 1000, the conventional driver previously used in these patients. The only potentially serious problem with the HeartPak was console failure in one case. The patient took appropriate backup measures, and the HeartPak was replaced. In no case did the device cause any adverse effects or interruption of LVAS support. Compared with HeartMate 1000, the HeartPak was more convenient, easier to operate, and allowed better patient mobility.

  18. Pak4 Is Required during Epithelial Polarity Remodeling through Regulating AJ Stability and Bazooka Retention at the ZA

    PubMed Central

    Walther, Rhian F.; Nunes de Almeida, Francisca; Vlassaks, Evi; Burden, Jemima J.; Pichaud, Franck

    2016-01-01

    Summary The ability of epithelial cells to assemble into sheets relies on their zonula adherens (ZA), a circumferential belt of adherens junction (AJ) material, which can be remodeled during development to shape organs. Here, we show that during ZA remodeling in a model neuroepithelial cell, the Cdc42 effector P21-activated kinase 4 (Pak4/Mbt) regulates AJ morphogenesis and stability through β-catenin (β-cat/Arm) phosphorylation. We find that β-catenin phosphorylation by Mbt, and associated AJ morphogenesis, is needed for the retention of the apical determinant Par3/Bazooka at the remodeling ZA. Importantly, this retention mechanism functions together with Par1-dependent lateral exclusion of Par3/Bazooka to regulate apical membrane differentiation. Our results reveal an important functional link between Pak4, AJ material morphogenesis, and polarity remodeling during organogenesis downstream of Par3. PMID:27052178

  19. Activity release from damaged fuel during the Paks-2 cleaning tank incident in the spent fuel storage pool

    NASA Astrophysics Data System (ADS)

    Hózer, Zoltán; Szabó, Emese; Pintér, Tamás; Varjú, Ilona Baracska; Bujtás, Tibor; Farkas, Gábor; Vajda, Nóra

    2009-07-01

    During crud removal operations the integrity of 30 fuel assemblies was lost at high temperature at the unit No. 2 of the Paks NPP. Part of the fission products was released from the damaged fuel into the coolant of the spent fuel storage pool. The gaseous fission products escaped through the chimney from the reactor hall. The volatile and non-volatile materials remained mainly in the coolant and were collected on the filters of water purification system. The activity release from damaged fuel rods during the Paks-2 cleaning tank incident was estimated on the basis of coolant activity concentration measurements and chimney activity data. The typical release rate of noble gases, iodine and caesium was 1-3%. The release of non-volatile fission products and actinides was also detected.

  20. The endocytic adapter E-Syt2 recruits the p21 GTPase activated kinase PAK1 to mediate actin dynamics and FGF signalling

    PubMed Central

    Jean, Steve; Tremblay, Michel G.; Herdman, Chelsea; Guillou, François; Moss, Tom

    2012-01-01

    Summary Fibroblast growth factor (FGF) signalling plays an essential role in early vertebrate development. However, the response to FGF requires endocytosis of the activated FGF receptor (FGFR) that is in part dependent on remodelling of the actin cytoskeleton. Recently we showed that the extended synaptotagmin family plasma membrane protein, E-Syt2, is an essential endocytic adapter for FGFR1. Here we show E-Syt2 is also an interaction partner for the p21-GTPase Activated Kinase PAK1. The phospholipid binding C2C domain of E-Syt2 specifically binds a site adjacent to the CRIB/GBD of PAK1. PAK1 and E-Syt2 selectively complex with FGFR1 and functionally cooperate in the FGF signalling. E-Syt2 binding suppresses actin polymerization and inhibits the activation of PAK1 by the GTPases Cdc42 and Rac. Interestingly, the E-Syt2 binding site on PAK1 extensively overlaps a site recently suggested to bind phospholipids. Our data suggest that PAK1 interacts with phospholipid membrane domains via E-Syt2, where it may cooperate in the E-Syt2-dependent endocytosis of activated FGFR1 by modulating cortical actin stability. PMID:23213466

  1. Overexpression of a novel activator of PAK4, the CDK5 kinase-associated protein CDK5RAP3, promotes hepatocellular carcinoma metastasis.

    PubMed

    Mak, Grace Wing-Yan; Chan, Mandy Man-Lok; Leong, Veronica Yee-Law; Lee, Joyce Man-Fong; Yau, Tai-On; Ng, Irene Oi-Lin; Ching, Yick-Pang

    2011-04-15

    The CDK5 kinase regulatory subunit-associated protein 3 (CDK5RAP3 or C53/LZAP) regulates apoptosis induced by genotoxic stress. Although CDK5RAP3 has been implicated in cancer progression, its exact role in carcinogenesis is not well established. In this article, we report that CDK5RAP3 has an important prometastatic function in hepatocarcinogenesis. An examination of human hepatocellular carcinoma (HCC) samples revealed at least twofold overexpression of CDK5RAP3 transcripts in 58% (39/67) of HCC specimens when compared with corresponding nontumorous livers. CDK5RAP3 overexpression was associated with more aggressive biological behavior. In HCC cell lines, stable overexpression of CDK5RAP3 promoted, and small interfering RNA-mediated knockdown inhibited, tumorigenic activity and metastatic potential. We found that overexpression of CDK5RAP3 and p21-activated protein kinase 4 (PAK4) correlated in human HCCs, and that CDK5RAP3 was a novel binding partner of PAK4, and this binding enhanced PAK4 activity. siRNA-mediated knockdown of PAK4 in CDK5RAP3-expressing HCC cells reversed the enhanced cell invasiveness mediated by CDK5RAP3 overexpression, implying that PAK4 is essential for CDK5RAP3 function. Taken together, our findings reveal that CDK5RAP3 is widely overexpressed in HCC and that overexpression of CDK5RAP3 promotes HCC metastasis through PAK4 activation.

  2. Artepillin C (ARC) in Brazilian green propolis selectively blocks oncogenic PAK1 signaling and suppresses the growth of NF tumors in mice.

    PubMed

    Messerli, Shanta M; Ahn, Mok-Ryeon; Kunimasa, Kazuhiro; Yanagihara, Miyako; Tatefuji, Tomoki; Hashimoto, Ken; Mautner, Victor; Uto, Yoshihiro; Hori, Hitoshi; Kumazawa, Shigenori; Kaji, Kazuhiko; Ohta, Toshiro; Maruta, Hiroshi

    2009-03-01

    There are mainly three types of propolis whose major anticancer ingredients are entirely different: (1) CAPE (caffeic acid phenethyl ester)-based propolis in Europe, Far East and New Zealand, (2) artepillin C (ARC)-based Brazilian green propolis and (3) Brazilian red propolis. It was shown previously that NF (neurofibromatosis)-associated tumors require the kinase PAK1 for their growth, and CAPE-based propolis extracts such as Bio 30 suppress completely the growth of NF tumors in vivo by blocking PAK1 signaling. Also it was demonstrated that ARC suppresses angiogenesis, suggesting the possibility that ARC also blocks oncogenic PAK1 signaling. Here it is shown for the first time that both ARC and green propolis extract (GPE) indeed block the PAK1 signaling selectively, without affecting another kinase known as AKT. Furthermore, it was confirmed that ARC as well as GPE suppress almost completely the growth of human NF tumor xenografts in mice, as does Bio 30. These results suggest that both CAPE-based and ARC-based propolis extracts are natural anti-PAK1 remedies and could be among the first effective NF therapeutics available on the market. Since more than 70% of human cancers such as breast and prostate cancers require the kinase PAK1 for their growth, it is quite possible that GPE could be potentially useful for the treatment of these cancers, as is Bio 30. (c) 2008 John Wiley & Sons, Ltd.

  3. Towards the Seismic Hazard Reassessment of Paks NPP (Hungary) Site: Seismicity and Sensitivity Studies

    NASA Astrophysics Data System (ADS)

    Toth, Laszlo; Monus, Peter; Gyori, Erzsebet; Grenerczy, Gyula; Janos Katona, Tamas; Kiszely, Marta

    2015-04-01

    In context of extension of Paks Nuclear Power Plant by new units, a comprehensive site seismic hazard evaluation program has been developed that is already approved by the Hungarian Authorities. This includes a 3D seismic survey, drilling of several deep boreholes, extensive geological mapping, and geophysical investigations at the site and its vicinity, as well as on near regional, and regional scale. Furthermore, all relevant techniques of modern space geodesy (GPS, PSInSAR) will be also utilized to construct a new seismotectonic model. The implementation of the project is still in progress. In the presentation, some important elements of the new seismic hazard assessment are highlighted, and some results obtained in the preliminary phase of the program are presented and discussed. The first and most important component of the program is the compilation of the seismological database that is developed on different time scale zooming on different event recurrence rates such as paleo-earthquakes (10-1/a). In 1995, Paks NPP installed and started to operate a sensitive microseismic monitoring network capable for locating earthquakes as small as magnitude 2.0 within about 100 km of the NPP site. During the two decades of operation, the microseismic monitoring network located some 2,000 earthquakes within the region of latitude 45.5 - 49 N and longitude 16 - 23 E. Out of the total number of events, 130 earthquakes were reported as 'felt events'. The largest earthquake was an event of ML 4.8, causing significant damage in the epicenter area. The results of microseismic monitoring provide valuable data for seismotectonic modelling and results in more accurate earthquake recurrence equations. The first modern PSHA of Paks NPP site was carried out in 1995. Complex site characterization project was implemented and hazard curves had been evaluated for 10-3 - 10-5 annual frequency. As a follow-up, PSHA results have been reviewed and updated in the frame of periodic safety

  4. Oncogenic epithelial cell-derived exosomes containing Rac1 and PAK2 induce angiogenesis in recipient endothelial cells

    PubMed Central

    Gopal, Shashi K.; Greening, David W.; Hanssen, Eric G.; Zhu, Hong-Jian; Simpson, Richard J.; Mathias, Rommel A.

    2016-01-01

    The metastatic cascade describes the escape of primary tumour cells to distant secondary sites. Cells at the leading tumour edge are thought to undergo epithelial-mesenchymal transition (EMT), to enhance their motility and invasion for spreading. Whether EMT cells directly promote tumour angiogenesis, and the role of exosomes (30-150 nm extracellular vesicles) remains largely unknown. We examined the functional effects of exosomes from MDCK cells, MDCK cells stably expressing YBX1 (MDCKYBX1, intermediate EMT), and Ras-transformed MDCK cells (21D1 cells, complete EMT). 2F-2B cell motility and tube formation (length and branching) was significantly increased following supplementation with MDCKYBX1 or 21D1 exosomes, but not MDCK exosomes. Next, Matrigel™ plugs containing exosome-supplemented 2F-2B cells were subcutaneously injected into mice. Systemic perfusion was only observed for plugs supplemented with MDCKYBX1 or 21D1 exosomes. Comparative proteomics revealed that 21D1 exosomes contained VEGF-associated proteins, while MDCKYBX1 exosomes were enriched with activated Rac1 and PAK2. To validate, 2F-2B cells and HUVECs were pre-treated with PAK inhibitors prior to exosome supplementation. PAK inhibition nullified the effects of MDCKYBX1 exosomes by reducing the tube length and branching to baseline levels. By contrast, the effects of 21D1 exosomes were not significantly decreased. Our results demonstrate for the first time that oncogenic cells undergoing EMT can communicate with endothelial cells via exosomes, and establish exosomal Rac1/PAK2 as angiogenic promoters that may function from early stages of the metastatic cascade. PMID:26919098

  5. p21-activated Kinases (PAKs) Mediate the Phosphorylation of PREX2 Protein to Initiate Feedback Inhibition of Rac1 GTPase.

    PubMed

    Barrows, Douglas; Schoenfeld, Sarah M; Hodakoski, Cindy; Silkov, Antonina; Honig, Barry; Couvillon, Anthony; Shymanets, Aliaksei; Nürnberg, Bernd; Asara, John M; Parsons, Ramon

    2015-11-27

    Phosphatidylinositol 3,4,5-trisphosphate (PIP3)-dependent Rac exchanger 2 (PREX2) is a guanine nucleotide exchange factor (GEF) for the Ras-related C3 botulinum toxin substrate 1 (Rac1) GTPase, facilitating the exchange of GDP for GTP on Rac1. GTP-bound Rac1 then activates its downstream effectors, including p21-activated kinases (PAKs). PREX2 and Rac1 are frequently mutated in cancer and have key roles within the insulin-signaling pathway. Rac1 can be inactivated by multiple mechanisms; however, negative regulation by insulin is not well understood. Here, we show that in response to being activated after insulin stimulation, Rac1 initiates its own inactivation by decreasing PREX2 GEF activity. Following PREX2-mediated activation of Rac1 by the second messengers PIP3 or Gβγ, we found that PREX2 was phosphorylated through a PAK-dependent mechanism. PAK-mediated phosphorylation of PREX2 reduced GEF activity toward Rac1 by inhibiting PREX2 binding to PIP3 and Gβγ. Cell fractionation experiments also revealed that phosphorylation prevented PREX2 from localizing to the cellular membrane. Furthermore, the onset of insulin-induced phosphorylation of PREX2 was delayed compared with AKT. Altogether, we propose that second messengers activate the Rac1 signal, which sets in motion a cascade whereby PAKs phosphorylate and negatively regulate PREX2 to decrease Rac1 activation. This type of regulation would allow for transient activation of the PREX2-Rac1 signal and may be relevant in multiple physiological processes, including diseases such as diabetes and cancer when insulin signaling is chronically activated.

  6. p21-activated Kinases (PAKs) Mediate the Phosphorylation of PREX2 Protein to Initiate Feedback Inhibition of Rac1 GTPase*

    PubMed Central

    Barrows, Douglas; Schoenfeld, Sarah M.; Hodakoski, Cindy; Silkov, Antonina; Honig, Barry; Couvillon, Anthony; Shymanets, Aliaksei; Nürnberg, Bernd; Asara, John M.; Parsons, Ramon

    2015-01-01

    Phosphatidylinositol 3,4,5-trisphosphate (PIP3)-dependent Rac exchanger 2 (PREX2) is a guanine nucleotide exchange factor (GEF) for the Ras-related C3 botulinum toxin substrate 1 (Rac1) GTPase, facilitating the exchange of GDP for GTP on Rac1. GTP-bound Rac1 then activates its downstream effectors, including p21-activated kinases (PAKs). PREX2 and Rac1 are frequently mutated in cancer and have key roles within the insulin-signaling pathway. Rac1 can be inactivated by multiple mechanisms; however, negative regulation by insulin is not well understood. Here, we show that in response to being activated after insulin stimulation, Rac1 initiates its own inactivation by decreasing PREX2 GEF activity. Following PREX2-mediated activation of Rac1 by the second messengers PIP3 or Gβγ, we found that PREX2 was phosphorylated through a PAK-dependent mechanism. PAK-mediated phosphorylation of PREX2 reduced GEF activity toward Rac1 by inhibiting PREX2 binding to PIP3 and Gβγ. Cell fractionation experiments also revealed that phosphorylation prevented PREX2 from localizing to the cellular membrane. Furthermore, the onset of insulin-induced phosphorylation of PREX2 was delayed compared with AKT. Altogether, we propose that second messengers activate the Rac1 signal, which sets in motion a cascade whereby PAKs phosphorylate and negatively regulate PREX2 to decrease Rac1 activation. This type of regulation would allow for transient activation of the PREX2-Rac1 signal and may be relevant in multiple physiological processes, including diseases such as diabetes and cancer when insulin signaling is chronically activated. PMID:26438819

  7. SparsePak: A Formatted Fiber Field Unit for The WIYN Telescope Bench Spectrograph. II. On-Sky Performance

    NASA Astrophysics Data System (ADS)

    Bershady, Matthew A.; Andersen, David R.; Verheijen, Marc A. W.; Westfall, Kyle B.; Crawford, Steven M.; Swaters, Rob A.

    2005-02-01

    We present a performance analysis of SparsePak and the WIYN Bench Spectrograph for precision studies of stellar and ionized gas kinematics of external galaxies. We focus on spectrograph configurations with echelle and low-order gratings yielding spectral resolutions of ~10,000 between 500 and 900 nm. These configurations are of general relevance to the spectrograph performance. Benchmarks include spectral resolution, sampling, vignetting, scattered light, and an estimate of the system absolute throughput. Comparisons are made to other, existing, fiber feeds on the WIYN Bench Spectrograph. Vignetting and relative throughput are found to agree with a geometric model of the optical system. An aperture-correction protocol for spectrophotometric standard-star calibrations has been established using independent WIYN imaging data and the unique capabilities of the SparsePak fiber array. The WIYN point-spread function is well fitted by a Moffat profile with a constant power-law outer slope of index -4.4. We use SparsePak commissioning data to debunk a long-standing myth concerning sky-subtraction with fibers: by properly treating the multifiber data as a ``long-slit'' it is possible to achieve precision sky-subtraction with a signal-to-noise performance as good or better than conventional long-slit spectroscopy. No beam-switching is required, and hence the method is efficient. Finally, we give several examples of science measurements that SparsePak now makes routine. These include Hα velocity fields of low surface brightness disks, gas and stellar velocity-fields of nearly face-on disks, and stellar absorption-line profiles of galaxy disks at spectral resolutions of ~24,000.

  8. Mangrove Colonization: Mangrove Progression Over the Growing Pak Phanang (SE Thailand) Mud Flat

    NASA Astrophysics Data System (ADS)

    Panapitukkul, N.; Duarte, C. M.; Thampanya, U.; Kheowvongsri, P.; Srichai, N.; Geertz-Hansen, O.; Terrados, J.; Boromthanarath, S.

    1998-07-01

    A combination of remote sensing techniques and in situmeasurements along a chronosequence was used to elucidate the rate of progression of the mangrove forest in the Pak Phanang Bay (SE Thailand), a large bay with an extended and rapidly accreting mud flat. The examination of black and white aerial photographs of the forest in 1966, 1974, 1989 and 1995, and satellite images in 1985, 1990 and 1994 revealed that the mangrove forest located in the eastern bank of the bay was progressing over the mud flat. The rate of progression was estimated, from examination of changes in the position of the forest edge with time in the series of images, to average 38·6 m year -1over the 28-year interval encompassed by the images. Mangrove progression rates were fastest between 1966 and 1974 and slowest between 1974 and 1985, remaining uniform at about 30 m year -1thereafter. The in situexamination of vegetation along transects in the area of fastest mangrove progression showed an average progression rate of 53·12±5·86 m year -1, quite similar to the estimate (48·4 m year -1) derived from remote sensing techniques for the area where the transects were surveyed. Avicennia albawas found to dominate the vegetation at the progressing edge of the mangrove, followed by Sonneratia caseolaris, with Rhizophora apiculatabeing present only occasionally. The fast colonization of A. albaover the mud flat was supported by a large export flux of mangrove propagules from the channels draining the mangrove forest, which averaged 3715±920 and 1900±808 fruits day -1in each of the channels examined. Extrapolation of the long-term mean mangrove progression rate observed along the eastern bank of the Pak Phanang Bay suggested that this mangrove forest will increase by 33 ha year -1. These results provide evidence that natural mangrove colonization can be a rapid process if sufficient propagules of the pioneer species ( A. albaand S. caseolaris) are available, and point, therefore, to alternative

  9. Specification of Dendritogenesis Site in Drosophila aCC Motoneuron by Membrane Enrichment of Pak1 through Dscam1.

    PubMed

    Kamiyama, Daichi; McGorty, Ryan; Kamiyama, Rie; Kim, Michael D; Chiba, Akira; Huang, Bo

    2015-10-12

    Precise positioning of dendritic branches is a critical step in the establishment of neuronal circuitry. However, there is limited knowledge on how environmental cues translate into dendrite initiation or branching at a specific position. Here, through a combination of mutation, RNAi, and imaging experiments, we found that a Dscam-Dock-Pak1 hierarchical interaction defines the stereotypical dendrite growth site in the Drosophila aCC motoneuron. This interaction localizes the Cdc42 effector Pak1 to the plasma membrane at the dendrite initiation site before the activation of Cdc42. Ectopic expression of membrane-anchored Pak1 overrides this spatial specification of dendritogenesis, confirming its function in guiding Cdc42 signaling. We further discovered that Dscam1 localization in aCC occurs through an inter-neuronal contact that involves Dscam1 in the partner MP1 neuron. These findings elucidate a mechanism by which Dscam1 controls neuronal morphogenesis through spatial regulation of Cdc42 signaling and, subsequently, cytoskeletal remodeling. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. PAK1 and CtBP1 Regulate the Coupling of Neuronal Activity to Muscle Chromatin and Gene Expression

    PubMed Central

    Thomas, Jean-Luc; Ravel-Chapuis, Aymeric; Valente, Carmen; Corda, Daniela; Méjat, Alexandre

    2015-01-01

    Acetylcholine receptor (AChR) expression in innervated muscle is limited to the synaptic region. Neuron-induced electrical activity participates in this compartmentalization by promoting the repression of AChR expression in the extrasynaptic regions. Here, we show that the corepressor CtBP1 (C-terminal binding protein 1) is present on the myogenin promoter together with repressive histone marks. shRNA-mediated downregulation of CtBP1 expression is sufficient to derepress myogenin and AChR expression in innervated muscle. Upon denervation, CtBP1 is displaced from the myogenin promoter and relocates to the cytoplasm, while repressive histone marks are replaced by activating ones concomitantly to the activation of myogenin expression. We also observed that upon denervation the p21-activated kinase 1 (PAK1) expression is upregulated, suggesting that phosphorylation by PAK1 may be involved in the relocation of CtBP1. Indeed, preventing CtBP1 Ser158 phosphorylation induces CtBP1 accumulation in the nuclei and abrogates the activation of myogenin and AChR expression. Altogether, these findings reveal a molecular mechanism to account for the coordinated control of chromatin modifications and muscle gene expression by presynaptic neurons via a PAK1/CtBP1 pathway. PMID:26416879

  11. Cultivar-Specific Changes in Primary and Secondary Metabolites in Pak Choi (Brassica Rapa, Chinensis Group) by Methyl Jasmonate

    PubMed Central

    Kim, Moo Jung; Chiu, Yu-Chun; Kim, Na Kyung; Park, Hye Min; Lee, Choong Hwan; Juvik, John A.; Ku, Kang-Mo

    2017-01-01

    Glucosinolates, their hydrolysis products and primary metabolites were analyzed in five pak choi cultivars to determine the effect of methyl jasmonate (MeJA) on metabolite flux from primary metabolites to glucosinolates and their hydrolysis products. Among detected glucosinolates (total 14 glucosinolates; 9 aliphatic, 4 indole and 1 aromatic glucosinolates), indole glucosinolate concentrations (153–229%) and their hydrolysis products increased with MeJA treatment. Changes in the total isothiocyanates by MeJA were associated with epithiospecifier protein activity estimated as nitrile formation. Goitrin, a goitrogenic compound, significantly decreased by MeJA treatment in all cultivars. Changes in glucosinolates, especially aliphatic, significantly differed among cultivars. Primary metabolites including amino acids, organic acids and sugars also changed with MeJA treatment in a cultivar-specific manner. A decreased sugar level suggests that they might be a carbon source for secondary metabolite biosynthesis in MeJA-treated pak choi. The result of the present study suggests that MeJA can be an effective agent to elevate indole glucosinolates and their hydrolysis products and to reduce a goitrogenic compound in pak choi. The total glucosinolate concentration was the highest in “Chinese cabbage” in the control group (32.5 µmol/g DW), but indole glucosinolates increased the greatest in “Asian” when treated with MeJA. PMID:28481284

  12. Industrial-hygiene walk-through survey report of Tetra Pak, Inc. , Denton, Texas

    SciTech Connect

    McCammon, C.S.; Krishnan, E.R.; Goodman, R.J.

    1987-06-05

    A walk-through industrial-hygiene survey was conducted at the Tetra Pak Denton facility in Denton, Texas to determine possible employee exposure to acrylates or methacrylates. Acrylated inks and coatings have been used at this facility since 1984 to produce aseptic flexible packaging material for the food industry. There were three offset printing press lines used at the company, each with five offset presses to apply different colored inks. As many as five separate wet-on-wet applications of acrylated ink formulations sometimes preceded the coating application. After the inks were applied they were passed through an offset blanket coater. Following this step an electron-beam-curing unit radiated the inks and coating. Environmental enclosures surrounded the offset-printing presses in order to cut down on noise and mist. Of the 176 employees at this company, 37 had potential contact with acrylates. No air monitoring has been conducted at this facility for acrylates. One case of dermatitis arose at the company since 1984 and possibly was related directly to skin contact with acrylates. There was a safety program in place at the company and personnel records were maintained for each employee.

  13. Reactor Dosimetry Aspects of the Service Life Extension of the Hungarian Paks NPP

    NASA Astrophysics Data System (ADS)

    Zsolnay, Eva M.; Czifrus, Szabolcs; Fehér, Sándor; Hordósy, Gábor; Keresztúri, András; Kresz, Norbert; Oszvald, Ferenc

    2016-02-01

    The service life of the Hungarian Paks Nuclear Power Plant (NPP) will be extended from the originally planned 30 years to 50 years. To improve the reliability of the results obtained in frame of the old reactor pressure vessel (RPV) surveillance programme, new methods have been developed, and based on them, the old exposition data have been re-evaluated for all the four reactor units. At the same time, a new RPV surveillance programme has been developed and introduced, and long term irradiations have been performed to determine the radiation damage of the surveillance specimens due to the high fast neutron exposition. Neutron transport calculations have been performed with a validated neutron transport code system to determine the fast neutron exposition of the RPVs during the extended service life. The cavity dosimetry is in the introductory phase. This paper presents the new developments in the field of the RPV surveillance dosimetry and summarises the results obtained. According to the results the service life of the NPP can safely be extended for the planned 50 years.

  14. Geochemistry and tectonic setting of the Central Loei volcanic rocks, Pak Chom area, Loei, northeastern Thailand

    NASA Astrophysics Data System (ADS)

    Panjasawatwong, Y.; Zaw, Khin; Chantaramee, S.; Limtrakun, P.; Pirarai, K.

    2006-01-01

    The Central Loei volcanic rocks, as evidenced by those in the Pak Chom area, were formed in the Late Devonian-Early Carboniferous and can be separated into three magmatic groups: transitional tholeiitic basalt, tholeiitic microgabbro and calc-alkalic basalt/andesite on the basis of immobile-element contents and ratios of least altered samples. All the tholeiitic microgabbro possibly occurred as dikes. Chemically, the transitional tholeittic basalt and tholeiitic microgabbro have higher abundances of TiO 2, Ni and Cr relative to the calc-alkalic basalt/andesite at similar values for FeO*/MgO; they also contain higher Ti/Zr but lower Zr/Nb. The transitional tholeiitic basalt has higher concentrations of P 2O 5 and Nb relative to the tholeiitic microgabbro at similar levels of FeO*/MgO, and also has higher ratios of Nb/Y and Ti/V, but lower values for Ti/Zr and Zr/Nb. In terms of chondrite normalized REE and N-MORB normalized patterns, the transitional tholeiitic basalt, tholeiitic basalt and calc-alkalic basalt/andesite are analogous to those from North Atlantic, Southwest Indian Ridge and New Britain Arc. On this basis, the Central Loei volcanic rocks are comprised of MORBs and oceanic island-arc lavas. These arc lavas may have erupted on an oceanic basement in the same ocean basin as those in the Chiang Rai-Chiang Mai volcanic belt.

  15. Chronic exposure to cigarette smoke leads to activation of p21 (RAC1)-activated kinase 6 (PAK6) in non-small cell lung cancer cells

    PubMed Central

    Syed, Nazia; Solanki, Hitendra S.; Puttamallesh, Vinuth N.; Balaji, Sai A.; Nanjappa, Vishalakshi; Datta, Keshava K.; Babu, Niraj; Renuse, Santosh; Patil, Arun H.; Izumchenko, Evgeny; Prasad, T.S. Keshava; Chang, Xiaofei; Rangarajan, Annapoorni; Sidransky, David; Pandey, Akhilesh; Gowda, Harsha; Chatterjee, Aditi

    2016-01-01

    Epidemiological data clearly establishes cigarette smoking as one of the major cause for lung cancer worldwide. Recently, targeted therapy has become one of the most preferred modes of treatment for cancer. Though certain targeted therapies such as anti-EGFR are in clinical practice, they have shown limited success in lung cancer patients who are smokers. This demands discovery of alternative drug targets through systematic investigation of cigarette smoke-induced signaling mechanisms. To study the signaling events activated in response to cigarette smoke, we carried out SILAC-based phosphoproteomic analysis of H358 lung cancer cells chronically exposed to cigarette smoke. We identified 1,812 phosphosites, of which 278 phosphosites were hyperphosphorylated (≥ 3-fold) in H358 cells chronically exposed to cigarette smoke. Our data revealed hyperphosphorylation of S560 within the conserved kinase domain of PAK6. Activation of PAK6 is associated with various processes in cancer including metastasis. Mechanistic studies revealed that inhibition of PAK6 led to reduction in cell proliferation, migration and invasion of the cigarette smoke treated cells. Further, siRNA mediated silencing of PAK6 resulted in decreased invasive abilities in a panel of non-small cell lung cancer (NSCLC) cells. Consistently, mice bearing tumor xenograft showed reduced tumor growth upon treatment with PF-3758309 (group II PAK inhibitor). Immunohistochemical analysis revealed overexpression of PAK6 in 66.6% (52/78) of NSCLC cases in tissue microarrays. Taken together, our study indicates that PAK6 is a promising novel therapeutic target for NSCLC, especially in smokers. PMID:27542207

  16. PKA-induced phosphorylation of ERα at serine 305 and high PAK1 levels is associated with sensitivity to tamoxifen in ER-positive breast cancer.

    PubMed

    Kok, Marleen; Zwart, Wilbert; Holm, Caroline; Fles, Renske; Hauptmann, Michael; Van't Veer, Laura J; Wessels, Lodewyk F A; Neefjes, Jacques; Stål, Olle; Linn, Sabine C; Landberg, Göran; Michalides, Rob

    2011-01-01

    Phosphorylation of estrogen receptor α at serine 305 (ERαS305-P) by protein kinase A (PKA) or p21-activated kinase 1 (PAK1) has experimentally been associated with tamoxifen sensitivity. Here, we investigated the clinical application of this knowledge to predict tamoxifen resistance in ER-positive breast cancer patients. Using immunohistochemistry, a score including PAK1 and co-expression of PKA and ERαS305-P (PKA/ERαS305-P) was developed on a training set consisting of 103 patients treated with tamoxifen for metastatic disease, and validated on 231 patients randomized between adjuvant tamoxifen or no treatment. In the training set, PAK1 levels were associated with tumor progression after tamoxifen (HR 1.57, 95% CI 0.99-2.48), as was co-expression of PKA and ERαS305-P (HR 2.00, 95% CI 1.14-3.52). In the validation set, a significant tamoxifen benefit was found among the 73% patients negative for PAK1 and PKA/ERαS305-P (HR 0.54, 95% CI 0.34-0.87), while others (27%) were likely to have no benefit from tamoxifen (HR 0.88, 95% 0.42-1.82). The test for interaction showed a significant difference in recurrence-free survival between groups defined by PAK1 and PKA/ERαS305-P (P = 0.037). Elevated PAK1 and PKA/ERαS305-P appeared to influence tamoxifen sensitivity. Both PAK1 and PKA/ERαS305-P levels were associated with sensitivity to tamoxifen in breast tumors and the combination of these variables should be considered in predicting tamoxifen benefit.

  17. Rainfall Trends over the Indo-Pak Summer Monsoon and Related Large-Scale Dynamics

    NASA Astrophysics Data System (ADS)

    Latif, Muhammad; Syed, Faisal; Hannachi, Abdel

    2016-04-01

    The study of regional rainfall trends over South Asia is critically important for food security and infrastructure. This study investigates the presence of trends in seasonal and sub-seasonal (June through September-JJAS) rainfall obtained from multiple observed datasets. The obtained results identified a dipole-type structure in rainfall trends over the region north of the Indo-Pak subcontinent, where significant increasing trends are seen over the core monsoon region of Pakistan and significant decreasing trends are observed over the central-north India and adjacent areas. The study strongly suggests that strengthening of Vertically Integrated Meridional Moisture Transport (VIMMT) over the Arabian Sea is likely reason for the trend of rainfall in the core monsoon region of Pakistan. In contrast, over the central-north India region, the rainfall trends are significantly decreasing due to the weakening of IMT over the Bay of Bengal. The leading EOF clearly shows the strengthening (weakening) patterns of VIMMT over the Arabian Sea (Bay of Bengal) in seasonal and sub-seasonal interannual time-scales. The regression analysis between the principal components and rainfall confirms the dipole pattern over the region. Our results also suggest that the Circumglobal Teleconnection in upper troposphere influence in maintaining the mean rainfall over Pakistan via cross-equatorial flow of moisture into the Arabian Sea. We also investigate seasonal JJAS rainfall trends using historical and climate change (RCP4.5 and RCP8.5) simulations from a set of regional climate models from Coupled Model Intercomparison Project (CMIP5). Trends and asymmetry of seasonal rainfall show great variability across models. Meridional moisture transport and associated large-scale dynamics will also be discussed.

  18. PAK2 is an effector of TSC1/2 signaling independent of mTOR and a potential therapeutic target for Tuberous Sclerosis Complex

    PubMed Central

    Alves, Maria M.; Fuhler, Gwenny M.; Queiroz, Karla C.S.; Scholma, Jetse; Goorden, Susan; Anink, Jasper; Arnold Spek, C.; Hoogeveen-Westerveld, Marianne; Bruno, Marco J.; Nellist, Mark; Elgersma, Ype; Aronica, Eleonora; Peppelenbosch, Maikel P.

    2015-01-01

    Tuberous sclerosis complex (TSC) is caused by inactivating mutations in either TSC1 or TSC2 and is characterized by uncontrolled mTORC1 activation. Drugs that reduce mTOR activity are only partially successful in the treatment of TSC, suggesting that mTOR-independent pathways play a role in disease development. Here, kinome profiles of wild-type and Tsc2−/− mouse embryonic fibroblasts (MEFs) were generated, revealing a prominent role for PAK2 in signal transduction downstream of TSC1/2. Further investigation showed that the effect of the TSC1/2 complex on PAK2 is mediated through RHEB, but is independent of mTOR and p21RAC. We also demonstrated that PAK2 over-activation is likely responsible for the migratory and cell cycle abnormalities observed in Tsc2−/− MEFs. Finally, we detected high levels of PAK2 activation in giant cells in the brains of TSC patients. These results show that PAK2 is a direct effector of TSC1-TSC2-RHEB signaling and a new target for rational drug therapy in TSC. PMID:26412398

  19. PAK2 is an effector of TSC1/2 signaling independent of mTOR and a potential therapeutic target for Tuberous Sclerosis Complex.

    PubMed

    Alves, Maria M; Fuhler, Gwenny M; Queiroz, Karla C S; Scholma, Jetse; Goorden, Susan; Anink, Jasper; Spek, C Arnold; Hoogeveen-Westerveld, Marianne; Bruno, Marco J; Nellist, Mark; Elgersma, Ype; Aronica, Eleonora; Peppelenbosch, Maikel P

    2015-09-28

    Tuberous sclerosis complex (TSC) is caused by inactivating mutations in either TSC1 or TSC2 and is characterized by uncontrolled mTORC1 activation. Drugs that reduce mTOR activity are only partially successful in the treatment of TSC, suggesting that mTOR-independent pathways play a role in disease development. Here, kinome profiles of wild-type and Tsc2(-/-) mouse embryonic fibroblasts (MEFs) were generated, revealing a prominent role for PAK2 in signal transduction downstream of TSC1/2. Further investigation showed that the effect of the TSC1/2 complex on PAK2 is mediated through RHEB, but is independent of mTOR and p21RAC. We also demonstrated that PAK2 over-activation is likely responsible for the migratory and cell cycle abnormalities observed in Tsc2(-/-) MEFs. Finally, we detected high levels of PAK2 activation in giant cells in the brains of TSC patients. These results show that PAK2 is a direct effector of TSC1-TSC2-RHEB signaling and a new target for rational drug therapy in TSC.

  20. Increased expression of microRNA-221 inhibits PAK1 in endothelial progenitor cells and impairs its function via c-Raf/MEK/ERK pathway

    SciTech Connect

    Zhang, Xiaoping; Mao, Haian; Chen, Jin-yuan; Wen, Shengjun; Li, Dan; Ye, Meng; Lv, Zhongwei

    2013-02-15

    Highlights: ► MicroRNA-221 is upregulated in the endothelial progenitor cells of atherosclerosis patients. ► PAK1 is a direct target of microRNA-221. ► MicroRNA-221 inhibits EPCs proliferation through c-Raf/MEK/ERK pathway. -- Abstract: Coronary artery disease (CAD) is associated with high mortality and occurs via endothelial injury. Endothelial progenitor cells (EPCs) restore the integrity of the endothelium and protect it from atherosclerosis. In this study, we compared the expression of microRNAs (miRNAs) in EPCs in atherosclerosis patients and normal controls. We found that miR-221 expression was significantly up-regulated in patients compared with controls. We predicted and identified p21/Cdc42/Rac1-activated kinase 1 (PAK1) as a novel target of miR-221 in EPCs. We also demonstrated that miR-221 targeted a putative binding site in the 3′UTR of PAK1, and absence of this site was inversely associated with miR-221 expression in EPCs. We confirmed this relationship using a luciferase reporter assay. Furthermore, overexpression of miR-221 in EPCs significantly decreased EPC proliferation, in accordance with the inhibitory effects induced by decreased PAK1. Overall, these findings demonstrate that miR-221 affects the MEK/ERK pathway by targeting PAK1 to inhibit the proliferation of EPCs.

  1. Flagellin Glycosylation in Pseudomonas aeruginosa PAK Requires the O-antigen Biosynthesis Enzyme WbpO*s

    PubMed Central

    Miller, Wayne L.; Matewish, Mauricia J.; McNally, David J.; Ishiyama, Noboru; Anderson, Erin M.; Brewer, Dyanne; Brisson, Jean-Robert; Berghuis, Albert M.; Lam, Joseph S.

    2010-01-01

    Pseudomonas aeruginosa PAK (serotype O6) produces a single polar, glycosylated flagellum composed of a-type flagellin. To determine whether or not flagellin glycosylation in this serotype requires O-antigen genes, flagellin was isolated from the wild type, three O-antigen-deficient mutants wbpL, wbpO, and wbpP, and a wbpO mutant complemented with a plasmid containing a wild-type copy of wbpO. Flagellin from the wbpO mutant was smaller (42 kDa) than that of the wild type (45 kDa), or other mutants strains, and exhibited an altered isoelectric point (pI 4.8) when compared with PAK flagellin (pI 4.6). These differences were because of the truncation of the glycan moiety in the wbpO-flagellin. Thus, flagellin glycosylation in P. aeruginosa PAK apparently requires a functional WbpO but not WbpP. Because WbpP was previously proposed to catalyze a metabolic step in the biosynthesis of B-band O-antigen that precedes the action of WbpO, these results prompted us to reevaluate the two-step pathway catalyzed by WbpO and WbpP. Results from WbpO-WbpP-coupled enzymatic assays showed that either WbpO or WbpP is capable of initiating the two-step pathway; however, the kinetic parameters favored the WbpO reaction to occur first, converting UDP-N-acetyl-D-glucosamine to UDP-N-acetyl-D-glucuronic acid prior to the conversion to UDP-N-acetyl-D-galacturonic acid by WbpP. This is the first report to show that a C4 epimerase could utilize UDP-N-acetylhexuronic acid as a substrate. PMID:18065759

  2. Threonine 209 phosphorylation on RUNX3 by Pak1 is a molecular switch for its dualistic functions.

    PubMed

    Kumar, A; Singhal, M; Chopra, C; Srinivasan, S; Surabhi, R P; Kanumuri, R; Tentu, S; Jagadeeshan, S; Sundaram, S; Ramanathan, K; Shankar Pitani, R; Muthuswamy, B; Abhijit, S; Nair, A S; Venkatraman, G; Rayala, S K

    2016-09-15

    P21 Activated Kinase 1 (Pak1), an oncogenic serine/threonine kinase, is known to have a significant role in the regulation of cytoskeleton and cellular morphology. Runx3 was initially known for its role in tumor suppressor function, but recent studies have reported the oncogenic role of Runx3 in various cancers. However, the mechanism that controls the paradoxical functions of Runx3 still remains unclear. In this study, we show that Runx3 is a physiologically interacting substrate of Pak1. We identified the site of phosphorylation in Runx3 as Threonine 209 by mass spectrometry analysis and site-directed mutagenesis, and further confirmed the same with a site-specific antibody. Results from our functional studies showed that Threonine 209 phosphorylation in Runx3 alters its subcellular localization by protein mislocalization from the nucleus to the cytoplasm and subsequently converses its biological functions. This was further supported by in vivo tumor xenograft studies in nude mouse models which clearly demonstrated that PANC-28 cells transfected with the Runx3-T209E clone showed high tumorigenic potential as compared with other clones. Our results from clinical samples also suggest that Threonine 209 phosphorylation by Pak1 could be a potential therapeutic target and of great clinical relevance with implications for Runx3 inactivation in cancer cells where Runx3 is known to be oncogenic. The findings presented in this study provide evidence of Runx3-Threonine 209 phosphorylation as a molecular switch in dictating the tissue-specific dualistic functions of Runx3 for the first time.

  3. A vertebrate-specific Chp-PAK-PIX pathway maintains E-cadherin at adherens junctions during zebrafish epiboly.

    PubMed

    Tay, Hwee Goon; Ng, Yuen Wai; Manser, Ed

    2010-04-12

    In early vertebrate development, embryonic tissues modulate cell adhesiveness and acto-myosin contractility to correctly orchestrate the complex processes of gastrulation. E-cadherin (E-cadh) is the earliest expressed cadherin and is needed in the mesendodermal progenitors for efficient migration. Regulatory mechanisms involving directed E-cadh trafficking have been invoked downstream of Wnt11/5 signaling. This non-canonical Wnt pathway regulates RhoA-ROK/DAAM1 to control the acto-myosin network. However, in this context nothing is known of the intracellular signals that participate in the correct localization of E-cadh, other than a need for Rab5c signaling. By studying loss of Chp induced by morpholino-oligonucleotide injection in zebrafish, we find that the vertebrate atypical Rho-GTPase Chp is essential for the proper disposition of cells in the early embryo. The underlying defect is not leading edge F-actin assembly (prominent in the cells of the envelope layer), but rather the failure to localize E-cadh and beta-catenin at the adherens junctions. Loss of Chp results in delayed epiboly that can be rescued by mRNA co-injection, and phenocopies zebrafish E-cadh mutants. This new signaling pathway involves activation of an effector kinase PAK, and involvement of the adaptor PAK-interacting exchange factor PIX. Loss of signaling by any of the three components results in similar underlying defects, which is most prominent in the epithelial-like envelope layer. Our current study uncovers a developmental pathway involving Chp/PAK/PIX signaling, which helps co-ordinate E-cadh disposition to promote proper cell adhesiveness, and coordinate movements of the three major cell layers in epiboly. Our data shows that without Chp signaling, E-cadh shifts to intracellular vesicles rather than the adhesive contacts needed for directed cell movement. These events may mirror the requirement for PAK2 signaling essential for the proper formation of the blood-brain barrier.

  4. A Vertebrate-Specific Chp-PAK-PIX Pathway Maintains E-Cadherin at Adherens Junctions during Zebrafish Epiboly

    PubMed Central

    Tay, Hwee Goon; Ng, Yuen Wai; Manser, Ed

    2010-01-01

    Background In early vertebrate development, embryonic tissues modulate cell adhesiveness and acto-myosin contractility to correctly orchestrate the complex processes of gastrulation. E-cadherin (E-cadh) is the earliest expressed cadherin and is needed in the mesendodermal progenitors for efficient migration [1], [2]. Regulatory mechanisms involving directed E-cadh trafficking have been invoked downstream of Wnt11/5 signaling [3]. This non-canonical Wnt pathway regulates RhoA-ROK/DAAM1 to control the acto-myosin network. However, in this context nothing is known of the intracellular signals that participate in the correct localization of E-cadh, other than a need for Rab5c signaling [3]. Methodology/Principal Findings By studying loss of Chp induced by morpholino-oligonucleotide injection in zebrafish, we find that the vertebrate atypical Rho-GTPase Chp is essential for the proper disposition of cells in the early embryo. The underlying defect is not leading edge F-actin assembly (prominent in the cells of the envelope layer), but rather the failure to localize E-cadh and β-catenin at the adherens junctions. Loss of Chp results in delayed epiboly that can be rescued by mRNA co-injection, and phenocopies zebrafish E-cadh mutants [4], [5]. This new signaling pathway involves activation of an effector kinase PAK, and involvement of the adaptor PAK-interacting exchange factor PIX. Loss of signaling by any of the three components results in similar underlying defects, which is most prominent in the epithelial-like envelope layer. Conclusions/Significance Our current study uncovers a developmental pathway involving Chp/PAK/PIX signaling, which helps co-ordinate E-cadh disposition to promote proper cell adhesiveness, and coordinate movements of the three major cell layers in epiboly. Our data shows that without Chp signaling, E-cadh shifts to intracellular vesicles rather than the adhesive contacts needed for directed cell movement. These events may mirror the requirement

  5. Slit stimulation recruits Dock and Pak to the roundabout receptor and increases Rac activity to regulate axon repulsion at the CNS midline.

    PubMed

    Fan, Xueping; Labrador, Juan Pablo; Hing, Huey; Bashaw, Greg J

    2003-09-25

    Drosophila Roundabout (Robo) is the founding member of a conserved family of repulsive axon guidance receptors that respond to secreted Slit proteins. Here we present evidence that the SH3-SH2 adaptor protein Dreadlocks (Dock), the p21-activated serine-threonine kinase (Pak), and the Rac1/Rac2/Mtl small GTPases can function during Robo repulsion. Loss-of-function and genetic interaction experiments suggest that limiting the function of Dock, Pak, or Rac partially disrupts Robo repulsion. In addition, Dock can directly bind to Robo's cytoplasmic domain, and the association of Dock and Robo is enhanced by stimulation with Slit. Furthermore, Slit stimulation can recruit a complex of Dock and Pak to the Robo receptor and trigger an increase in Rac1 activity. These results provide a direct physical link between the Robo receptor and an important cytoskeletal regulatory protein complex and suggest that Rac can function in both attractive and repulsive axon guidance.

  6. Determination of response of real-time SidePak AM510 monitor to secondhand smoke, other common indoor aerosols, and outdoor aerosol.

    PubMed

    Jiang, Ruo-Ting; Acevedo-Bolton, Viviana; Cheng, Kai-Chung; Klepeis, Neil E; Ott, Wayne R; Hildemann, Lynn M

    2011-06-01

    The amount of light scattered by airborne particles inside an aerosol photometer will vary not only with the mass concentration, but also with particle properties such as size, shape, and composition. This study conducted controlled experiments to compare the measurements of a real-time photometer, the SidePak AM510 monitor (SidePak), with gravimetric mass. PM sources tested were outdoor aerosols, and four indoor combustion sources: cigarettes, incense, wood chips, and toasting bread. The calibration factor for rescaling the SidePak measurements to agree with gravimetric mass was similar for the cigarette and incense sources, but different for burning wood chips and toasting bread. The calibration factors for ambient urban aerosols differed substantially from day to day, due to variations in the sources and composition of outdoor PM. A field evaluation inside a casino with active smokers yielded calibration factors consistent with those obtained in the controlled experiments with cigarette smoke.

  7. Comparison of peritoneal adhesion formation in bowel retraction by cotton towels versus the silicone lap pak device in a rabbit model.

    PubMed

    Liu, Brian G; Ruben, Dawn S; Renz, Wolfgang; Santillan, Antonio; Kubisen, Steven J; Harmon, John W

    2011-01-01

    Manipulation of cotton operating room towels within the abdominal cavity in open abdominal surgery has been associated with the formation of peritoneal adhesions. In a rabbit model, the use of standard cotton operating room towels is compared to the Lap Pak, a silicone bowel-packing device, to determine the potential for reducing the risk of adhesions. Thirty rabbits were randomly assigned to 3 groups. The rabbits underwent a sham surgery with incision only (n = 10), placement of operating room towels (n = 10), or placement of a Lap Pak (n = 10). After 14 days, the rabbits were sacrificed and the peritoneal cavity explored for adhesions. The number, tenacity, ease of dissection, and density of adhesions were recorded, and the adhesions quantitatively graded using a Modified Hopkins Adhesion scoring system. The operating room towel group had an average adhesion score of 2.5, and 8 (80%) rabbits developed adhesions. The sham group had an average adhesion score of 0.3 and one rabbit (10%) developed adhesions. The Lap Pak group had an average adhesion score of 0.2 and 1 rabbit (10%) developed adhesions. The frequency and severity of adhesions in the operating room towel group were significantly greater from that of the baseline sham group. There was no significant difference between the Lap Pak and sham groups. In this rabbit laparotomy model, the use of the Lap Pak to retract the bowels resulted in significantly fewer adhesions compared to cotton operating room towels. Lap Pak may be beneficial for bowel packing in general abdominal surgeries.

  8. Comparison of Peritoneal Adhesion Formation in Bowel Retraction by Cotton Towels Versus the Silicone Lap Pak Device in a Rabbit Model

    PubMed Central

    Liu, Brian G.; Ruben, Dawn S.; Renz, Wolfgang; Santillan, Antonio; Kubisen, Steven J.; Harmon, John W.

    2011-01-01

    Objective: Manipulation of cotton operating room towels within the abdominal cavity in open abdominal surgery has been associated with the formation of peritoneal adhesions. In a rabbit model, the use of standard cotton operating room towels is compared to the Lap Pak, a silicone bowel-packing device, to determine the potential for reducing the risk of adhesions. Methods: Thirty rabbits were randomly assigned to 3 groups. The rabbits underwent a sham surgery with incision only (n = 10), placement of operating room towels (n = 10), or placement of a Lap Pak (n = 10). After 14 days, the rabbits were sacrificed and the peritoneal cavity explored for adhesions. The number, tenacity, ease of dissection, and density of adhesions were recorded, and the adhesions quantitatively graded using a Modified Hopkins Adhesion scoring system. Results: The operating room towel group had an average adhesion score of 2.5, and 8 (80%) rabbits developed adhesions. The sham group had an average adhesion score of 0.3 and one rabbit (10%) developed adhesions. The Lap Pak group had an average adhesion score of 0.2 and 1 rabbit (10%) developed adhesions. The frequency and severity of adhesions in the operating room towel group were significantly greater from that of the baseline sham group. There was no significant difference between the Lap Pak and sham groups. Conclusions: In this rabbit laparotomy model, the use of the Lap Pak to retract the bowels resulted in significantly fewer adhesions compared to cotton operating room towels. Lap Pak may be beneficial for bowel packing in general abdominal surgeries. PMID:22096614

  9. Biochemical characterization of the Cool (Cloned-out-of-Library)/Pix (Pak-interactive exchange factor) proteins.

    PubMed

    Baird, Daniel; Feng, Qiyu; Cerione, Richard A

    2006-01-01

    The Cool (Cloned out of Library)/Pix (Pak interactive exchange factor) proteins have been implicated in a diversity of biological activities, ranging from pathways initiated by growth factors and chemoattractants to X-linked mental retardation. Initially discovered through yeast two-hybrid and biochemical analyses as binding partners for the Cdc42/Rac-target/effector, Pak (p21 activated kinase), the sequences for the Cool/Pix proteins revealed a DH (Dbl homology) domain. Because the DH domain is the limit functional unit for stimulating guanine nucleotide exchange on Rho family GTP-binding proteins, it was assumed that the Cool/Pix proteins would act as guanine nucleotide exchange factors (GEFs) for the Rho proteins. Of the three known isoforms, (p50Cool-1, p85Cool-1/beta-Pix, and 90Cool-2/alpha-Pix), only Cool-2/alpha-Pix has exhibited significant GEF activity. A number of experimental techniques have been used to characterize Cool-2, and in vitro analysis has revealed that its GEF activity is under tight control through intramolecular interactions involving several binding partners. Here we describe the biochemical methods used to study the Cool/Pix proteins and, in particular, the regulation of the GEF activity of Cool-2/alpha-Pix.

  10. Increased Circulating Endothelial Microparticles Associated with PAK4 Play a Key Role in Ventilation-Induced Lung Injury Process

    PubMed Central

    Pan, Shuming; Fei, Aihua; Jing, Lihong; Zhang, Xiangyu

    2017-01-01

    Inappropriate mechanical ventilation (MV) can result in ventilator-induced lung injury (VILI). Probing mechanisms of VILI and searching for effective methods are current areas of research focus on VILI. The present study aimed to probe into mechanisms of endothelial microparticles (EMPs) in VILI and the protective effects of Tetramethylpyrazine (TMP) against VILI. In this study, C57BL/6 and TLR4KO mouse MV models were used to explore the function of EMPs associated with p21 activated kinases-4 (PAK-4) in VILI. Both the C57BL/6 and TLR4 KO groups were subdivided into a mechanical ventilation (MV) group, a TMP + MV group, and a control group. After four hours of high tidal volume (20 ml/kg) MV, the degree of lung injury and the protective effects of TMP were assessed. VILI inhibited the cytoskeleton-regulating protein of PAK4 and was accompanied by an increased circulating EMP level. The intercellular junction protein of β-catenin was also decreased accompanied by a thickening alveolar wall, increased lung W/D values, and neutrophil infiltration. TMP alleviated VILI via decreasing circulating EMPs, stabilizing intercellular junctions, and alleviating neutrophil infiltration. PMID:28261612

  11. Molluscan associations from the Pak Phanang Bay (SW Gulf of Thailand) as a record of natural and anthropogenic changes

    NASA Astrophysics Data System (ADS)

    Negri, Mauro Pietro; Sanfilippo, Rossana; Basso, Daniela; Rosso, Antonietta; Di Geronimo, Sebastiano Italo

    2014-08-01

    Recent environmental changes in the Pak Phanang Bay (SW Gulf of Thailand) are investigated by means of mollusc assemblages. The present-day water depth within the bay slightly exceeds 2 m at low tide and the seafloor is almost entirely muddy, except for the outermost part of the embayment which is directly influenced by the longshore drift that is building the Laem Talumpuk sand spit. A multivariate analysis of the molluscan fauna recovered at 16 sampling stations within the bay delineates three thanatofacies and two biofacies. The Bay Mouth thanatofacies, including several infralittoral species, is distributed around the bay entrance; the Tidal Flat thanatofacies, characterized by few brackish and freshwater taxa, occurs in the inner part of the bay; the Channel thanatofacies includes a mixed fauna and is found along the long axis of the bay. All thanatofacies are not older than a few decades. The two biofacies are significantly less diverse than their dead counterparts, and are simply identified as Bay Mouth biofacies and Inner Bay biofacies. The faunal evolution, combined with bathymetric and sedimentological data, confirms that the embayment is undergoing a confinement process. The inner bay has evolved into an undifferentiated tidal flat hosting an oligospecific fauna. The confinement trend and the consequent siltation of the bay, mainly due to natural geomorphologic processes acting since centuries, are likely to have sped up in recent years by interaction with some human activities; among these, the deforestation in the upper Pak Phanang basin and the construction of Uthokaviphatprasit Watergate.

  12. Reversed-phase liquid chromatography of radiolabeled peptides using a C18 guard-PAK precolumn system

    SciTech Connect

    Carriere, P.D.; Bennett, H.P. )

    1989-03-01

    In order to avoid radioactive contamination of high-performance liquid chromatography columns and injectors, we have investigated the use of a Guard-PAK precolumn system for the chromatography of ({sup 125}I) labeled peptides. Two gonadotropin-releasing hormone analogs: (1) (D-Ala6-des-Gly10)-GnRH (GnRH-(Ala6)) and (2) (D-Ser(TBu)6-des-Gly10)-GnRH (GnRH-(Ser6)) and rat prolactin (r-PRL) were radiolabeled with {sup 125}I and subjected to reversed-phase liquid chromatography using a C18 Guard-PAK precolumn system. Major peak fractions of purified ({sup 125}I)GnRH-(Ala6), ({sup 125}I)GnRH-(Ser6), and ({sup 125}I)r-PRL eluted at 24%, 28%, and 55% acetonitrile, respectively. Purified ({sup 125}I)GnRH analogs showed specific high affinity binding to rat anterior pituitary gland membranes (specific activity: 1500-1700 Ci/mmol). Purified ({sup 125}I)r-PRL showed high affinity binding to r-PRL antibody by RIA (specific activity: 70-75 microCi/micrograms). This rapid and efficient chromatographic method should be useful in the separation of a wide range of radiolabeled protein and peptide molecules.

  13. Effects of low light on photosynthetic properties, antioxidant enzyme activity, and anthocyanin accumulation in purple pak-choi (Brassica campestris ssp. Chinensis Makino)

    PubMed Central

    Li, Xiaofeng; Zhai, Wen; Liu, Yang; Gao, Qianqian; Liu, Jinping; Ren, Li; Chen, Huoying; Zhu, Yuying

    2017-01-01

    Anthocyanins are secondary metabolites that contribute to red, blue, and purple colors in plants and are affected by light, but the effects of low light on the physiological responses of purple pak-choi plant leaves are still unclear. In this study, purple pak-choi seedlings were exposed to low light by shading with white gauze and black shading in a phytotron. The responses in terms of photosynthetic properties, carbohydrate metabolism, antioxidant enzyme activity, anthocyanin biosynthetic enzyme activity, and the relative chlorophyll and anthocyanin content of leaves were measured. The results showed that chlorophyll b, intracellular CO2 content, stomatal conductance and antioxidant activities of guaiacol peroxidase, catalase and superoxide dismutase transiently increased in the shade treatments at 5 d. The malondialdehyde content also increased under low light stress, which damages plant cells. With the extension of shading time (at 15 d), the relative chlorophyll a, anthocyanin and soluble protein contents, net photosynthetic rate, transpiration rate, stomata conductance, antioxidant enzyme activities, and activities of four anthocyanin biosynthetic enzymes decreased significantly. Thus, at the early stage of low light treatment, the chlorophyll b content increased to improve photosynthesis. When the low light treatment was extended, antioxidant enzyme activity and the activity of anthocyanin biosynthesis enzymes were inhibited, causing the purple pak-choi seedlings to fade from purple to green. This study provides valuable information for further deciphering genetic mechanisms and improving agronomic traits in purple pak-choi under optimal light requirements. PMID:28609452

  14. Effects of low light on photosynthetic properties, antioxidant enzyme activity, and anthocyanin accumulation in purple pak-choi (Brassica campestris ssp. Chinensis Makino).

    PubMed

    Zhu, Hongfang; Li, Xiaofeng; Zhai, Wen; Liu, Yang; Gao, Qianqian; Liu, Jinping; Ren, Li; Chen, Huoying; Zhu, Yuying

    2017-01-01

    Anthocyanins are secondary metabolites that contribute to red, blue, and purple colors in plants and are affected by light, but the effects of low light on the physiological responses of purple pak-choi plant leaves are still unclear. In this study, purple pak-choi seedlings were exposed to low light by shading with white gauze and black shading in a phytotron. The responses in terms of photosynthetic properties, carbohydrate metabolism, antioxidant enzyme activity, anthocyanin biosynthetic enzyme activity, and the relative chlorophyll and anthocyanin content of leaves were measured. The results showed that chlorophyll b, intracellular CO2 content, stomatal conductance and antioxidant activities of guaiacol peroxidase, catalase and superoxide dismutase transiently increased in the shade treatments at 5 d. The malondialdehyde content also increased under low light stress, which damages plant cells. With the extension of shading time (at 15 d), the relative chlorophyll a, anthocyanin and soluble protein contents, net photosynthetic rate, transpiration rate, stomata conductance, antioxidant enzyme activities, and activities of four anthocyanin biosynthetic enzymes decreased significantly. Thus, at the early stage of low light treatment, the chlorophyll b content increased to improve photosynthesis. When the low light treatment was extended, antioxidant enzyme activity and the activity of anthocyanin biosynthesis enzymes were inhibited, causing the purple pak-choi seedlings to fade from purple to green. This study provides valuable information for further deciphering genetic mechanisms and improving agronomic traits in purple pak-choi under optimal light requirements.

  15. Glucosinolate Accumulation and Related Gene Expression in Pak Choi (Brassica rapa L. ssp. chinensis var. communis [N. Tsen & S.H. Lee] Hanelt) in Response to Insecticide Application.

    PubMed

    Zhu, Biao; Yang, Jing; He, Yong; Zang, Yunxiang; Zhu, Zhujun

    2015-11-11

    Glucosinolates and their breakdown products are well-known for their cancer-chemoprotective functions and biocidal activities against pathogens and generalist herbivores. Insecticides are commonly used in the production of pak choi (Brassica rapa L. ssp. chinensis var. communis [N. Tsen & S.H. Lee] Hanelt). We studied the effects of four commonly used insecticides, namely, β-cypermethrin, acephate, pymetrozine, and imidacloprid, on glucosinolate metabolism in pak choi. All insecticides significantly increased both the transcription of glucosinolate biosynthetic genes and the aliphatic and total glucosinolate accumulations in pak choi. β-Cypermethrin and acephate caused gradual and continuous up-regulation of gene expression from 0.5 to 24 h after treatment, whereas pymetrozine and imidacloprid did so more rapidly, reaching a peak at 1 h and returning to normal at 3 h. Our findings indicate that the four insecticides affect glucosinolate metabolism in pak choi plants to various degrees and suggest that glucosinolates may be involved in plant insecticide metabolism.

  16. Test Plan for Lockheed Idaho Technologies Company (LITCO), ARROW-PAK Packaging, Docket 95-40-7A, Type A Container

    SciTech Connect

    Kelly, D.L.

    1995-10-23

    This report documents the U.S. Department of Transportation Specification 7A Type A (DOT-7A) compliance testing to be followed for qualification of the Lockheed Idaho Technologies Company, ARROW-PAK, for use as a Type A Packaging. The packaging configuration being tested is intended for transportation of radioactive solids, Form No. 1, Form No. 2, and Form No. 3.

  17. Regulation of insulin signaling by the phosphatidylinositol 3,4,5-triphosphate phosphatase SKIP through the scaffolding function of Pak1.

    PubMed

    Ijuin, Takeshi; Takenawa, Tadaomi

    2012-09-01

    Skeletal muscle and kidney-enriched inositol polyphosphate phosphatase (SKIP) has previously been implicated in the regulation of insulin signaling in skeletal muscle. Here, we present the first report of the mechanisms by which SKIP specifically suppresses insulin signaling and the subsequent glucose uptake. Upon insulin stimulation, SKIP is translocated to the membrane ruffles, where it binds to the active form of Pak1, which mediates multiple protein complex formation with phosphatidylinositol 3,4,5-triphosphate (PIP(3)) effectors such as Akt2, PDK1, and Rac1; this leads to inactivation of these proteins. SKIP also promotes the inhibition of Rac1-dependent kinase activity and the scaffolding function of Pak1, which results in the dissociation of Akt2 and PDK1 from Pak1. Thus, specific suppression of insulin signaling is achieved via the spatiotemporal regulation of SKIP through the scaffolding function of Pak1. These interactions are the foundation of the specific and prominent role of SKIP in the regulation of insulin signaling.

  18. ETV TEST REPORT OF CONTROL OF BIOAEROSOLS IN HVAC SYSTEMS GLASFLOSS INDUSTRIES Z-PAK SERIES S, MODEL ZPS24241295BO

    EPA Science Inventory

    The Environmental Technology Verification report discusses the technology and performance of the Z-Pak Series S, Model ZPS24241295B0 air filter for dust and bioaerosol filtration manufactured by Glasfloss Industries, Inc. The pressure drop across the filter was 91 Pa clean and 34...

  19. Hexavalent chromium stress enhances the uptake of nitrate but reduces the uptake of ammonium and glycine in pak choi (Brassica chinensis L.).

    PubMed

    Ma, Qingxu; Cao, Xiaochuang; Ma, Jinzhao; Tan, Xiaoli; Xie, Yinan; Xiao, Han; Wu, Lianghuan

    2017-05-01

    Chromium (Cr) pollution affects plant growth and biochemical processes, so, the relative uptake of glycine, nitrate, and ammonium by pak choi (Brassica chinensis) seedlings in treatments with 0mgL(-1) and 10mgL(-1) Cr (VI) were detected by substrate-specific (15)N-labelling in a sterile environment. The short-term uptake of (15)N-labelled sources and (15)N-enriched amino acids were detected by gas chromatography mass spectrometry to explore the mechanism by which Cr stress affects glycine uptake and metabolism, which showing that Cr stress hindered the uptake of ammonium and glycine but increased significantly the uptake of nitrate. Cr stress did not decrease the active or passive uptake of glycine, but it inhibited the conversion of glycine to serine in pak choi roots, indicating that the metabolism of glycine to serine in roots, rather than the root uptake, was the limiting step in glycine contribution to total N uptake in pak choi. Since Cr affects the relative uptake of different N sources, a feasible way to reduce Cr-induced stress is application of selective fertilization, in particular nitrate, in pak choi cultivation on Cr-polluted soil. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. ETV TEST REPORT OF CONTROL OF BIOAEROSOLS IN HVAC SYSTEMS GLASFLOSS INDUSTRIES Z-PAK SERIES S, MODEL ZPS24241295BO

    EPA Science Inventory

    The Environmental Technology Verification report discusses the technology and performance of the Z-Pak Series S, Model ZPS24241295B0 air filter for dust and bioaerosol filtration manufactured by Glasfloss Industries, Inc. The pressure drop across the filter was 91 Pa clean and 34...

  1. 1,2,3-Triazolyl ester of Ketorolac: A "Click Chemistry"-based highly potent PAK1-blocking cancer-killer.

    PubMed

    Nguyen, Binh Cao Quan; Takahashi, Hideaki; Uto, Yoshihiro; Shahinozzaman, M D; Tawata, Shinkichi; Maruta, Hiroshi

    2017-01-27

    An old anti-inflammatory/analgesic drug called Toradol is a racemic form of Ketorolac (50% R-form and 50% S-form) that blocks the oncogenic RAC-PAK1-COX-2 (cyclooxygenase-2) signaling, through the direct inhibition of RAC by the R-form and of COX-2 by the S-form, eventually down-regulating the production of prostaglandins. However, due to its COOH moiety which is clearly repulsive to negatively-charged phospholipid-based plasma membrane, its cell-permeability is rather poor (the IC50 against the growth of human cancer cells such as A549 is around 13 μM). In an attempt to boost its anti-cancer activity, hopefully by increasing its cell-permeability through abolishing the negative charge, yet keeping its water-solubility, here we synthesized a 1,2,3-triazolyl ester of Toradol through "Click Chemistry". The resultant water-soluble "azo" derivative called "15K" was found to be over 500 times more potent than Toradol with the IC50 around 24 nM against the PAK1-dependent growth of A549 cancer cells, inactivating PAK1 in cell culture with the apparent IC50 around 65 nM, and inhibiting COX-2 in vitro with the IC50 around 6 nM. Furthermore, the Click Chemistry boosts the anti-cancer activity of Ketorolac by 5000 times against the PAK1-independent growth of B16F10 melanoma cells. Using a multi-drug-resistant (MDR) cancer cell line (EMT6), we found that the esterization of Ketorolac boosts its cell-permeability by at least 10 folds. Thus, the Click Chemistry dramatically boosts the anti-cancer activity of Ketorolac, at least in three ways: increasing its cell-permeability, the anti-PAK1 activity of R-form and anti-COX-2 activity of S-form. The resultant "15K" is so far among the most potent PAK1-blockers, and therefore would be potentially useful for the therapy of many different PAK1-dependent diseases/disorders such as cancers.

  2. Performance of para-Pak Ultra ECOFIX compared with Para-Pak Ultra formalin/mercuric chloride-based polyvinyl alcohol for concentration and permanent stained smears of stool parasites.

    PubMed

    Fedorko, D P; Williams, E C; Nelson, N A; Mazyck, T D; Hanson, K L; Cartwright, C P

    2000-05-01

    ECOFIX is a mercury and formalin-free fecal preservative that can be used for concentration of stool specimens and preparation of permanently-stained slides. In this study, the standard two-vial ParaPak Ultra system was compared with ECOFIX Ultra for the detection of intestinal parasites. A total of 261 specimens in 92 sets (77 with 3 specimens, 15 with 2 specimens) were collected in ECOFIX, formalin, and low viscosity polyvinyl alcohol (LV-PVA). Concentrations were performed from ECOFIX using Hemo-De and saline and from formalin using ethyl acetate and formalin. To prepare permanently-stained smears, ECOSTAIN (a modification of Wheatley's trichrome stain) was used on ECOFIX material and Wheatley's trichrome stain was used on specimens preserved in PVA. A total of 157 protozoa and helminths were detected; 132 (84.1%) were recovered in formalin/PVA and 129 (82.2%) in ECOFIX. In permanently-stained smears, 139 protozoa were observed, 116 (83.5%) in PVA-preserved material and 117 (84.2%) in ECOFIX. Fecal concentration yielded 111 parasites (103 protozoa and 8 helminths), of which 98 (88.3%) were detected in formalin-fixed stool and 48 (43.2%) in ECOFIX. Significantly fewer ECOFIX-preserved concentrates were positive for Blastocystis hominis (35 versus 15, p-value <0.001) and Endolimax nana (19 versus 2, p-value <0.001). In conclusion, use of the ECOFIX Ultra collection device in combination with ECOSTAIN resulted in largely comparable recovery of enteric parasites to the conventional two-vial ParaPak Ultra system when both sedimentation-concentration and permanently stained smears were performed, and 2-3 specimens per patient were evaluated.

  3. Hair Growth Promoting and Anticancer Effects of p21-activated kinase 1 (PAK1) Inhibitors Isolated from Different Parts of Alpinia zerumbet.

    PubMed

    Taira, Nozomi; Nguyen, Binh Cao Quan; Tawata, Shinkichi

    2017-01-14

    PAK1 (p21-activated kinase 1) is an emerging target for the treatment of hair loss (alopecia) and cancer; therefore, the search for PAK1 blockers to treat these PAK1-dependent disorders has received much attention. In this study, we evaluated the anti-alopecia and anticancer effects of PAK1 inhibitors isolated from Alpinia zerumbet (alpinia) in cell culture. The bioactive compounds isolated from alpinia were found to markedly promote hair cell growth. Kaempferol-3-O-β-d-glucuronide (KOG) and labdadiene, two of the isolated compounds, increased the proliferation of human follicle dermal papilla cells by approximately 117%-180% and 132%-226%, respectively, at 10-100 μM. MTD (2,5-bis(1E,3E,5E)-6-methoxyhexa-1,3,5-trien-1-yl)-2,5-dihydrofuran) and TMOQ ((E)-2,2,3,3-tetramethyl-8-methylene-7-(oct-6-en-1-yl)octahydro-1H-quinolizine) showed growth-promoting activity around 164% and 139% at 10 μM, respectively. The hair cell proliferation induced by these compounds was significantly higher than that of minoxidil, a commercially available treatment for hair loss. Furthermore, the isolated compounds from alpinia exhibited anticancer activity against A549 lung cancer cells with IC50 in the range of 67-99 μM. Regarding the mechanism underlying their action, we hypothesized that the anti-alopecia and anticancer activities of these compounds could be attributed to the inhibition of the oncogenic/aging kinase PAK1.

  4. p21-Activated Kinase 1 (Pak1) Phosphorylates BAD Directly at Serine 111 In Vitro and Indirectly through Raf-1 at Serine 112

    PubMed Central

    Zhuo, Ya; Field, Jeffrey

    2011-01-01

    Background Cell survival depends on the balance between protective and apoptotic signals. When the balance of signals tips towards apoptosis, cells undergo programmed cell death. This balance has profound implications in diseases including cancer. Oncogenes and tumor suppressors are mutated to promote cell survival during tumor development, and many chemotherapeutic drugs kill tumor cells by stimulating apoptosis. BAD is a pro-apoptotic member of the Bcl-2 family of proteins, which can be phosphorylated on numerous sites to modulate binding to Bcl-2 and 14-3-3 proteins and inhibit its pro-apoptotic activities. One of the critical phosphorylation sites is the serine 112 (S112), which can be phosphorylated by several kinases including Pak1. Methodology/Principal Findings We mapped the Pak phosphorylation sites by making serine to alanine mutations in BAD and testing them as substrates in in vitro kinase assays. We found that the primary phosphorylation site is not S112 but serine 111 (S111), a site that is sometimes found phosphorylated in vivo. In transfection assays of HEK293T cells, we showed that Pak1 required Raf-1 to stimulate phosphorylation on S112. Mutating either S111 or S112 to alanine enhanced binding to Bcl-2, but the double mutant S111/112A bound better to Bcl-2. Moreover, BAD phosphorylation at S111 was observed in several other cell lines, and treating one of them with the Pak1 inhibitor 2,2′-Dihydroxy-1,1′-dinaphthyldisulfide (IPA-3) reduced phosphorylation primarily at S112 and to a smaller extent at S111, while Raf inhibitors only reduced phosphorylation at S112. Conclusion/Significance Together, these findings demonstrate that Pak1 phosphorylates BAD directly at S111, but phosphorylated S112 through Raf-1. These two sites of BAD serve as redundant regulatory sites for Bcl-2 binding. PMID:22096607

  5. A Population Growth Trend Analysis for Neotricula aperta, the Snail Intermediate Host of Schistosoma mekongi, after Construction of the Pak-Mun Dam

    PubMed Central

    Attwood, Stephen W.; Upatham, E. Suchart

    2013-01-01

    Background The Pak-Mun dam is a controversial hydro-power project on the Mun River in Northeast Thailand. The dam is sited in a habitat of the freshwater snail Neotricula aperta, which is the intermediate host for the parasitic blood-fluke Schistosoma mekongi causing Mekong schistosomiasis in humans in Cambodia and Laos. Few data are available which can be used to assess the effects of water resource development on N. aperta. The aim of this study was to obtain data and to analyze the possible impact of the dam on N. aperta population growth. Methodology/Principal Findings Estimated population densities were recorded for an N. aperta population in the Mun River 27 km upstream of Pak-Mun, from 1990 to 2011. The Pak-Mul dam began to operate in 1994. Population growth was modeled using a linear mixed model expression of a modified Gompertz stochastic state-space exponential growth model. The N. aperta population was found to be quite stable, with the estimated growth parameter not significantly different from zero. Nevertheless, some marked changes in snail population density were observed which were coincident with changes in dam operation policy. Conclusions/Significance The study found that there has been no marked increase in N. aperta population growth following operation of the Pak-Mun dam. The analysis did indicate a large and statistically significant increase in population density immediately after the dam came into operation; however, this increase was not persistent. The study has provided the first vital baseline data on N. aperta population behavior near to the Pak-Mun dam and suggests that the operation policy of the dam may have an impact on snail population density. Nevertheless, additional studies are required for other N. aperta populations in the Mun River and for an extended time series, to confirm or refine the findings of this work. PMID:24244775

  6. Near real time observational data collection for SPRUCE experiment- PakBus protocol for slow satellite connections

    NASA Astrophysics Data System (ADS)

    Krassovski, Misha; Hanson, Paul; Riggs, Jeff

    2017-04-01

    Climate change studies are one of the most important aspects of modern science and related experiments are getting bigger and more complex. One such experiment is the Spruce and Peatland Responses Under Climatic and Environmental Change experiment (SPRUCE, http://mnspruce.ornl.gov) conducted in in northern Minnesota, 40 km north of Grand Rapids, in the USDA Forest Service Marcell Experimental Forest (MEF). The SPRUCE experimental mission is to assess ecosystem-level biological responses of vulnerable, high carbon terrestrial ecosystems to a range of climate warming manipulations and an elevated CO2 atmosphere. This manipulation experiment generates a lot of observational data and requires a reliable onsite data collection system, dependable methods to transfer data to a robust scientific facility, and real-time monitoring capabilities. This publication shares our experience of establishing near real time data collection and monitoring system via a satellite link using PakBus protocol.

  7. DNA methylation of a novel PAK4 locus influences ototoxicity susceptibility following cisplatin and radiation therapy for pediatric embryonal tumors.

    PubMed

    Brown, Austin L; Foster, Kayla L; Lupo, Philip J; Peckham-Gregory, Erin C; Murray, Jeffrey C; Okcu, M Fatih; Lau, Ching C; Rednam, Surya P; Chintagumpala, Murali; Scheurer, Michael E

    2017-10-01

    Ototoxicity is a common adverse side effect of platinum chemotherapy and cranial radiation therapy; however, individual susceptibility is highly variable. Therefore, our objective was to conduct an epigenome-wide association study to identify differentially methylated cytosine-phosphate-guanine (CpG) sites associated with ototoxicity susceptibility among cisplatin-treated pediatric patients with embryonal tumors. Samples were collected for a discovery cohort (n = 62) and a replication cohort (n = 18) of medulloblastoma and primitive neuroectodermal tumor patients. Posttreatment audiograms were evaluated using the International Society of Paediatric Oncology (SIOP) Boston Ototoxicity Scale. Genome-wide associations between CpG methylation and ototoxicity were examined using multiple linear regression, controlling for demographic and treatment factors. The mean cumulative dose of cisplatin was 330 mg/m2 and the mean time from end of therapy to the last available audiogram was 6.9 years. In the discovery analysis of 435233 CpG sites, 6 sites were associated with ototoxicity grade (P < 5 × 10-5) after adjusting for confounders. Differential methylation at the top CpG site identified in the discovery cohort (cg14010619, PAK4 gene) was replicated (P = 0.029) and reached genome-wide significance (P = 2.73 × 10-8) in a combined analysis. These findings were robust to a sensitivity analysis evaluating other potential confounders. We identified and replicated a novel CpG methylation loci (cg14010619) associated with ototoxicity severity. Methylation at cg14010619 may modify PAK4 activity, which has been implicated in cisplatin resistance in malignant cell lines.

  8. Final evaluation report for Lockheed Idaho Technologies Company, ARROW-PAK packaging, Docket 95-40-7A, Type A container

    SciTech Connect

    Kelly, D.L.

    1995-11-01

    The report documents the U.S. Department of Transportation Specification 7A Type A (DOT-7A) compliance test results of the ARROW-PAK packaging. The ARROW-PAK packaging system consists of Marlex M-8000 Driscopipe (Series 8000 [gas] or Series 8600 [industrial]) resin pipe, manufactured by Phillips-Driscopipe, Inc., and is sealed with two dome-shaped end caps manufactured from the same materials. The patented sealing process involves the use of electrical energy to heat opposing faces of the pipe and end caps, and hydraulic rams to press the heated surfaces together. This fusion process produces a homogeneous bonding of the end cap to the pipe. The packaging may be used with or without the two internal plywood spacers. This packaging was evaluated and tested in October 1995. The packaging configuration described in this report is designed to ship Type A quantities of solid radioactive materials, Form No. 1, Form No. 2, and Form No. 3.

  9. Hispidin and related herbal compounds from Alpinia zerumbet inhibit both PAK1-dependent melanogenesis in melanocytes and reactive oxygen species (ROS) production in adipocytes.

    PubMed

    Be Tu, Pham Thi; Chompoo, Jamnian; Tawata, Shinkichi

    2015-06-01

    Recently several compounds from Okinawa plants including Alpinia zerumbet (alpinia) were shown to inhibit directly the oncogenic/ageing kinase PAK1 (p21-activated kinase 1). Furthermore, it was recently revealed that both PAK1 and PAK4 (p21-activated kinase 4) are equally essential for the melanogenesis in melanoma cells. Thus, in this study, we tested if several alpinia compounds inhibit the melanogenesis in melanoma (B16F10) cells, as well as the PAK1-dependent up-regulation of both reactive oxygen species (ROS) and nitric oxide (NO) in cultured adipocytes (3T3-L1) without any cytotoxicity. The effect of alpinia compounds on the melanogenesis was measured by both the melanin content and intracellular tyrosinase activity in melanoma cells treated with 3-isobutyl-1-methylxanthine (IBMX), a melanogenesis stimulating hormone. We found that (1E,3E,5E)-6-methoxyhexa-1,3,5-trien-1-yl)-2,5-dihydrofuran (MTD), 5,6-dehydrokawain (DK), labdadiene, hispidin and dihydro-5,6-dehydrokawain (DDK) at 50 μg/mL reduced the melanin content by 63-79%. The MTD, DK and hispidin, at 50 μg/mL, inhibited tyrosinase activity by 70-83% in melanoma cells. Among these compounds, labdadiene, MTD, (E)-2,2,3,3-Tetramethyl8-methylene-7-(oct-6-en-1-yl)octahydro-1H-quinolizine (TMOQ) and hispidin strongly inhibited the ROS production. Hispidin, labdadiene and MTD at 20 μg/mL inhibited NO production by over 70%. These findings altogether suggest that some of these alpinia compounds could be potentially useful for the prevention or treatment of hyperpigmentation and obesity.

  10. Molecular evolution, characterization, and expression analysis of SnRK2 gene family in Pak-choi (Brassica rapa ssp. chinensis)

    PubMed Central

    Huang, Zhinan; Tang, Jun; Duan, Weike; Wang, Zhen; Song, Xiaoming; Hou, Xilin

    2015-01-01

    The sucrose non-fermenting 1-related protein kinase 2 (SnRK2) family members are plant-specific serine/threonine kinases that are involved in the plant response to abiotic stress and abscisic acid (ABA)-dependent plant development. Further understanding of the evolutionary history and expression characteristics of these genes will help to elucidate the mechanisms of the stress tolerance in Pak-choi, an important green leafy vegetable in China. Thus, we investigated the evolutionary patterns, footprints and conservation of SnRK2 genes in selected plants and later cloned and analyzed SnRK2 genes in Pak-choi. We found that this gene family was preferentially retained in Brassicas after the Brassica-Arabidopsis thaliana split. Next, we cloned and sequenced 13 SnRK2 from both cDNA and DNA libraries of stress-induced Pak-choi, which were under conditions of ABA, salinity, cold, heat, and osmotic treatments. Most of the BcSnRK2s have eight exons and could be divided into three groups. The subcellular localization predictions suggested that the putative BcSnRK2 proteins were enriched in the nucleus. The results of an analysis of the expression patterns of the BcSnRK2 genes showed that BcSnRK2 group III genes were robustly induced by ABA treatments. Most of the BcSnRK2 genes were activated by low temperature, and the BcSnRK2.6 genes responded to both ABA and low temperature. In fact, most of the BcSnRK2 genes showed positive or negative regulation under ABA and low temperature treatments, suggesting that they may be global regulators that function at the intersection of multiple signaling pathways to play important roles in Pak-choi stress responses. PMID:26557127

  11. Signaling of the p21-activated kinase (PAK1) coordinates insulin-stimulated actin remodeling and glucose uptake in skeletal muscle cells

    PubMed Central

    Tunduguru, Ragadeepthi; Chiu, Tim T.; Ramalingam, Latha; Elmendorf, Jeffrey S.; Klip, Amira; Thurmond, Debbie C.

    2015-01-01

    Skeletal muscle accounts for ~80% of postprandial glucose clearance, and skeletal muscle glucose clearance is crucial for maintaining insulin sensitivity and euglycemia. Insulin-stimulated glucose clearance/uptake entails recruitment of glucose transporter 4 (GLUT4) to the plasma membrane (PM) in a process that requires cortical F-actin remodeling; this process is dysregulated in Type 2 Diabetes. Recent studies have implicated PAK1 as a required element in GLUT4 recruitment in mouse skeletal muscle in vivo, although its underlying mechanism of action and requirement in glucose uptake remains undetermined. Toward this, we have employed the PAK1 inhibitor, IPA3, in studies using L6-GLUT4-myc muscle cells. IPA3 fully ablated insulin-stimulated GLUT4 translocation to the PM, corroborating the observation of ablated insulin-stimulated GLUT4 accumulation in the PM of skeletal muscle from PAK1−/− knockout mice. IPA3-treatment also abolished insulin-stimulated glucose uptake into skeletal myotubes. Mechanistically, live-cell imaging of myoblasts expressing the F-actin biosensor LifeAct-GFP treated with IPA3 showed blunting of the normal insulin-induced cortical actin remodeling. This blunting was underpinned by a loss of normal insulin-stimulated cofilin dephosphorylation in IPA3-treated myoblasts. These findings expand upon the existing model of actin remodeling in glucose uptake, by placing insulin-stimulated PAK1 signaling as a required upstream step to facilitate actin remodeling and subsequent cofilin dephosphorylation. Active, dephosphorylated cofilin then provides the G-actin substrate for continued F-actin remodeling to facilitate GLUT4 vesicle translocation for glucose uptake into the skeletal muscle cell. PMID:25199455

  12. Signaling of the p21-activated kinase (PAK1) coordinates insulin-stimulated actin remodeling and glucose uptake in skeletal muscle cells.

    PubMed

    Tunduguru, Ragadeepthi; Chiu, Tim T; Ramalingam, Latha; Elmendorf, Jeffrey S; Klip, Amira; Thurmond, Debbie C

    2014-11-15

    Skeletal muscle accounts for ∼ 80% of postprandial glucose clearance, and skeletal muscle glucose clearance is crucial for maintaining insulin sensitivity and euglycemia. Insulin-stimulated glucose clearance/uptake entails recruitment of glucose transporter 4 (GLUT4) to the plasma membrane (PM) in a process that requires cortical F-actin remodeling; this process is dysregulated in Type 2 Diabetes. Recent studies have implicated PAK1 as a required element in GLUT4 recruitment in mouse skeletal muscle in vivo, although its underlying mechanism of action and requirement in glucose uptake remains undetermined. Toward this, we have employed the PAK1 inhibitor, IPA3, in studies using L6-GLUT4-myc muscle cells. IPA3 fully ablated insulin-stimulated GLUT4 translocation to the PM, corroborating the observation of ablated insulin-stimulated GLUT4 accumulation in the PM of skeletal muscle from PAK1(-/-) knockout mice. IPA3-treatment also abolished insulin-stimulated glucose uptake into skeletal myotubes. Mechanistically, live-cell imaging of myoblasts expressing the F-actin biosensor LifeAct-GFP treated with IPA3 showed blunting of the normal insulin-induced cortical actin remodeling. This blunting was underpinned by a loss of normal insulin-stimulated cofilin dephosphorylation in IPA3-treated myoblasts. These findings expand upon the existing model of actin remodeling in glucose uptake, by placing insulin-stimulated PAK1 signaling as a required upstream step to facilitate actin remodeling and subsequent cofilin dephosphorylation. Active, dephosphorylated cofilin then provides the G-actin substrate for continued F-actin remodeling to facilitate GLUT4 vesicle translocation for glucose uptake into the skeletal muscle cell. Copyright © 2014 Elsevier Inc. All rights reserved.

  13. FTY720 prevents ischemia/reperfusion injury-associated arrhythmias in an ex vivo rat heart model via activation of Pak1/Akt signaling.

    PubMed

    Egom, E Eroume A; Ke, Yunbo; Musa, Hanny; Mohamed, Tamer M A; Wang, Tao; Cartwright, Elizabeth; Solaro, R John; Lei, Ming

    2010-02-01

    Recent studies demonstrated a role of sphingosine-1-phosphate (S1P) in the protection against the stress of ischemia/reperfusion (I/R) injury. In experiments reported here, we have investigated the signaling through the S1P cascade by FTY720, a sphingolipid drug candidate displaying structural similarity to S1P, underlying the S1P cardioprotective effect. In ex vivo rat heart and isolated sinoatrial node models, FTY720 significantly prevented arrhythmic events associated with I/R injury including premature ventricular beats, VT, and sinus bradycardia as well as A-V conduction block. Real-time PCR and Western blot analysis demonstrated the expression of the S1P receptor transcript pools and corresponding proteins including S1P1, S1P2, and S1P3 in tissues dissected from sinoatrial node, atrium and ventricle. FTY720 (25 nM) significantly blunted the depression of the levels of phospho-Pak1 and phospho-Akt with ischemia and with reperfusion. There was a significant increase in phospho-Pak1 levels by 35%, 199%, and 205% after 5, 10, and 15 min of treatment with 25 nM FTY720 compared with control nontreated myocytes. However, there was no significant difference in the levels of total Pak1 expression between nontreated and FTY720 treated. Phospho-Akt levels were increased by 44%, 63%, and 61% after 5, 10, and 15 min of treatment with 25 nM FTY720, respectively. Our data provide the first evidence that FTY720 prevents I/R injury-associated arrhythmias and indicate its potential significance as an important and new agent protecting against I/R injury. Our data also indicate, for the first time, that the cardioprotective effect of FTY720 is likely to involve activation of signaling through the Pak1.

  14. Phytoavailability of Cadmium (Cd) to Pak Choi (Brassica chinensis L.) Grown in Chinese Soils: A Model to Evaluate the Impact of Soil Cd Pollution on Potential Dietary Toxicity

    PubMed Central

    Yang, Xiaoe; Xiao, Wendan; Stoffella, Peter J.; Saghir, Aamir; Azam, Muhammad; Li, Tingqiang

    2014-01-01

    Food chain contamination by soil cadmium (Cd) through vegetable consumption poses a threat to human health. Therefore, an understanding is needed on the relationship between the phytoavailability of Cd in soils and its uptake in edible tissues of vegetables. The purpose of this study was to establish soil Cd thresholds of representative Chinese soils based on dietary toxicity to humans and develop a model to evaluate the phytoavailability of Cd to Pak choi (Brassica chinensis L.) based on soil properties. Mehlich-3 extractable Cd thresholds were more suitable for Stagnic Anthrosols, Calcareous, Ustic Cambosols, Typic Haplustalfs, Udic Ferrisols and Periudic Argosols with values of 0.30, 0.25, 0.18, 0.16, 0.15 and 0.03 mg kg−1, respectively, while total Cd is adequate threshold for Mollisols with a value of 0.86 mg kg−1. A stepwise regression model indicated that Cd phytoavailability to Pak choi was significantly influenced by soil pH, organic matter, total Zinc and Cd concentrations in soil. Therefore, since Cd accumulation in Pak choi varied with soil characteristics, they should be considered while assessing the environmental quality of soils to ensure the hygienically safe food production. PMID:25386790

  15. Cloning and partial characterization of regulated promoters from Lactococcus lactis Tn917-lacZ integrants with the new promoter probe vector, pAK80.

    PubMed Central

    Israelsen, H; Madsen, S M; Vrang, A; Hansen, E B; Johansen, E

    1995-01-01

    Transposon Tn917-LTV1 was used to produce a collection of Lactococcus lactis strains with fusion of a promoterless lacZ gene to chromosomal loci. Screening 2,500 Tn917-LTV1 integrants revealed 222 that express beta-galactosidase on plates at 30 degrees C. Pulsed-field gel electrophoresis revealed Tn917-LTV1 insertions in at least 13 loci in 15 strains analyzed. Integrants in which beta-galactosidase expression was regulated by temperature or pH and/or arginine concentration were isolated. In most cases, the regulation observed on plates was reproducible in liquid medium. One integrant, PA170, produces beta-galactosidase at pH 5.2 but not at pH 7.0, produces more beta-galactosidase at 15 degrees C than at 30 degrees C, and has increased beta-galactosidase activity in the stationary phase. DNA fragments potentially carrying promoters from selected Lactococcus lactis integrants were cloned in Escherichia coli. A new promoter probe vector, pAK80, containing promoterless beta-galactosidase genes from Leuconostoc mesenteroides subsp. cremoris and the Lactococcus lactis subsp. lactis biovar diacetylactis citrate plasmid replication region was constructed, and the lactococcal fragments were inserted. Plasmid pAK80 was capable of detecting and discriminating even weak promoters in Lactococcus lactis. When inserted in pAK80, the promoter cloned from PA170 displayed a regulated expression of beta-galactosidase analogous to the regulation observed in PA170. PMID:7618865

  16. Phytoavailability of cadmium (Cd) to Pak choi (Brassica chinensis L.) grown in Chinese soils: a model to evaluate the impact of soil Cd pollution on potential dietary toxicity.

    PubMed

    Rafiq, Muhammad Tariq; Aziz, Rukhsanda; Yang, Xiaoe; Xiao, Wendan; Stoffella, Peter J; Saghir, Aamir; Azam, Muhammad; Li, Tingqiang

    2014-01-01

    Food chain contamination by soil cadmium (Cd) through vegetable consumption poses a threat to human health. Therefore, an understanding is needed on the relationship between the phytoavailability of Cd in soils and its uptake in edible tissues of vegetables. The purpose of this study was to establish soil Cd thresholds of representative Chinese soils based on dietary toxicity to humans and develop a model to evaluate the phytoavailability of Cd to Pak choi (Brassica chinensis L.) based on soil properties. Mehlich-3 extractable Cd thresholds were more suitable for Stagnic Anthrosols, Calcareous, Ustic Cambosols, Typic Haplustalfs, Udic Ferrisols and Periudic Argosols with values of 0.30, 0.25, 0.18, 0.16, 0.15 and 0.03 mg kg-1, respectively, while total Cd is adequate threshold for Mollisols with a value of 0.86 mg kg-1. A stepwise regression model indicated that Cd phytoavailability to Pak choi was significantly influenced by soil pH, organic matter, total Zinc and Cd concentrations in soil. Therefore, since Cd accumulation in Pak choi varied with soil characteristics, they should be considered while assessing the environmental quality of soils to ensure the hygienically safe food production.

  17. Separation properties of aluminium-plastic laminates in post-consumer Tetra Pak with mixed organic solvent.

    PubMed

    Zhang, S F; Zhang, L L; Luo, K; Sun, Z X; Mei, X X

    2014-04-01

    The separation properties of the aluminium-plastic laminates in postconsumer Tetra Pak structure were studied in this present work. The organic solvent blend of benzene-ethyl alcohol-water was used as the separation reagent. Then triangle coordinate figure analysis was taken to optimize the volume proportion of various components in the separating agent and separation process. And the separation temperature of aluminium-plastic laminates was determined by the separation time, efficiency, and total mass loss of products. The results show that cost-efficient separations perform best with low usage of solvents at certain temperatures, for certain times, and within a certain range of volume proportions of the three components in the solvent agent. It is also found that similar solubility parameters of solvents and polyethylene adhesives (range 26.06-34.85) are a key factor for the separation of the aluminium-plastic laminates. Such multisolvent processes based on the combined-system concept will be vital to applications in the recycling industry.

  18. Experimental investigation and modelling of tritium washout by precipitation in the area of the nuclear power plant of Paks, Hungary.

    PubMed

    Köllo, Z; Palcsu, L; Major, Z; Papp, L; Molnár, M; Ranga, T; Dombóvári, P; Manga, L

    2011-01-01

    Tritium occurs in nature in trace amounts, but its concentration is changing due to natural and artificial sources. Studies focusing on natural tritium have to take into account the effect of artificial sources. Also, the impact of tritium is an important issue in environmental protection, e.g. in connection with the emissions from nuclear power plants. The present work focuses on the rain washout of tritium emitted from the Paks nuclear power plant in Hungary. Rainwater collectors were placed around the plant and after a period of precipitation, rainwater was collected and analysed for tritium content. Samples were analysed using low-level liquid scintillation counting, with some also subject to the more accurate (3)He ingrowth method. The results clearly show the trace of the tritium plume emitted from the plant; however, values are only about one order of magnitude higher than environmental background levels. A washout model was devised to estimate the distribution of tritium around the plant. The model gives slightly higher concentrations than those measured in the field, but in general the agreement is satisfactory. The modelled values demonstrate that the effect of the plant on rainwater tritium levels is negligible over a distance of some kilometres.

  19. An approach to Moho discontinuity recovery from on-orbit GOCE data with application over Indo-Pak region

    NASA Astrophysics Data System (ADS)

    Eshagh, Mehdi; Hussain, Matloob

    2016-10-01

    In this research, a modified form of Vening Meinesz-Moritz (VMM) theory of isostasy for the second-order radial derivative of gravitational potential, measured from the Gravity field and steady-state Ocean Circulation Explorer (GOCE), is developed for local Moho depth recovery. An integral equation is organised for inverting the GOCE data to compute a Moho model in combination with topographic/bathymetric heights of SRTM30, sediment and consolidated crystalline basement and the laterally-varying density contrast model of CRUST1.0. A Moho model from EGM2008 to degree and order 180 is also computed based on the same principle for the purpose of comparison. In addition, we compare both of them with the 3 available seismic Moho models; two global and one regional over the Indo-Pak region. Numerical results show that our GOCE-based Moho model is closer to the all seismic models than that of EGM2008. The model is closest to the regional one with a standard deviation of 5.5 km and a root mean squares error of 7.8 km, which is 2.3 km smaller than the corresponding one based on EGM2008.

  20. P-REX1 amplification promotes progression of cutaneous melanoma via the PAK1/P38/MMP-2 pathway.

    PubMed

    Wang, Jinhua; Hirose, Hajime; Du, Guanhua; Chong, Kelly; Kiyohara, Eiji; Witz, Isaac P; Hoon, Dave S B

    2017-10-28

    P-REX1 (PIP3-dependent Rac exchange factor-1) is a guanine nucleotide exchange factor that activates Rac by catalyzing exchange of GDP for GTP bound to Rac. Aberrant up-regulation of P-REX1 expression has a role in metastasis however, copy number (CN) and function of P-REX1 in cutaneous melanoma are unclear. To explore the role of P-REX1 in melanoma, SNP 6.0 and Exon 1.0 ST microarrays were assessed. There was a higher CN (2.82-fold change) of P-REX1 in melanoma cells than in melanocytes, and P-REX1 expression was significantly correlated with P-REX1 CN. When P-REX1 was knocked down in cells by P-REX1 shRNA, proliferation, colony formation, 3D matrigel growth, and migration/invasiveness were inhibited. Loss of P-REX1 inhibited cell proliferation by inhibiting cyclin D1, blocking cell cycle, and increased cell apoptosis by reducing expression of the protein survivin. Knockdown of P-REX1 expression inhibited cell migration/invasiveness by disrupting P-REX1/RAC1/PAK1/p38/MMP-2 pathway. Assessment of patient tumors and disease outcome demonstrated lower distant metastasis-free survival among AJCC stage I/II/III patients with high P-REX1 expression compared to patients with low P-REX1 expression. These results suggest P-REX1 plays an important role in tumor progression and a potential theranostic target. Copyright © 2017. Published by Elsevier B.V.

  1. Activation of the PAK-Related Kinase by Human Immunodeficiency Virus Type 1 Nef in Primary Human Peripheral Blood Lymphocytes and Macrophages Leads to Phosphorylation of a PIX-p95 Complex

    PubMed Central

    Brown, Amanda; Wang, Xia; Sawai, Earl; Cheng-Mayer, Cecilia

    1999-01-01

    Human immunodeficiency virus type 1 (HIV-1) Nef enhances virus replication in both primary T lymphocytes and monocyte-derived macrophages. This enhancement phenotype has been linked to the ability of Nef to modulate the activity of cellular kinases. We find that despite the reported high-affinity interaction between Nef and the Src kinase Hck in vitro, a Nef-Hck interaction in the context of HIV-1-infected primary macrophages is not detectable. However, Nef binding and activation of the PAK-related kinase and phosphorylation of its substrate could be readily detected in both infected primary T lymphocytes and macrophages. Furthermore, we show that this substrate is a complex composed of the recently characterized PAK interacting partner PIX (PAK-interacting guanine nucleotide exchange factor) and its tightly associated p95 protein. PAK and PIX-p95 appear to be differentially activated and phosphorylated depending on the intracellular environment in which nef is expressed. These results identify the PIX-p95 complex as a novel effector of Nef in primary cells and suggest that the regulation of the PAK signaling pathway may differ in T cells and macrophages. PMID:10559302

  2. p21-Activated kinase 2 (PAK2) inhibits TGF-β signaling in Madin-Darby canine kidney (MDCK) epithelial cells by interfering with the receptor-Smad interaction.

    PubMed

    Yan, Xiaohua; Zhang, Junyu; Sun, Qinyu; Tuazon, Polygena T; Wu, Xiaoping; Traugh, Jolinda A; Chen, Ye-Guang

    2012-04-20

    TGF-β (transforming growth factor β) plays a variety of cellular functions mainly through the Smad pathway. Phosphorylation of the carboxyl SXS motif in R-Smads (Smad2 and Smad3) by the type I receptor TβRI is a key step for their activation. It has been reported that the serine/threonine kinase PAK2 (p21-activated kinase 2) can mediate TGF-β signaling in mesenchymal cells. Here, we report that PAK2 restricts TGF-β-induced Smad2/3 activation and transcriptional responsiveness in MDCK epithelial cells. Mechanistically, PAK2 associates with Smad2 and Smad3 in a kinase activity-dependent manner and blocks their activation. PAK2 phosphorylates Smad2 at Ser-417, which is adjacent to the L3 loop that contributes to the TβRI-R-Smad association. Consistently, substitution of Ser-417 with glutamic acid attenuates the interaction of Smad2 with TβRI. Together, our results indicate that PAK2 negatively modulate TGF-β signaling by attenuating the receptor-Smad interaction and thus Smad activation.

  3. p21-activated Kinase 2 (PAK2) Inhibits TGF-β Signaling in Madin-Darby Canine Kidney (MDCK) Epithelial Cells by Interfering with the Receptor-Smad Interaction*

    PubMed Central

    Yan, Xiaohua; Zhang, Junyu; Sun, Qinyu; Tuazon, Polygena T.; Wu, Xiaoping; Traugh, Jolinda A.; Chen, Ye-Guang

    2012-01-01

    TGF-β (transforming growth factor β) plays a variety of cellular functions mainly through the Smad pathway. Phosphorylation of the carboxyl SXS motif in R-Smads (Smad2 and Smad3) by the type I receptor TβRI is a key step for their activation. It has been reported that the serine/threonine kinase PAK2 (p21-activated kinase 2) can mediate TGF-β signaling in mesenchymal cells. Here, we report that PAK2 restricts TGF-β-induced Smad2/3 activation and transcriptional responsiveness in MDCK epithelial cells. Mechanistically, PAK2 associates with Smad2 and Smad3 in a kinase activity-dependent manner and blocks their activation. PAK2 phosphorylates Smad2 at Ser-417, which is adjacent to the L3 loop that contributes to the TβRI-R-Smad association. Consistently, substitution of Ser-417 with glutamic acid attenuates the interaction of Smad2 with TβRI. Together, our results indicate that PAK2 negatively modulate TGF-β signaling by attenuating the receptor-Smad interaction and thus Smad activation. PMID:22393057

  4. Autocrine VEGF and IL-8 Promote Migration via Src/Vav2/Rac1/PAK1 Signaling in Human Umbilical Vein Endothelial Cells.

    PubMed

    Ju, Li; Zhou, Zhiwen; Jiang, Bo; Lou, Yue; Guo, Xirong

    2017-01-01

    Pro-angiogenic factors VEGF and IL-8 play a major role in modulating the migratory potential of endothelial cells. The goal of this study was to investigate the effect of autocrine VEGF and IL-8 in the form of self-conditioned medium (CM) on human umbilical vein endothelial cells (HUVECs). Enzyme-linked immunosorbent assay (ELISA) examined the automatic secretion of VEGF and IL-8 protein by HUVECs. Western blot, small interfering RNA (siRNA), pulldown and Transwell assays were used to explore the role and the mechanism of autocrine VEGF and IL-8 in migration of HUVECs. Neutralizing VEGF and IL-8 in CM significantly abrogated CM-induced migration of HUVECs. Autocrine VEGF and IL-8 increased Src phosphorylation, Rac1 activity and PAK1 phosphorylation in a time dependent manner. Additionally, blocking Rac1 activity with Rac1 siRNA largely abolished autocrine VEGF and IL-8-induced cell migration. Vav2 siRNA suppressed autocrine VEGF and IL-8-induced Rac1 activation and cell migration. Furthermore, blocking Src signaling with PP2, a specific inhibitor for Src, markedly prevented autocrine VEGF and IL-8-induced Vav2 and Rac1 activation as well as consequently cell migration. PAK1 siRNA also significantly abolished autocrine VEGF and IL-8-induced cell migration. We demonstrated for the first time that autocrine VEGF and IL-8 promoted endothelial cell migration via the Src/Vav2/Rac1/PAK1 signaling pathway. This finding reveals the molecular mechanism in the increase of endothelial cell migration induced by autocrine growth factors and cytokines, which is expected to provide a novel therapeutic target in vascular diseases. © 2017 The Author(s)Published by S. Karger AG, Basel.

  5. The role of remote and regional boundary forcing in the evolution of 2010 summer monsoon heavy rainfall over northwest Indo-Pak region

    NASA Astrophysics Data System (ADS)

    Priya, Pattancheri; Mujumdar, Milind; P, Sabin T.; Pascal, Terray; Krishnan, Raghavan

    2015-04-01

    The evolution of sub-tropical south Asian heavy rainfall during 2010 summer and its relationship with tropical boundary forcing, is diagnosed in this study. Interestingly, the summer time Indo-Pacific SST during 2010 evolved as a combination of three dominant modes of variability. In addition to the strongest La Niña event on long-term record, warming over tropical Indian Ocean and a negative phase of the Indian Ocean Dipole (IOD) phenomenon were also prominent during 2010 boreal summer. A high resolution AGCM is used in this study to distinguish the role of remote and regional tropical SST boundary forcing on the heavy rainfall over northwest Indo-Pak region. The remote boundary forcing over Pacific seems to induce a westward shift of the large-scale monsoon circulation and significantly weakens the convection over Bay of Bengal, but is not sufficient for a realistic simulation of the flood event during 2010. The intensification of northward moisture transport from Arabian Sea into the subtropical Indo-Pak region leading to positive rainfall anomalies as observed in 2010 could be attributed to regional boundary forcing over Indian Ocean. The warmer SST anomalies over southeast Indian Ocean significantly strengthen the cyclonic convergence (divergence) over the region (Bay of Bengal and central and western Indian Ocean). Furthermore, divergence over equatorial Indian Ocean together with positive SST anomalies over north Arabian Sea enhances regional convection and promotes northward moisture transport, which is important for explaining the rainfall anomalies over northwest Indo-Pak region during 2010.

  6. An inherited duplication at the gene p21 Protein-Activated Kinase 7 (PAK7) is a risk factor for psychosis

    PubMed Central

    Morris, Derek W.; Pearson, Richard D.; Cormican, Paul; Kenny, Elaine M.; O'Dushlaine, Colm T.; Perreault, Louis-Philippe Lemieux; Giannoulatou, Eleni; Tropea, Daniela; Maher, Brion S.; Wormley, Brandon; Kelleher, Eric; Fahey, Ciara; Molinos, Ines; Bellini, Stefania; Pirinen, Matti; Strange, Amy; Freeman, Colin; Thiselton, Dawn L.; Elves, Rachel L.; Regan, Regina; Ennis, Sean; Dinan, Timothy G.; McDonald, Colm; Murphy, Kieran C.; O'Callaghan, Eadbhard; Waddington, John L.; Walsh, Dermot; O'Donovan, Michael; Grozeva, Detelina; Craddock, Nick; Stone, Jennifer; Scolnick, Ed; Purcell, Shaun; Sklar, Pamela; Coe, Bradley; Eichler, Evan E.; Ophoff, Roel; Buizer, Jacobine; Szatkiewicz, Jin; Hultman, Christina; Sullivan, Patrick; Gurling, Hugh; Mcquillin, Andrew; St Clair, David; Rees, Elliott; Kirov, George; Walters, James; Blackwood, Douglas; Johnstone, Mandy; Donohoe, Gary; O'Neill, Francis A.; Kendler, Kenneth S.; Gill, Michael; Riley, Brien P.; Spencer, Chris C. A.; Corvin, Aiden

    2014-01-01

    Identifying rare, highly penetrant risk mutations may be an important step in dissecting the molecular etiology of schizophrenia. We conducted a gene-based analysis of large (>100 kb), rare copy-number variants (CNVs) in the Wellcome Trust Case Control Consortium 2 (WTCCC2) schizophrenia sample of 1564 cases and 1748 controls all from Ireland, and further extended the analysis to include an additional 5196 UK controls. We found association with duplications at chr20p12.2 (P = 0.007) and evidence of replication in large independent European schizophrenia (P = 0.052) and UK bipolar disorder case-control cohorts (P = 0.047). A combined analysis of Irish/UK subjects including additional psychosis cases (schizophrenia and bipolar disorder) identified 22 carriers in 11 707 cases and 10 carriers in 21 204 controls [meta-analysis Cochran–Mantel–Haenszel P-value = 2 × 10−4; odds ratio (OR) = 11.3, 95% CI = 3.7, ∞]. Nineteen of the 22 cases and 8 of the 10 controls carried duplications starting at 9.68 Mb with similar breakpoints across samples. By haplotype analysis and sequencing, we identified a tandem ∼149 kb duplication overlapping the gene p21 Protein-Activated Kinase 7 (PAK7, also called PAK5) which was in linkage disequilibrium with local haplotypes (P = 2.5 × 10−21), indicative of a single ancestral duplication event. We confirmed the breakpoints in 8/8 carriers tested and found co-segregation of the duplication with illness in two additional family members of one of the affected probands. We demonstrate that PAK7 is developmentally co-expressed with another known psychosis risk gene (DISC1) suggesting a potential molecular mechanism involving aberrant synapse development and plasticity. PMID:24474471

  7. Phosphorylation of p85 beta PIX, a Rac/Cdc42-specific guanine nucleotide exchange factor, via the Ras/ERK/PAK2 pathway is required for basic fibroblast growth factor-induced neurite outgrowth.

    PubMed

    Shin, Eun-Young; Shin, Kyung-Sun; Lee, Chan-Soo; Woo, Kyung-Nam; Quan, Song-Hua; Soung, Nak-Kyun; Kim, Young Gyu; Cha, Choong Ik; Kim, Seung-Ryul; Park, Dongeun; Bokoch, Gary M; Kim, Eung-Gook

    2002-11-15

    Guanine nucleotide exchange factors (GEFs) have been implicated in growth factor-induced neuronal differentiation through the activation of small GTPases. Although phosphorylation of these GEFs is considered an activation mechanism, little is known about the upstream of PAK-interacting exchange factor (PIX), a member of the Dbl family of GEFs. We report here that phosphorylation of p85 betaPIX/Cool/p85SPR is mediated via the Ras/ERK/PAK2 pathway. To understand the role of p85 betaPIX in basic fibroblast growth factor (bFGF)-induced neurite outgrowth, we established PC12 cell lines that overexpress the fibroblast growth factor receptor-1 in a tetracycline-inducible manner. Treatment with bFGF induces the phosphorylation of p85 betaPIX, as determined by metabolic labeling and mobility shift upon gel electrophoresis. Interestingly, phosphorylation of p85 betaPIX is inhibited by PD98059, a specific MEK inhibitor, suggesting the involvement of the ERK cascade. PAK2, a major PAK isoform in PC12 cells as well as a binding partner of p85 betaPIX, also functions upstream of p85 betaPIX phosphorylation. Surprisingly, PAK2 directly binds to ERK, and its activation is dependent on ERK. p85 betaPIX specifically localizes to the lamellipodia at neuronal growth cones in response to bFGF. A mutant form of p85 betaPIX (S525A/T526A), in which the major phosphorylation sites are replaced by alanine, shows significant defect in targeting. Moreover, expression of the mutant p85 betaPIX efficiently blocks PC12 cell neurite outgrowth. Our study defines a novel signaling pathway for bFGF-induced neurite outgrowth that involves activation of the PAK2-p85 betaPIX complex via the ERK cascade and subsequent translocation of this complex.

  8. Physical, mechanical and hydration kinetics of particleboards manufactured with woody biomass (Cupressus lusitanica, Gmelina arborea, Tectona grandis), agricultural resources, and Tetra Pak packages.

    PubMed

    Moya, Róger; Camacho, Diego; Oporto, Gloria S; Soto, Roy F; Mata, Julio S

    2014-02-01

    Lignocellulosic wastes resulting from agricultural activities as well as Tetra Pak residues from urban centres can cause significant levels of pollution. A possible action to minimize this problem is to use them in the production of particleboards. The purpose of this study was to evaluate the physical, mechanical, and hydration properties of particleboards manufactured with the mixture of woody biomass (Cupressus lusitanica, Gmelina arborea, and Tectona grandis) and either agricultural wastes [pineapple leaves (Ananas comosus) and palm residues (Elaeis guineensis)] or Tetra Pak residues (TP). The results show that the particleboards prepared with TP and woody biomass can reduce the swelling and water absorption in up to 40% and 50% compared with particleboards without TP. Also, these particleboards had increased flexure resistance and shear stress (up to 100%) compared with those without TP. On the contrary, particleboards prepared with pineapple leaves in combination with woody biomass showed the lowest mechanical properties, particularly for tensile strength, hardness, glue-line shear, and nail and screw evaluation.

  9. Induced production of 1-methoxy-indol-3-ylmethyl glucosinolate by jasmonic acid and methyl jasmonate in sprouts and leaves of pak choi (Brassica rapa ssp. chinensis).

    PubMed

    Wiesner, Melanie; Hanschen, Franziska S; Schreiner, Monika; Glatt, Hansruedi; Zrenner, Rita

    2013-07-18

    Pak choi plants (Brassica rapa ssp. chinensis) were treated with different signaling molecules methyl jasmonate, jasmonic acid, linolenic acid, and methyl salicylate and were analyzed for specific changes in their glucosinolate profile. Glucosinolate levels were quantified using HPLC-DAD-UV, with focus on induction of indole glucosinolates and special emphasis on 1-methoxy-indol-3-ylmethyl glucosinolate. Furthermore, the effects of the different signaling molecules on indole glucosinolate accumulation were analyzed on the level of gene expression using semi-quantitative realtime RT-PCR of selected genes. The treatments with signaling molecules were performed on sprouts and mature leaves to determine ontogenetic differences in glucosinolate accumulation and related gene expression. The highest increase of indole glucosinolate levels, with considerable enhancement of the 1-methoxy-indol-3-ylmethyl glucosinolate content, was achieved with treatments of sprouts and mature leaves with methyl jasmonate and jasmonic acid. This increase was accompanied by increased expression of genes putatively involved in the indole glucosinolate biosynthetic pathway. The high levels of indole glucosinolates enabled the plant to preferentially produce the respective breakdown products after tissue damage. Thus, pak choi plants treated with methyl jasmonate or jasmonic acid, are a valuable tool to analyze the specific protection functions of 1-methoxy-indole-3-carbinole in the plants defense strategy in the future.

  10. Induced Production of 1-Methoxy-indol-3-ylmethyl Glucosinolate by Jasmonic Acid and Methyl Jasmonate in Sprouts and Leaves of Pak Choi (Brassica rapa ssp. chinensis)

    PubMed Central

    Wiesner, Melanie; Hanschen, Franziska S.; Schreiner, Monika; Glatt, Hansruedi; Zrenner, Rita

    2013-01-01

    Pak choi plants (Brassica rapa ssp. chinensis) were treated with different signaling molecules methyl jasmonate, jasmonic acid, linolenic acid, and methyl salicylate and were analyzed for specific changes in their glucosinolate profile. Glucosinolate levels were quantified using HPLC-DAD-UV, with focus on induction of indole glucosinolates and special emphasis on 1-methoxy-indol-3-ylmethyl glucosinolate. Furthermore, the effects of the different signaling molecules on indole glucosinolate accumulation were analyzed on the level of gene expression using semi-quantitative realtime RT-PCR of selected genes. The treatments with signaling molecules were performed on sprouts and mature leaves to determine ontogenetic differences in glucosinolate accumulation and related gene expression. The highest increase of indole glucosinolate levels, with considerable enhancement of the 1-methoxy-indol-3-ylmethyl glucosinolate content, was achieved with treatments of sprouts and mature leaves with methyl jasmonate and jasmonic acid. This increase was accompanied by increased expression of genes putatively involved in the indole glucosinolate biosynthetic pathway. The high levels of indole glucosinolates enabled the plant to preferentially produce the respective breakdown products after tissue damage. Thus, pak choi plants treated with methyl jasmonate or jasmonic acid, are a valuable tool to analyze the specific protection functions of 1-methoxy-indole-3-carbinole in the plants defense strategy in the future. PMID:23873294

  11. BET Bromodomain Suppression Inhibits VEGF-induced Angiogenesis and Vascular Permeability by Blocking VEGFR2-mediated Activation of PAK1 and eNOS

    PubMed Central

    Huang, Mingcheng; Qiu, Qian; Xiao, Youjun; Zeng, Shan; Zhan, Mingying; Shi, Maohua; Zou, Yaoyao; Ye, Yujin; Liang, Liuqin; Yang, Xiuyan; Xu, Hanshi

    2016-01-01

    The tyrosine kinase receptor vascular endothelial growth factor receptor 2 (VEGFR2) is a critical modulator of angiogenesis. Increasing evidence indicate the important role of bromodomain and extra-terminal domain (BET) of chromatin adaptors in regulating tumor growth and inflammatory response. However, whether BET proteins have a role in angiogenesis and endothelial permeability is unclear. In this study, we observed that treatment with JQ1, a specific BET inhibitor, suppressed in vitro tube formation of human umbilical vein endothelial cells (HUVECs) and in vivo angiogenesis in a Matrigel plug and oxygen-induced retinopathy neovascularization. JQ1 attenuated the VEGF-induced decrease in TEER in HUVECs and prevented Evans blue dye leakage in the VEGF-induced Miles assay in athymic Balb/c nude mice. BET inhibition with JQ1 or shRNA for Brd2 or Brd4 suppressed VEGF-induced migration, proliferation, and stress fiber formation of HUVECs. Furthermore, BET inhibition suppressed phosphorylation of VEGFR2 and PAK1, as well as eNOS activation in VEGF-stimulated HUVECs. Inhibition with VEGFR2 and PAK1 also reduced migration and proliferation, and attenuated the VEGF-induced decrease in TEER. Thus, our observations suggest the important role of BET bromodomain in regulating VEGF-induced angiogenesis. Strategies that target the BET bromodomain may provide a new therapeutic approach for angiogenesis-related diseases. PMID:27044328

  12. Chagas' disease diagnosis: a multicentric evaluation of Chagas Stat-Pak, a rapid immunochromatographic assay with recombinant proteins of Trypanosoma cruzi.

    PubMed

    Luquetti, Alejandro O; Ponce, Carlos; Ponce, Elisa; Esfandiari, Javan; Schijman, Alejandro; Revollo, Susana; Añez, Nestor; Zingales, Bianca; Ramgel-Aldao, Rafael; Gonzalez, Antonio; Levin, Mariano J; Umezawa, Eufrosina S; Franco da Silveira, José

    2003-08-01

    A rapid serologic test for diagnosis of T. cruzi infection (Chagas Stat Pak) was developed using recombinant proteins in an immunochromatographic assay. This cassette format test was evaluated first in blind with a panel of 393 coded serum samples. The Chagas Stat-Pak identified 197 infected (98.5% sensitivity) and 183 non-infected individuals (94.8% specificity). A second evaluation was performed with 352 sera from four Latin America countries tested independently in each country, showing a sensitivity of 100% and specificity of 98.6%. A third set of tests comparing sera with plasma and eluates from filter paper as well as serum preserved in 50% glycerol did show identical results as those obtained with serum. This rapid test (15 min) uses one device per sample, does not require refrigeration nor a laboratory structure or specialized skills to be performed, accepts different types of samples and may be stored for long periods of time for result checking and documentation. These attributes together with the high sensitivity and specificity demonstrated herein, make this test a suitable tool for field studies, small laboratories and emergencies at blood banks in the countryside of endemic areas.

  13. Comparison of CampyPak II with standard 5% oxygen and candle jars for growth of Campylobacter jejuni from human feces.

    PubMed

    Wang, W L; Luechtefeld, N W; Blaser, M J; Reller, L B

    1982-08-01

    To determine optimal temperature and atmospheric conditions for isolating Campylobacter jejuni from fecal specimens of humans, we studied six laboratory isolates and 19 fecal specimens that were known to contain C. jejuni. We compared incubations in 5% oxygen, the CampyPak II (BBL Microbiology Systems, Cockeysville, Md.) with 6 plates per jar (CP-6) and 12 plates per jar (CP-12), and candle jars at 37 and 42 degrees C. At both temperatures, the colony sizes for the laboratory strains were larger in the 5% O2 and the CP-6 than under the other two conditions. For the primary isolations, CP-12 failed to detect one and two campylobacters at 42 and 37 degrees C, respectively, whereas the candle jar failed to detect one at 42 degrees C and four at 37 degrees C. Colony size was again larger in the 5% O2 and the CP-6. For all four atmospheric conditions tested, colonies were significantly larger at 42 degrees C than at 37 degrees C. These studies showed that incubation at 42 degrees C in either 5% O2 or the CampyPak II with six plates per jar was optimal for primary isolation of C. jejuni from fecal specimens of humans. The candle jars incubated at 42 degrees C appeared to be satisfactory for primary isolation of C. jejuni from human feces, but incubation at 37 degrees C was not acceptable.

  14. Mini-Pak Fencing.

    ERIC Educational Resources Information Center

    Stucky, Franklin; And Others

    The document presents unit plans which offer lists of experiences and competencies to be learned in the construction, planning, and building of fences. The lessons are comprised of seven areas: (1) planning the location and arrangement of fences, (2) fencing and fence building equipment, (3) types of fences, (4) constructing fences, (5) end and…

  15. Will the Need-Based Planning of Health Human Resources Currently Undertaken in Several Countries Lead to Excess Supply and Inefficiency? A Comment on Basu and Pak.

    PubMed

    Birch, Stephen; Tomblin Murphy, Gail; MacKenzie, Adrian; Whittaker, William; Mason, Thomas

    2017-06-01

    Basu and Pak (2014) argue that need-based workforce planning models would not maximize social welfare, and use of need-based models would result in inefficiency. They propose that planning be based on service utilization to incorporate preferences or other socioeconomic factors. We show that the analysis is based on inappropriate considerations of the nature of healthcare demand, a misrepresentation of need-based approaches and misunderstanding publicly funded healthcare system objectives. We explain how current levels of utilization emerge from workload and income interests of providers that underlie utilization-based models and are incompatible with public goals of maximizing health gains. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  16. A protein related to p21-activated kinase (PAK) that is involved in neurogenesis in the Drosophila adult central nervous system.

    PubMed

    Melzig, J; Rein, K H; Schäfer, U; Pfister, H; Jäckle, H; Heisenberg, M; Raabe, T

    1998-11-05

    Brains are organized by the developmental processes generating them. The embryonic neurogenic phase of Drosophila melanogaster has been studied in detail at the genetic, cellular and molecular level. In contrast, much of what is known of postembryonic brain development has been gathered by neuroanatomical and gene expression studies. The molecular mechanisms underlying cellular diversity and structural organisation in the adult brain, such as the establishment of the correct neuroblast number, the spatial and temporal control of neuroblast proliferation, cell fate determination, and the generation of the precise pattern of neuronal connectivity, are largely unknown. In a screen for viable mutations affecting adult central brain structures, we isolated the mushroom bodies tiny (mbt) gene of Drosophila, which encodes a protein related to p21-activated kinase (PAK). We show that mutations in mbt primarily interfere with the generation or survival of the intrinsic cells (Kenyon cells) of the mushroom body, a paired neuropil structure in the adult brain involved in learning and memory.

  17. Characterization and Comparative Genomic Analyses of Pseudomonas aeruginosa Phage PaoP5: New Members Assigned to PAK_P1-like Viruses

    PubMed Central

    Shen, Mengyu; Le, Shuai; Jin, Xiaolin; Li, Gang; Tan, Yinling; Li, Ming; Zhao, Xia; Shen, Wei; Yang, Yuhui; Wang, Jing; Zhu, Hongbin; Li, Shu; Rao, Xiancai; Hu, Fuquan; Lu, Shuguang

    2016-01-01

    As a potential alternative to antibiotics, phages can be used to treat multi-drug resistant bacteria. As such, the biological characteristics of phages should be investigated to utilize them as effective antimicrobial agents. In this study, phage PaoP5, a lytic virus that infects Pseudomonas aeruginosa PAO1, was isolated and genomically characterized. PaoP5 comprises an icosahedral head with an apex diameter of 69 nm and a contractile tail with a length of 120 nm. The PaoP5 genome is a linear dsDNA molecule containing 93,464 base pairs (bp) with 49.51% G + C content of 11 tRNA genes and a 1,200 bp terminal redundancy. A total of 176 protein-coding genes were predicted in the PaoP5 genome. Nine PaoP5 structural proteins were identified. Three hypothetical proteins were determined as structural. Comparative genomic analyses revealed that seven new Pseudomonas phages, namely, PaoP5, K8, C11, vB_PaeM_C2-10_Ab02, vB_PaeM_C2-10_Ab08, vB_PaeM_C2-10_Ab10, and vB_PaeM_C2-10_Ab15, were similar to PAK_P1-like viruses. Phylogenetic and pan-genome analyses suggested that the new phages should be assigned to PAK_P1-like viruses, which possess approximately 100 core genes and 150 accessory genes. This work presents a detailed and comparative analysis of PaoP5 to enhance our understanding of phage biology. PMID:27659070

  18. Paxillin LD4 motif binds PAK and PIX through a novel 95-kD ankyrin repeat, ARF-GAP protein: A role in cytoskeletal remodeling.

    PubMed

    Turner, C E; Brown, M C; Perrotta, J A; Riedy, M C; Nikolopoulos, S N; McDonald, A R; Bagrodia, S; Thomas, S; Leventhal, P S

    1999-05-17

    Paxillin is a focal adhesion adaptor protein involved in the integration of growth factor- and adhesion-mediated signal transduction pathways. Repeats of a leucine-rich sequence named paxillin LD motifs (Brown M.C., M.S. Curtis, and C.E. Turner. 1998. Nature Struct. Biol. 5:677-678) have been implicated in paxillin binding to focal adhesion kinase (FAK) and vinculin. Here we demonstrate that the individual paxillin LD motifs function as discrete and selective protein binding interfaces. A novel scaffolding function is described for paxillin LD4 in the binding of a complex of proteins containing active p21 GTPase-activated kinase (PAK), Nck, and the guanine nucleotide exchange factor, PIX. The association of this complex with paxillin is mediated by a new 95-kD protein, p95PKL (paxillin-kinase linker), which binds directly to paxillin LD4 and PIX. This protein complex also binds to Hic-5, suggesting a conservation of LD function across the paxillin superfamily. Cloning of p95PKL revealed a multidomain protein containing an NH2-terminal ARF-GAP domain, three ankyrin-like repeats, a potential calcium-binding EF hand, calmodulin-binding IQ motifs, a myosin homology domain, and two paxillin-binding subdomains (PBS). Green fluorescent protein- (GFP-) tagged p95PKL localized to focal adhesions/complexes in CHO.K1 cells. Overexpression in neuroblastoma cells of a paxillin LD4 deletion mutant inhibited lamellipodia formation in response to insulin-like growth fac- tor-1. Microinjection of GST-LD4 into NIH3T3 cells significantly decreased cell migration into a wound. These data implicate paxillin as a mediator of p21 GTPase-regulated actin cytoskeletal reorganization through the recruitment to nascent focal adhesion structures of an active PAK/PIX complex potentially via interactions with p95PKL.

  19. Ciliary neurotrophic factor upregulates follistatin and Pak1, causes overexpression of muscle differentiation related genes and downregulation of established atrophy mediators in skeletal muscle.

    PubMed

    Tsompanidis, Alexandros; Vafiadaki, Elizabeth; Blüher, Susann; Kalozoumi, Georgia; Sanoudou, Despina; Mantzoros, Christos S

    2016-06-01

    The Ciliary Neurotrophic Factor (CNTF) is a pluripotent cytokine with anorexigenic actions in the hypothalamus that improves insulin sensitivity, increases energy expenditure and induces weight loss. Since CNTF also has an established myotrophic role, we sought to examine whether skeletal muscle contributes to the CNTF-induced metabolic improvement and identify the molecular mechanisms mediating these effects. We used a mouse model of diet-induced obesity, to which high or low CNTF doses were administered for 7days. Whole transcriptome expression levels were analyzed in dissected soleus muscles using microarrays and data were then confirmed using qRT-PCR. We demonstrate that CNTF administration significantly downregulates leptin, while it upregulates follistatin and Pak1; a molecule associated with insulin sensitization in skeletal muscle. A significant overexpression of muscle differentiation related genes and downregulation of established atrophy mediators was observed. The overall gene expression changes suggest an indirect, beneficial effect of CNTF on metabolism, energy expenditure and insulin sensitivity, exerted by the pronounced stimulation of muscle growth, with similarities to the described effect of follistatin and the activation of the Akt pathway in skeletal muscle. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. Monitoring of tritium, 60Co and 137Cs in the vicinity of the warm water outlet of the Paks Nuclear Power Plant, Hungary.

    PubMed

    Janovics, R; Bihari, Á; Papp, L; Dezső, Z; Major, Z; Sárkány, K E; Bujtás, T; Veres, M; Palcsu, L

    2014-02-01

    Danube water, sediment and various aquatic organisms (snail, mussel, predatory and omnivorous fish) were collected upstream (at a background site) and downstream of the outlet of the warm water channel of Paks Nuclear Power Plant. Gamma emitters, tissue free-water tritium (TFWT) and total organically-bound tritium (T-OBT) measurements were performed. A slight contribution of the power plant to the natural tritium background concentration was measured in water samples from the Danube section downstream of the warm water channel. Sediment samples also contained elevated tritium concentrations, along with a detectable amount of (60)Co. In the case of biota samples, TFWT exhibited only a very slight difference compared to the tritium concentration of the Danube water, however, the OBT was higher than the tritium concentration in the Danube, independent of the origin of the samples. The elevated OBT concentration in the mollusc samples downstream of the warm water channel may be attributed to the excess emission from the nuclear power plant. The whole data set obtained was used for dose rate calculations and will be contributed to the development of the ERICA database.

  1. An experimentally based computer search identifies unstructured membrane-binding sites in proteins: application to class I myosins, PAKS, and CARMIL.

    PubMed

    Brzeska, Hanna; Guag, Jake; Remmert, Kirsten; Chacko, Susan; Korn, Edward D

    2010-02-19

    Programs exist for searching protein sequences for potential membrane-penetrating segments (hydrophobic regions) and for lipid-binding sites with highly defined tertiary structures, such as PH, FERM, C2, ENTH, and other domains. However, a rapidly growing number of membrane-associated proteins (including cytoskeletal proteins, kinases, GTP-binding proteins, and their effectors) bind lipids through less structured regions. Here, we describe the development and testing of a simple computer search program that identifies unstructured potential membrane-binding sites. Initially, we found that both basic and hydrophobic amino acids, irrespective of sequence, contribute to the binding to acidic phospholipid vesicles of synthetic peptides that correspond to the putative membrane-binding domains of Acanthamoeba class I myosins. Based on these results, we modified a hydrophobicity scale giving Arg- and Lys-positive, rather than negative, values. Using this basic and hydrophobic scale with a standard search algorithm, we successfully identified previously determined unstructured membrane-binding sites in all 16 proteins tested. Importantly, basic and hydrophobic searches identified previously unknown potential membrane-binding sites in class I myosins, PAKs and CARMIL (capping protein, Arp2/3, myosin I linker; a membrane-associated cytoskeletal scaffold protein), and synthetic peptides and protein domains containing these newly identified sites bound to acidic phospholipids in vitro.

  2. Targeted genetic dependency screen facilitates identification of actionable mutations in FGFR4, MAP3K9, and PAK5 in lung cancer.

    PubMed

    Fawdar, Shameem; Trotter, Eleanor W; Li, Yaoyong; Stephenson, Natalie L; Hanke, Franziska; Marusiak, Anna A; Edwards, Zoe C; Ientile, Sara; Waszkowycz, Bohdan; Miller, Crispin J; Brognard, John

    2013-07-23

    Approximately 70% of patients with non-small-cell lung cancer present with late-stage disease and have limited treatment options, so there is a pressing need to develop efficacious targeted therapies for these patients. This remains a major challenge as the underlying genetic causes of ~50% of non-small-cell lung cancers remain unknown. Here we demonstrate that a targeted genetic dependency screen is an efficient approach to identify somatic cancer alterations that are functionally important. By using this approach, we have identified three kinases with gain-of-function mutations in lung cancer, namely FGFR4, MAP3K9, and PAK5. Mutations in these kinases are activating toward the ERK pathway, and targeted depletion of the mutated kinases inhibits proliferation, suppresses constitutive activation of downstream signaling pathways, and results in specific killing of the lung cancer cells. Genomic profiling of patients with lung cancer is ushering in an era of personalized medicine; however, lack of actionable mutations presents a significant hurdle. Our study indicates that targeted genetic dependency screens will be an effective strategy to elucidate somatic variants that are essential for lung cancer cell viability.

  3. Growth hormone-releasing hormone receptor antagonists inhibit human gastric cancer through downregulation of PAK1–STAT3/NF-κB signaling

    PubMed Central

    Gan, Jinfeng; Ke, Xiurong; Jiang, Jiali; Dong, Hongmei; Yao, Zhimeng; Lin, Yusheng; Lin, Wan; Wu, Xiao; Yan, Shumei; Zhuang, Yixuan; Chu, Wai Kit; Cai, Renzhi; Zhang, Xianyang; Cheung, Herman S.; Block, Norman L.; Pang, Chi Pui; Schally, Andrew V.; Zhang, Hao

    2016-01-01

    Gastric cancer (GC) ranks as the fourth most frequent in incidence and second in mortality among all cancers worldwide. The development of effective treatment approaches is an urgent requirement. Growth hormone-releasing hormone (GHRH) and GHRH receptor (GHRH-R) have been found to be present in a variety of tumoral tissues and cell lines. Therefore the inhibition of GHRH-R was proposed as a promising approach for the treatment of these cancers. However, little is known about GHRH-R and the relevant therapy in human GC. By survival analyses of multiple cohorts of GC patients, we identified that increased GHRH-R in tumor specimens correlates with poor survival and is an independent predictor of patient prognosis. We next showed that MIA-602, a highly potent GHRH-R antagonist, effectively inhibited GC growth in cultured cells. Further, this inhibitory effect was verified in multiple models of human GC cell lines xenografted into nude mice. Mechanistically, GHRH-R antagonists target GHRH-R and down-regulate the p21-activated kinase 1 (PAK1)-mediated signal transducer and activator of transcription 3 (STAT3)/nuclear factor-κB (NF-κB) inflammatory pathway. Overall, our studies establish GHRH-R as a potential molecular target in human GC and suggest treatment with GHRH-R antagonist as a promising therapeutic intervention for this cancer. PMID:27930339

  4. The metastasis suppressor NDRG1 modulates the phosphorylation and nuclear translocation of β-catenin through mechanisms involving FRAT1 and PAK4.

    PubMed

    Jin, Runsen; Liu, Wensheng; Menezes, Sharleen; Yue, Fei; Zheng, Minhua; Kovacevic, Zaklina; Richardson, Des R

    2014-07-15

    N-myc downstream-regulated gene 1 (NDRG1) is a potent metastasis suppressor that has been demonstrated to inhibit the transforming growth factor β (TGF-β)-induced epithelial-to-mesenchymal transition (EMT) by maintaining the cell-membrane localization of E-cadherin and β-catenin in prostate and colon cancer cells. However, the precise molecular mechanism remains unclear. In this investigation, we demonstrate that NDRG1 inhibits the phosphorylation of β-catenin at Ser33/37 and Thr41 and increases the levels of non-phosphorylated β-catenin at the plasma membrane in DU145 prostate cancer cells and HT29 colon cancer cells. The mechanism of inhibiting β-catenin phosphorylation involves the NDRG1-mediated upregulation of the GSK3β-binding protein FRAT1, which prevents the association of GSK3β with the Axin1-APC-CK1 destruction complex and the subsequent phosphorylation of β-catenin. Additionally, NDRG1 is shown to modulate the WNT-β-catenin pathway by inhibiting the nuclear translocation of β-catenin. This is mediated through an NDRG1-dependent reduction in the nuclear localization of p21-activated kinase 4 (PAK4), which is known to act as a transporter for β-catenin nuclear translocation. The current study is the first to elucidate a unique molecular mechanism involved in the NDRG1-dependent regulation of β-catenin phosphorylation and distribution.

  5. Update of the tectonic model for the Pannonian basin: a contribution to the seismic hazard reassessment of the Paks NPP (Hungary)

    NASA Astrophysics Data System (ADS)

    Horváth, Ferenc; Tóth, Tamás; Wórum, Géza; Koroknai, Balázs; Kádi, Zoltán; Kovács, Gábor; Balázs, Attila; Visnovitz, Ferenc

    2015-04-01

    The planned construction of two new units at the site of the Paks NPP requires a comprehensive site investigation including complete reassessment of the seismic hazard according to the Hungarian as well as international standards. Following the regulations of the Specific Safety Guide no. 9 (IAEA 2010), the approved Hungarian Geological Investigation Program (HGIP) includes integrated geological-geophysical studies at different scales. The regional study aims at to elaborate a new synthesis of all published data for the whole Pannonian basin. This task is nearly completed and the main outcomes have already been published (Horváth et al. 2015). The near regional study is in progress and addresses the construction of a new tectonic model for the circular area with 50 km radius around the NPP using a wealth of unpublished oil company seismic and borehole data. The site vicinity study has also been started with a core activity of 300 km² 3D seismic data acquisition, processing and interpretation assisted by a series of additional geophysical surveys, new drillings and geological mapping. This lecture will present a few important results of the near regional study, which sheds new light on the intricate tectonic evolution of the Mid-Hungarian Fault Zone (MHFZ), which is a strongly deformed belt between the Alcapa and Tisza-Dacia megatectonic units. The nuclear power plant is located at the margin of the Tisza unit near to the southern edge of the MHFZ. Reassessment of seismic hazard at the site of the NPP requires better understanding of the Miocene to Recent tectonic evolution of this region in the central part of the Pannonian basin. Early to Middle Miocene was a period of rifting with formation of 1 to 3 km deep half-grabens filled with terrestrial to marine deposits and large amount of rift-related volcanic material. Graben fill became strongly deformed as a consequence of juxtaposition of the two megatectonic units leading to strong compression and development of

  6. Documentation and Control of Flow Separation on a Low Pressure Turbine Linear Cascade of Pak-B Blades Using Plasma Actuators

    NASA Technical Reports Server (NTRS)

    Corke, Thomas c.; Thomas, FLint, O.; Huang, Junhui

    2007-01-01

    This work involved the documentation and control of flow separation that occurs over low pressure turbine (LPT) blades at low Reynolds numbers. A specially constructed linear cascade was utilized to study the flow field over a generic LPT cascade consisting of Pratt & Whitney "Pak-B" shaped blades. Flow visualization, surface pressure measurements, LDV measurements, and hot-wire anemometry were conducted to examine the flow fields with and without separation control. Experimental conditions were chosen to give a range of chord Reynolds numbers (based on axial chord and inlet velocity) from 10,000 to 100,000, and a range of freestream turbulence intensities from u'/U(infinity) = 0.08 to 2.85 percent. The blade pressure distributions were measured and used to identify the region of separation that depends on Reynolds number and the turbulence intensity. Separation control was performed using dielectric barrier discharge (DBD) plasma actuators. Both steady and unsteady actuation were implemented and found to work well. The comparison between the steady and unsteady actuators showed that the unsteady actuators worked better than the steady ones. For the steady actuators, it was found that the separated region is significantly reduced. For the unsteady actuators, where the signal was pulsed, the separation was eliminated. The total pressure losses (a low Reynolds number) was reduced by approximately a factor of two. It was also found that lowest plasma duty cycle (10 percent in this work) was as effective as the highest plasma duty cycle (50 percent in this work). The mechanisms of the steady and unsteady plasma actuators were studied. It was suggested by the experimental results that the mechanism for the steady actuators is turbulence tripping, while the mechanism for the unsteady actuators is to generate a train of spanwise structures that promote mixing.

  7. Natural or Organic Foods? [Project ECOLogy ELE Pak, Schmidt Pak].

    ERIC Educational Resources Information Center

    Schmidt, Linda

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit is designed for secondary students in home economics classes. The content of the units focuses on natural and organic foods, characteristics of the foods, and uses of the foods. The seven lessons in this unit are designed to last over a…

  8. From Rocks to Pots. [Project ECOLogy ELE Pak, Grim Pak].

    ERIC Educational Resources Information Center

    Grim, Dale

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit is designed for use by art classes at the secondary school level; it illustrates the availability of natural clay and provides the student with experiences such as digging the clay, locating desirable clays, preparing it for production,…

  9. Exponential Explosions! Today.... Tomorrow.... ? [Project ECOLogy ELE Pak, Jensen Pak].

    ERIC Educational Resources Information Center

    Jensen, Melanie

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit is designed for junior high school mathematics classes and emphasizes applications of exponents to problems of population growth and pollution. The nine lessons are designed for about eleven school days. Each lesson includes the concept…

  10. Your Nose Knows. [Project ECOLogy ELE Pak, Meaney Pak].

    ERIC Educational Resources Information Center

    Meaney, Marie

    This is one of a series of units for environmental education developed by the Highline Public Schools. This unit is designed for kindergarten pupils, but could be used effectively with primary pupils. The six lessons use the sense of smell to investigate various aspects of the earth, seasons, animals, and commercial use of fragrances. Each lesson…

  11. [Peabody's Time Machine. Project ECOLogy ELE Pak, Hirschel Pak].

    ERIC Educational Resources Information Center

    Hirschel, John

    This is one of a series of units for environmental education developed by the Highline Public Schools. This unit is designed for use by a substitute teacher for instruction of intermediate grade elementary school pupils. The kit provides three days of lesson plans. There is closure at the end of each lesson and each day. Much emphasis is placed on…

  12. Anthropology - Ecology. [Project ECOLogy ELE Pak, Skidmore Pak].

    ERIC Educational Resources Information Center

    Skidmore, Margaret

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit may be used as an introduction to the study of anthropology, the influence of ecology on the study of anthropology, and an introduction to the physical school environment. For best results, it should be used at the beginning of the…

  13. And Then - Recycling. [Project ECOLogy ELE Pak, Lewis Pak].

    ERIC Educational Resources Information Center

    Lewis, Dave

    This is one of a series of units for environmental education developed by the Highline Public Schools. There are seven concepts in this unit on recycling which is designed for grade five and six students. Each concept has one lesson that is comprised of several activities. Several suggested extra activities have been added to further the students'…

  14. River sinuosity changes as indicators of the possible neotectonic activity - a case study on the Danube River between Paks (Hungary) and Beograd (Serbia)

    NASA Astrophysics Data System (ADS)

    Petrovszki, Judit

    2010-05-01

    The meandering, pre-regulation river planforms of the Danube River, between Paks (Hungary) and Beograd (Serbia) was digitized from the map sheets of the Second Military Survey of the Habsburg Empire (Timár et al., 2006). These maps were surveyed before or simultaneously with the river control works, so it is possible to follow the natural riverbeds, the natural changing of the meandering structure. The sinuosity values were calculated with different window sizes, and displayed in a spectrum-like diagram (sinuosity spectra; after van Balen et al., 2008). The channel sinuosity of this river is analyzed in order to draw conclusions on the neotectonic activity of the western part of the Great Hungarian Plain. Several points of sinuosity change were identified. To prove that these are of neotectonic origin, a neotectonic map and seismic sections crossing the study area, were also analyzed. Significant sinuosity changes (low to high or high to low), spatially correlated to linear features identified in seismic survey sections or in tectonic maps (Horváth et al., 2006), indicate their neotectonic activity (Ouchi, 1985; Timár, 2003; Zámolyi et al., 2010). Upstream of the Hungarian-Serbian border, the Duna (Danube) has anabranching planform, the Baracskai-Duna is the main anabranch. There is a fault on the neotectonic map, crossing both rivers, and cause the decreasing of the sinuosity. The vertical activity of the structural line, which is more or less parallel to the international border, is verified by the sinuosity change. The direction of the change (from high to low sinuosity values) correlates with the normal fault character, shown on the map. Another significant sinuosity change occurs downstream of the Drava River confluence. The explanation of this change can be of two kinds. First, there is a known tectonic feature along the Drava River, with dextral faulting. The sinuosity increase could indicate a small active vertical component of this structural line

  15. Prolactin-Stimulated Activation of ERK1/2 Mitogen-Activated Protein Kinases is Controlled by PI3-Kinase/Rac/PAK Signaling Pathway in Breast Cancer Cells

    PubMed Central

    Aksamitiene, Edita; Achanta, Sirisha; Kolch, Walter; Kholodenko, Boris N.; Hoek, Jan B.; Kiyatkin, Anatoly

    2011-01-01

    There is strong evidence that deregulation of prolactin (PRL) signaling contributes to pathogenesis and chemoresistance of breast cancer. Therefore, understanding cross-talk between distinct signal transduction pathways triggered by activation of the prolactin receptor (PRL-R), is essential for elucidating the pathogenesis of metastatic breast cancer. In this study, we applied a sequential inhibitory analysis of various signaling intermediates to examine the hierarchy of protein interactions within the PRL signaling network and to evaluate the relative contributions of multiple signaling branches downstream of PRL-R to the activation of the extracellular signal-regulated kinases ERK1 and ERK2 in T47D and MCF-7 human breast cancer cells. Quantitative measurements of the phosphorylation/activation patterns of proteins showed that PRL simultaneously activated Src family kinases (SFKs) and the JAK/STAT, phosphoinositide-3 (PI3)-kinase/Akt and MAPK signaling pathways. The specific blockade or siRNA-mediated suppression of SFK/FAK, JAK2/STAT5, PI3-kinase/PDK1/Akt, Rac/PAK or Ras regulatory circuits revealed that (1) the PI3-kinase/Akt pathway is required for activation of the MAPK/ERK signaling cascade upon PRL stimulation; (2) PI3-kinase-mediated activation of the c-Raf-MEK1/2-ERK1/2 cascade occurs independent of signaling dowstream of STATs, Akt and PKC, but requires JAK2, SFKs and FAK activities; (3) activated PRL-R mainly utilizes the PI3-kinase-dependent Rac/PAK pathway rather than the canonical Shc/Grb2/SOS/Ras route to initiate and sustain ERK1/2 signaling. By interconnecting diverse signaling pathways PLR may enhance proliferation, survival, migration and invasiveness of breast cancer cells. PMID:21726627

  16. Impact of different feedstocks derived biochar amendment with cadmium low uptake affinity cultivar of pak choi (Brassica rapa ssb. chinensis L.) on phytoavoidation of Cd to reduce potential dietary toxicity.

    PubMed

    Yasmin Khan, Kiran; Ali, Barkat; Cui, Xiaoqiang; Feng, Ying; Yang, Xiaoe; Joseph Stoffella, Peter

    2017-07-01

    Biochar has become eco-friendly amendment used for phytoavoidation with low cadmium (Cd) accumulating cultivars of crops to ensure food safety in Cd contaminated soils. In this study, biochar with different waste feedstock material were evaluated for their effectiveness on essential trace metals mobility, Cd bioavailability and its accumulation in two contrasting Cd accumulating cultivars of pak choi (Brassica rapa ssp. chinensis L.) grown in Cd contaminated Mollisol soil. A greenhouse experiment was conducted with plants grown in Cd contaminated soil that had been amended with biochar derived from barley straw, tomato green waste, chicken manure, duck manure and swine manure at application rate of 0%, 2.5% and 5.0% (w/w). The results showed that soil pH was significantly increased by all treatments. Biochar increased plant dry biomass, micronutrients bioavailability with significant differences in the Cd sorption capacity, with the effectiveness higher with increasing biochar application rate. However, tomato green waste (TGW) and chicken manure (CM) derived biochar were more effective than the other biochar in reducing Cd mobilization in soil by 35-54% and 26-43% and reduced its accumulation in shoots of pak choi cultivars by 34-76% and 33-72% in low Cd accumulator cultivar and 64-85% and 55-80% in high Cd accumulator cultivar than the control. Overall, results indicate that TGW and CM biochar can efficiently immobilize Cd, thereby reducing bioavailability in Cd contaminated Mollisol soil to ensure food safety. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Hemopoietic cell kinase (Hck) and p21-activated kinase 2 (PAK2) are involved in the down-regulation of CD1a lipid antigen presentation by HIV-1 Nef in dendritic cells.

    PubMed

    Shinya, Eiji; Shimizu, Masumi; Owaki, Atsuko; Paoletti, Samantha; Mori, Lucia; De Libero, Gennaro; Takahashi, Hidemi

    2016-01-01

    Dendritic cells (DCs) play a major role in in vivo pathogenesis of HIV-1 infection. Therefore, DCs may provide a promising strategy to control and eventually overcome the fatal infection. Especially, immature DCs express all CD1s, the non-MHC lipid antigen -presenting molecules, and HIV-1 Nef down-regulates CD1 expression besides MHC. Moreover, CD1d-restricted CD4(+) NKT cells are infected by HIV-1, reducing the number of these cells in HIV-1-infected individuals. To understand the exact role of DCs and CD1-mediated immune response during HIV-1 infection, Nef down-regulation of CD1a-restricted lipid/glycolipid Ag presentation in iDCs was analyzed. We demonstrated the involvement of the association of Nef with hemopoietic cell kinase (Hck) and p21-activated kinase 2 (PAK2), and that Hck, which is expressed strongly in iDCs, augmented this mutual interaction. Hck might be another therapeutic target to preserve the function of HIV-1 infected DCs, which are potential reservoirs of HIV-1 even after antiretroviral therapy.

  18. Untersuchungen zur Sorptionsreversibilität von organischen Schadstoffen in Aktivkohle, Holzkohle und Zeolith Y-200

    NASA Astrophysics Data System (ADS)

    ElHaddad, Engy; Ensinger, Wolfgang; Schüth, Christoph

    2013-09-01

    Numerous studies have shown that sorption of organic contaminants in soils is dominated by the natural organic carbon content ( C org) of the soil. However, it is still under discussion whether sorption processes are fully reversible or whether an irreversibly sorbed contaminant fraction remains in the soil. This is especially important when considering soil remediation measures and its targets. In multi-stage sorption-desorption batch experiments with TCE, PCE, ortho-xylene and para-xylene and with the sorbents activated carbon, charcoal and a hydrophobic zeolite Y-200, the reversibility of sorption was studied. It could be shown that the structural features of the sorbents are of ample importance for the occurrence of a desorption-resistant fraction. While sorption was mainly reversible for the micro-porous zeolite Y-200 with a rigid pore network, charcoal and the activated carbon showed significant desorption hysteresis. However, following a subsequent sorption step, this fraction eventually desorbs and is re-mobilized.

  19. Food: The Challenge to Manage. [Project ECOLogy ELE Pak, Roush Pak].

    ERIC Educational Resources Information Center

    Roush, Judy

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit is designed for students at the senior high level who have a basic knowledge of nutrition, some experience in menu planning, and who are ready to put this knowledge of nutrition to work in selecting foods to attain maximum nutrition with…

  20. Eco-Kids Fly Off to the Forests. [Project ECOLogy ELE Pak, Bell Pak].

    ERIC Educational Resources Information Center

    Bell, Loretta

    This is one of a series of units for environmental education developed by the Highline Public Schools. This unit was designed for use with fourth-grade students; it focuses on three forest biomes. Each of the biomes has characteristics of its own. The unit includes eight lessons, as well as additional activities. The unit, which incorporates…

  1. Round and Round It Goes: A Study of Ecological Cycles. [Project ECOLogy ELE Pak, Roaa Pak].

    ERIC Educational Resources Information Center

    Ross, Catherine

    This is one of a series of units for environmental education developed by the Highline Public Schools. This unit, designed for third- and fourth-grade students, emphasizes cycles and focuses on the water, oxygen, and nutrient cycles. The eleven lessons in this unit are designed to take one-half to one hour each. Use of the extra activities would…

  2. ...About This Problem of Air Pollution... . [Project ECOLogy ELE Pak, Wright Pak].

    ERIC Educational Resources Information Center

    Wright, Jan

    This is one of a series of units for environmental education developed by the Highline Public Schools. The lessons in this unit are designed to help students discover causes, effects, and results of air pollution through involvement in various activities; it is recommended for intermediate grade elementary school pupils. The unit can be used…

  3. Eco-Kids: Experiment with Air on Spaceship Earth. [Project ECOLogy ELE Pak, Bell Pak].

    ERIC Educational Resources Information Center

    Bell, Loretta

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit, designed for intermediate grades in the elementary schools, is concerned with the study of air, air pollution, effects of air pollution, and ways of improving the quality of the air. Six lessons are included in the unit; most of the…

  4. This Land Is Your Land. [Project ECOLogy ELE Pak, Weber Pak].

    ERIC Educational Resources Information Center

    Weber, Lee

    This is one of a series of units for environmental education developed by the Highline Public Schools. The unit has been constructed for use by intermediate grade elementary school pupils. The seven lessons are designed to inform students about the land to develop a land ethic. The unit should be able to be completed in two to three weeks. The…

  5. Exploration with Garbage. [Project ECOLogy ELE Pak, Lund and Wolff Pak].

    ERIC Educational Resources Information Center

    Lund, Cherie; Wolff, Chanelle

    This is one of a series of units for environmental education developed by the Highline Public Schools. This unit is concerned with the topic of garbage. The eleven lessons explore what garbage is, problems of littering, ways to reduce garbage, and ways to use garbage. The materials were designed to be used with kindergarten pupils, but could be…

  6. Development of a simple valid method for the complete removal of insulin-like growth factor (IGF)-binding proteins from IGFs in human serum and other biological fluids: comparison with acid-ethanol treatment and C18 Sep-Pak separation.

    PubMed

    Mohan, S; Baylink, D J

    1995-02-01

    Insulin-like growth factor (IGF)s circulate in plasma as large mol wt proteins bound to specific proteins, termed IGF-binding proteins (IGFBPs). As IGFBPs have been shown to produce artifacts in IGF radioligand assays, various extraction procedures have been proposed to eliminate IGFBPs from biological samples before radioligand assays. Comparison of acid-ethanol and C18 Sep-Pak extraction methods, the two most widely used procedures for separation of IGFs from IGFBPs in human serum samples, with the established gold standard (Sephadex G-75 acid gel filtration) revealed that a significant amount of IGFBP activity survived the acid-ethanol extraction and C18 Sep-Pak separation techniques. We, therefore, have developed a simple novel method comprising a combination of two techniques, involving separation based on size and separation based on centrifugation. In this method, serum samples were acidified and applied to Bio-Spin columns containing BSA-pretreated Bio-Gel Polyacrylamide-10 (P-10). Upon centrifugation, IGFBPs eluted in the void volume. IGFs were then eluted with 1 mol/L acetic acid containing 0.1 mol/L NaCl upon subsequent centrifugation. The efficacy of Bio-Spin P-10 separation for the complete removal of IGFBPs was determined by Western ligand blot analysis and determination of IGFBP-3 levels by RIA in the extracted serum samples. The recovery of exogenously added IGF-I to the serum samples was greater than 90%. Comparison of IGF-I and IGF-II values determined in 12 human serum samples after Bio-Spin P-10 separation with those obtained after separation with the established gold standard method (Sephadex G-75) revealed a correlation greater than 0.9. In contrast to the established gold standard method, which is tedious and time consuming, Bio-Spin P-10 separation offers the advantage of speed, such that 50 or more samples can be processed in less than 4-6 h. Application of Bio-Gel P-10 gel filtration to determine the IGF-I and IGF-II levels in 14 normal

  7. The Teacher's Planning Pak and Guide to Individualized Instruction.

    ERIC Educational Resources Information Center

    Forte, Imogene; MacKenzie, Joy

    This book is for both elementary school teachers and students. Suggestions are provided for the teacher on ways to organize time, space, and materials with the emphasis on individualized instruction for the pupils. Materials for the students include activities and small questionnaires structured to reveal to the teacher each child's attitudes,…

  8. OPTIMIZING THE PAKS METHOD FOR MEASURING AIRBORNE ACROLEIN

    EPA Science Inventory

    Airborne acrolein is produced from the combustion of fuel and tobacco and is of concern due to its potential for respiratory tract irritation and other adverse health effects. DNPH active-sampling is a method widely used for sampling airborne aldehydes and ketones (carbonyls); ...

  9. OPTIMIZING THE PAKS METHOD FOR MEASURING AIRBORNE ACROLEIN

    EPA Science Inventory

    Airborne acrolein is produced from the combustion of fuel and tobacco and is of concern due to its potential for respiratory tract irritation and other adverse health effects. DNPH active-sampling is a method widely used for sampling airborne aldehydes and ketones (carbonyls); ...

  10. Ever Stop to Think Man's Survival Is Dependent on His Use of Food Resources? [Project ECOLogy ELE Pak, Nelson Pak].

    ERIC Educational Resources Information Center

    Nelson, Judy

    This is one of a series of units for environmental education developed by the Highline Public Schools. This unit is designed for senior high school students who have a basic knowledge of nutrition and some experience in menu planning. The five lessons provide experiences in selecting, preparing, and storing foods to attain maximum nutrition with a…

  11. Restoring fire suppressed Texas pak woodlands to historic conditions using prescribed fire

    Treesearch

    Jeff C. Sparks; Michael C. Stambaugh; Eric L. Keith

    2012-01-01

    Comparable to many oak ecosystems across the eastern United States, oak woodlands in Texas display characteristics of changing composition and structure due to altered fire regimes. Information describing historic fire regimes suggests woodlands underwent relatively frequent and repeated burning prior to major Euro-American influence in the early 19th century. Oak...

  12. PAKS: Parents-and-Kids Science. 24 Activities for Kids and Adults To Share.

    ERIC Educational Resources Information Center

    McKenzie, Danny L.

    This activity book designed for grades 1-3 provides teachers with ready-to-use materials designed to get parents and children excited about science, help establish a home-school connection, and provide interesting learning activities for children to share with adults. This program gets parents involved in developing their children's science…

  13. Satellite based classification (haze, fog) and affected area estimation over Indo - Pak Sub-Continent

    NASA Astrophysics Data System (ADS)

    Ghauri, Badar; Zafar, Sumaira

    2016-07-01

    Northern Pakistan and bordering Indian Punjab experience intense smog and fog during fall and winters. Environmentalists have been raising their voices over the situation and demanded control over regional emissions to save the livelihood of millions of dwellers whose trade, commerce and agriculture is at stake because of long smog/ fog spells.. This paper estimates the area affected by haze, smog and fog during 2006- 2010. MODIS (geo-referenced MODIS subsets India1, 2 &3) of the area in Pakistan and India from 2006 to 2010 for the period October to February) were analyzed using state of the art software ENVI 4.2 and ArcGIS 10.2. This process resulted in area belonging to each class that is; haze, smog and fog. On the basis of density, haze and fog cover was determined. Variations in fog cover, its density and identification of location of fog initiation process were also determined using near real time (30 minutes) METEOSAT-7 IODC data where actually fog formation started and then extended to the area of favorable conditions. Haze has been noticed to intensify due to massive burning of agricultural waste (rice husk) in India and Pakistan towards the end of October each year. MODIS thermal anomalies/fire data (MYD 14) were also used to verify this activity on the ground, which results in hazy conditions at regional level during fall months. Haze-affected area during 2006 to 2010 in Pakistan ranged from 155,000 Km2 to 354,000 Km2 and in India it ranged from 333,000 Km2 to 846,000 Km2. Similarly winter fog cover during this period in Pakistan varied from 136,000 Km2 to 381,000 Km2 and in India it was estimated at 327,000 Km2 to 566,000 Km2. This phenomenon was more prominent in India than in Pakistan where and fog cover was at least twice than that was observed in Pakistan. It has been noted that area covered by fog, smog and haze doubled during the study period in the region. Atmospheric dimming during autumn/ fall also reduces the mixing height leading to greater pollutants accumulation. So far no mitigation steps have been taken to combat this regional issue. Reduction in local emissions is highly recommended to save at least the lives of vulnerable (children, elderly, patients etc).

  14. Prevalence of Leishmania tropica in school boys of khyber agency, FATA near Pak-Afghan border.

    PubMed

    Qureshi, Naveeda Akhtar; Ali, Abid; Rashid, Umer; Tayyab-Ur-Rehman; Ali, Naeem

    2016-12-01

    In Pakistan leishmaniasis occurs periodically throughout the year and various out breaks are reported frequently. In continuation of our research on this neglected disease, the aim of present study is to explore: (1) the prevalence of cutaneous leishmaniasis in school boys; (2) Leishmania species identification in order to epidemiology and dynamics of the disease; (3) Identification of risk factors for Leishmaniasis especially for CL. The data was collected in August 2014 The experimental strategy involved a questionnaire for data collection and along with clinical diagnosis of 134 out of 9368 students for incidence of Leishmania spp. in 7 square kilometres area in the schoolboys at Tehsil Landi Kotal, District Khyber Agency, FATA Pakistan. The parasitological and molecular diagnosis of clinically suspected cutaneous leishmaniasis cases were performed using microscopical examination of Giemsa-stained smears of lesion exudates and minicircle kDNA semi nested PCR, respectively. Microscopy (x=1000) positive cases were 84/134 (62.6%) and 50/134 (37.4%) slides did not showed any presence of amastigotes of Leishmania spp. The samples were amplified using kDNA semi nested PCR and confirmed the presence of L. Tropica (Ac.no KT 985473). PCR positive cases were 97/134 (72.4%) and 37(27.6%) were negative. The prevalence of L. tropica in school boys was 1.4% (134/9368) in the total population studied (n=9368). The parasite prevalence might be greater as only male students were considered in the study due to ethical and social issues and limitations.

  15. The closure of Pak1-dependent macropinosomes requires the phosphorylation of CtBP1/BARS.

    PubMed

    Liberali, Prisca; Kakkonen, Elina; Turacchio, Gabriele; Valente, Carmen; Spaar, Alexander; Perinetti, Giuseppe; Böckmann, Rainer A; Corda, Daniela; Colanzi, Antonino; Marjomaki, Varpu; Luini, Alberto

    2008-04-09

    Membrane fission is an essential process in membrane trafficking and other cellular functions. While many fissioning and trafficking steps are mediated by the large GTPase dynamin, some fission events are dynamin independent and involve C-terminal-binding protein-1/brefeldinA-ADP ribosylated substrate (CtBP1/BARS). To gain an insight into the molecular mechanisms of CtBP1/BARS in fission, we have studied the role of this protein in macropinocytosis, a dynamin-independent endocytic pathway that can be synchronously activated by growth factors. Here, we show that upon activation of the epidermal growth factor receptor, CtBP1/BARS is (a) translocated to the macropinocytic cup and its surrounding membrane, (b) required for the fission of the macropinocytic cup and (c) phosphorylated on a specific serine that is a substrate for p21-activated kinase, with this phosphorylation being essential for the fission of the macropinocytic cup. Importantly, we also show that CtBP1/BARS is required for macropinocytic internalization and infection of echovirus 1. These results provide an insight into the molecular mechanisms of CtBP1/BARS activation in membrane fissioning, and extend the relevance of CtBP1/BARS-induced fission to human viral infection.

  16. Relationship Between Pak-Mediated Cell Death and Stress-Activated Kinase Signaling Breast Cancer

    DTIC Science & Technology

    2001-02-01

    Jurkat: T-lymphoblast cell line; HeLa: human cervical carcinoma; CHO : chinese hamster ovary ; ZR75, MDA23 1, SKBR-3: human breast cancer cell lines 16 RhoA...activation in Jurkat cells . J Immunol 1998 Jan 1; 160(1):7-1 1 12 Genomic Locus of GEF/H1/KIAA0651 GI 11427616: 870353 864048 861008 858631 858224...1: Schematic representation of the genomic locus of GEF-H1i/KIAA0651 as deduced from the working draft sequence of the GI 11427616 contig derived from

  17. STS-93 Columbia, Fit Check and Pre Pak in the O&C for Chandra

    NASA Technical Reports Server (NTRS)

    1999-01-01

    The primary objective of the STS-93 mission was to deploy the Advanced X-ray Astrophysical Facility, which had been renamed the Chandra X-ray Observatory in honor of the late Indian-American Nobel Laureate Subrahmanyan Chandrasekhar. The mission was launched at 12:31 on July 23, 1999 onboard the space shuttle Columbia. The mission was led by Commander Eileen Collins. The crew was Pilot Jeff Ashby and Mission Specialists Cady Coleman, Steve Hawley and Michel Tognini from the Centre National d'Etudes Spatiales (CNES). This videotape shows the astronauts getting into spacesuits, and inspecting the equipment.

  18. UTILIZING THE PAKS METHOD FOR MEASURING ACROLEIN AND OTHER ALDEHYDES IN DEARS

    EPA Science Inventory

    Acrolein is a hazardous air pollutant of high priority due to its high irritation potency and other potential adverse health effects. However, a reliable method is currently unavailable for measuring airborne acrolein at typical environmental levels. In the Detroit Exposure and A...

  19. Lipopeptide biosurfactant from Bacillus thuringiensis pak2310: A potential antagonist against Fusarium oxysporum.

    PubMed

    Deepak, R; Jayapradha, R

    2015-03-01

    The aims of the study were to evaluate the effects of a biosurfactant obtained from a novel Bacillus thuringiensis on Fusarium oxysporum to determine the morphological changes in the structure of the fungi and its biofilm in the presence of the biosurfactant and to evaluate the toxicity of the biosurfactant on HEp-2 human epithelial cell lines. The strain was screened and isolated from petroleum contaminated soil based on the E24 emulsification index. The biosurfactant was produced on glycerol, extracted using chloroform:methanol system and purified using HPLC. The purified fraction showing both surface activity (emulsification and oil-spread activity) and anti-fusarial activity (agar well diffusion method) was studied using FT-IR and MALDI-TOF MS, respectively. The minimum inhibitory concentration (MIC) and the biofilm inhibitory concentration (BIC) were determined using dilution method. The effect of biosurfactant on the morphology of Fusarium oxysporum was monitored using light microscopy and confocal laser scanning microscopy (for biofilm). The purified surfactant showed the presence of functional groups like that of surfactin in the FT-IR spectra and MALDI-TOF MS estimated the molecular weight as 700Da. The MIC and BIC were estimated to be 0.05 and 0.5mg/mL, respectively. The molecule was also non-toxic to HEp-2 cell lines at 10× MIC. A non-toxic and effective anti-Fusarium biosurfactant, that is both safe for human use and to the environment, has been characterized. The growth and metabolite production using glycerol (major byproduct of biodiesel and soap industries) also adds up to the efficiency and ecofriendly nature of this biosurfactant. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  20. Career Education Programs for Educable Mentally Retarded. Info-Pak 2, Selected Readings.

    ERIC Educational Resources Information Center

    Michigan State Univ., East Lansing. Regional Instructional Materials Center for Handicapped Children and Youth.

    The information packet contains six abridged readings on career education programs for educable mentally retarded (EMR) adolescents and young adults. A driver training program is discussed which serves special needs of EMR students and is based on the premise that travel independence provides more vocational opportunity. A guidebook presents facts…

  1. UTILIZING THE PAKS METHOD FOR MEASURING ACROLEIN AND OTHER ALDEHYDES IN DEARS

    EPA Science Inventory

    Acrolein is a hazardous air pollutant of high priority due to its high irritation potency and other potential adverse health effects. However, a reliable method is currently unavailable for measuring airborne acrolein at typical environmental levels. In the Detroit Exposure and A...

  2. Pattern of fall injuries in Pakistan: the Pakistan National Emergency Department Surveillance (Pak-NEDS) study

    PubMed Central

    2015-01-01

    Background We aimed to analyse the frequency and patterns of fall-related injuries presenting to the emergency departments (EDs) across Pakistan. Methods Pakistan National Emergency Departments surveillance system collected data from November 2010 to March 2011 on a 24/7 basis using a standardized tool in seven major EDs (five public and two private hospitals) in six major cities of Pakistan. For all patients presenting with fall-related injuries, we analysed data by intent with focus on unintentional falls. Simple frequencies were run for basic patient demographics, mechanism of falls, outcomes of fall injuries, mode of arrival to ED, investigations, and procedures with outcomes. Results There were 3335 fall-related injuries. In cases where intent was available, two-thirds (n = 1186, 65.3%) of fall injuries were unintentional. Among unintentional fall patients presenting to EDs, the majority (76.9%) were males and between 15-44 years of age (69%). The majority of the unintentional falls (n = 671, 56.6%) were due to slipping, followed by fall from height (n = 338, 28.5%). About two-thirds (n = 675, 66.6%) of fall injuries involved extremities, followed by head/neck (n = 257, 25.4%) and face (n = 99, 9.8%). Most of the patients were discharged from the hospital (n = 1059, 89.3%). There were 17 (1.3%) deaths among unintentional fall cases. Conclusion Falls are an important cause of injury-related visits to EDs in Pakistan. Most of the fall injury patients were men and in a productive age group. Fall injuries pose a burden on the healthcare system, especially emergency services, and future studies should therefore focus on safety measures at home and in workplaces to reduce this burden. PMID:26691821

  3. The Pak-U.S. Alliance in the Fight Against Terrorism: A Cost-Benefit Analysis

    DTIC Science & Technology

    2011-12-01

    97 3. Cultural and Environmental Blocks ................................................97 F. CHANGING PERCEPTIONS THROUGH MICRO...bringing substantial changes . Each sector and sub-sector should have centralized accountability at the USAID level. Besides development, it is evident...are able to promote commonalities, understand each other‘s limitations, change their policies and approach the issues with converging interests

  4. Changes in the distributions of juvenile horseshoe crabs (Arthropoda: Chelicerata) (2002-2014) related to environmental perturbations at Pak Nai and Ha Pak Nai, Deep Bay, Hong Kong SAR, China.

    PubMed

    Lee, Christine Nga-Wing; Morton, Brian

    2016-07-15

    A survey of juvenile Asian horseshoe crabs in 2002 on the mudflats along Hong Kong's north-western shoreline abutting Deep Bay identified two species, Tachypleus tridentatus and Carcinoscorpius rotundicauda, and assessed their population characteristics. Since the 2002 survey, there have been significant habitat changes to this natal site for the two species. By employing the same, but expanded, sampling protocol, a further survey was, therefore, conducted twelve years later in 2014. A general population decline was recorded for T. tridentatus whereas for C. rotundicauda there was an increase and its distribution had become more widespread. The distribution patterns of the two species were also shown to have changed. The potential factors that might be responsible for recorded changes in the species' population characteristics between 2002 and 2014 are related to anthropogenic perturbations, including environmental habitat alterations notably the building of a bridge linking Hong Kong to Shenzhen in China. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. Cooperative measures to support the Indo-Pak Agreement Reducing Risk from Accidents Relating to Nuclear Weapons.

    SciTech Connect

    Mishra, Sitakanta; Ahmed, Mansoor

    2014-04-01

    In 2012, India and Pakistan reaffirmed the Agreement on Reducing the Risk from Accidents Relating to Nuclear Weapons. Despite a history of mutual animosity and persistent conflict between the two countries, this agreement derives strength from a few successful nuclear confidence building measures that have stood the test of time. It also rests on the hope that the region would be spared a nuclear holocaust from an accidental nuclear weapon detonation that might be misconstrued as a deliberate use of a weapon by the other side. This study brings together two emerging strategic analysts from South Asia to explore measures to support the Agreement and further develop cooperation around this critical issue. This study briefly dwells upon the strategic landscape of nuclear South Asia with the respective nuclear force management structures, doctrines, and postures of India and Pakistan. It outlines the measures in place for the physical protection and safety of nuclear warheads, nuclear materials, and command and control mechanisms in the two countries, and it goes on to identify the prominent, emerging challenges posed by the introduction of new weapon technologies and modernization of the respective strategic forces. This is followed by an analysis of the agreement itself leading up to a proposed framework for cooperative measures that might enhance the spirit and implementation of the agreement.

  6. A School Improvement-Accountability Process Kit. PAK No. 4.4--Putting Staff Objectives Into Operation.

    ERIC Educational Resources Information Center

    Colorado State Dept. of Education, Denver. District Planning and Accountability Services.

    This Personalized Activity Kit focuses on one aspect of management by objectives--procedures for developing a plan that will help the participant meet his staff objectives. One way to report progress to one's superior is also suggested. Staff objectives state who, by job title, will do what. The plan sheet is made up of hows with deadline dates.…

  7. Bomb blast injuries: an exploration of patient characteristics and outcome using Pakistan National Emergency Departments Surveillance (Pak-NEDS) data

    PubMed Central

    2015-01-01

    Background Bomb blast injuries result in premature deaths and burdening of healthcare systems. The objective of this study was to explore the characteristics and outcome of patients presenting to the emergency departments in Pakistan with bomb blast injuries. Methods Active surveillance was conducted in seven major emergency departments of Pakistan from November 2010-March 2011. All the sites are tertiary care urban centers. All the patients who presented to the hospital's emergency department (ED) following a bomb blast injury as per self-report or the ambulance personnel were included in the study. Frequency of demographics, injury pattern, and outcomes were calculated. Results A total of 103 patients with bomb blast injuries presented to the selected emergency departments. The median age of patients was 30 years. Around three-fourth of the patients were males (n = 74, 74.7%). Most of the bomb blast patients were seen in Peshawar (n = 41, 39.8%) and Karachi city (n = 31, 30.1%) and the most common mode of arrival was non-ambulance transport (n = 71, 76.3%). Upper limb injuries (n = 12, 40%) were common in the under 18 age group and lower limb injuries (n = 31, 39.2%) in the 18 years and above group. There were a total of 8 (7.7%) deaths reported out of these 103 patients. Conclusion Bomb blast injuries in Pakistan generally affect young males. Non-ambulance transport is the most common way to access emergency departments (ED). Overall ED mortality is high and capturing data during a disaster in an emergency department is challenging. PMID:26692453

  8. Bomb blast injuries: an exploration of patient characteristics and outcome using Pakistan National Emergency Departments Surveillance (Pak-NEDS) data.

    PubMed

    Khan, Irum; Khan, Nadeem; Naeem, Rubaba; Kerai, Salima; Allen, Kate; Zia, Nukhba; Shahbaz, Sana; Afridi, Shiraz; Siddiqui, Emaduddin; Khan, Uzma; Hyder, Adnan A; Razzak, Junaid A

    2015-01-01

    Bomb blast injuries result in premature deaths and burdening of healthcare systems. The objective of this study was to explore the characteristics and outcome of patients presenting to the emergency departments in Pakistan with bomb blast injuries. Active surveillance was conducted in seven major emergency departments of Pakistan from November 2010-March 2011. All the sites are tertiary care urban centers. All the patients who presented to the hospital's emergency department (ED) following a bomb blast injury as per self-report or the ambulance personnel were included in the study. Frequency of demographics, injury pattern, and outcomes were calculated. A total of 103 patients with bomb blast injuries presented to the selected emergency departments. The median age of patients was 30 years. Around three-fourth of the patients were males (n = 74, 74.7%). Most of the bomb blast patients were seen in Peshawar (n = 41, 39.8%) and Karachi city (n = 31, 30.1%) and the most common mode of arrival was non-ambulance transport (n = 71, 76.3%). Upper limb injuries (n = 12, 40%) were common in the under 18 age group and lower limb injuries (n = 31, 39.2%) in the 18 years and above group. There were a total of 8 (7.7%) deaths reported out of these 103 patients. Bomb blast injuries in Pakistan generally affect young males. Non-ambulance transport is the most common way to access emergency departments (ED). Overall ED mortality is high and capturing data during a disaster in an emergency department is challenging.

  9. A School Improvement-Accountability Process Kit. PAK No. 3.2--Developing Educational Programs: Structures--Objectives--Budgets.

    ERIC Educational Resources Information Center

    Villars, Gerald K.

    The purpose of this Personalized Activity Kit is to provide an introduction to the concept of program budgeting by focusing on several key elements to be considered in the development of a program budgeting system. These elements are an examination of the school's organizational patterns in defining and designing the district's educational…

  10. Photocatalytic degradation of acephate in pak choi, Brassica chinensis, with Ce-doped TiO2.

    PubMed

    Liu, Xiangying; Wang, Lifeng; Zhou, Xiaomao; Liu, Kailin; Bai, Lianyang; Zhou, Xuguo

    2015-01-01

    The photocatalytic degradation of acephate was investigated using Ce-doped TiO2 (TiO2/Ce) hydrosol. In contrast to previous research conducted under artificial light in the laboratory, this study investigated the decomposition of acephate in a field trial. The results show that acephate can be efficiently degraded by the TiO2/Ce system under natural field conditions; the degradation efficiency was affected by the dosage of the photocatalyst and acephate. The optimum dosage of TiO2/Ce was 2400 g a.i.ha(-1), and the photodegradation efficiency of acephate reached 93.5% after 20 h at an acephate dosage of 675 g a.i.ha(-1). Ultra-performance liquid chromatography/mass spectrometry (UPLC/MS) analysis detected and identified four degradation products-methamidophos, phosphorothioic acid O,O,S-trimethyl ester, S-methyl methanethiosulfonate and phosphorous acid-that were formed during the TiO2/Ce photodegradation of acephate. Based on the structural identification of the degradation products, a probable photodegradation pathway was proposed, and the first decomposition step may be the cleavage of the C‒N bond of acephate. Subsequently, the P‒S and P‒O bonds may be oxidized gradually or simultaneously to complete the mineralization.

  11. High performance thin layer chromatographic screening for aflatoxins in poultry feed by using silica Sep-Paks

    SciTech Connect

    Kozloski, R.P.

    1986-06-01

    The need for a convenient, sensitive and highly selective method to test for aflatoxins in poultry feed samples led to the development of a method based on the CB procedure. The CB procedure requires large volumes of solvents, the preparation of silica gel of a specified moisture content and the use of large TLC plates, which have long development times. The desirability of running fortified control samples to monitor aflatoxin recovery and to serve as reference standards doubles the already high volume of solvents required. To overcome these difficulties, a number of changes in the CB method were explored. Sample size reduction was used in an attempt to decrease the volume of solvents required. The practicality of using commercially available silica Sep-Paps was investigated. The use of high performance thin layer chromatographic (HPTLC) plates with preadsorbent zones was tested to decrease spotting and development time. The capability of readily detecting aflatoxins B/sub 1/, G/sub 1/, B/sub 2/ and G/sub 2/ at 10, 10, 5 and 5 ppb (10/sup 9/), respectively, was set as the goal.

  12. Group II p21-activated kinases as therapeutic targets in gastrointestinal cancer

    PubMed Central

    Shao, Yang-Guang; Ning, Ke; Li, Feng

    2016-01-01

    P21-activated kinases (PAKs) are central players in various oncogenic signaling pathways. The six PAK family members are classified into group I (PAK1-3) and group II (PAK4-6). Focus is currently shifting from group I PAKs to group II PAKs. Group II PAKs play important roles in many fundamental cellular processes, some of which have particular significance in the development and progression of cancer. Because of their important functions, group II PAKs have become popular potential drug target candidates. However, few group II PAKs inhibitors have been reported, and most do not exhibit satisfactory kinase selectivity and “drug-like” properties. Isoform- and kinase-selective PAK inhibitors remain to be developed. This review describes the biological activities of group II PAKs, the importance of group II PAKs in the development and progression of gastrointestinal cancer, and small-molecule inhibitors of group II PAKs for the treatment of cancer. PMID:26811660

  13. Beilstein Without Tears: Education in the Use of the Literature of Organic Chemistry.

    ERIC Educational Resources Information Center

    Callaghan, Patricia M.; And Others

    1986-01-01

    The use of Beilstein ("Handbuch der Organischen Chemie") in the early stages of a second-year, one semester course in organic chemistry is described. Student literature projects, evaluation, use of ancillary literature, and a sample search are included. (JN)

  14. Group I p21-activated kinases regulate thyroid cancer cell migration and are overexpressed and activated in thyroid cancer invasion.

    PubMed

    McCarty, Samantha K; Saji, Motoyasu; Zhang, Xiaoli; Jarjoura, David; Fusco, Alfredo; Vasko, Vasyl V; Ringel, Matthew D

    2010-12-01

    p21-activated kinases (PAKs) are a family of serine/threonine kinases that regulate cytoskeletal dynamics and cell motility. PAKs are subdivided into group I (PAKs 1-3) and group II (PAKs 4-6) on the basis of structural and functional characteristics. Based on prior gene expression data that predicted enhanced PAK signaling in the invasive fronts of aggressive papillary thyroid cancers (PTCs), we hypothesized that PAKs functionally regulate thyroid cancer cell motility and are activated in PTC invasive fronts. We examined PAK isoform expression in six human thyroid cancer cell lines (BCPAP, KTC1, TPC1, FTC133, C643, and SW1746) by quantitative reverse transcription-PCR and western blot. All cell lines expressed PAKs 1-4 and PAK6 mRNA and PAKs 1-4 protein; PAK6 protein was variably expressed. Samples from normal and malignant thyroid tissues also expressed PAKs 1-4 and PAK6 mRNA; transfection with the group I (PAKs 1-3) PAK-specific p21 inhibitory domain molecular inhibitor reduced transwell filter migration by ∼50% without altering viability in all cell lines (P<0.05). BCPAP and FTC133 cells were transfected with PAK1, PAK2, or PAK3-specific small interfering RNA (siRNA); only PAK1 siRNA reduced migration significantly for both cell lines. Immunohistochemical analysis of seven invasive PTCs demonstrated an increase in PAK1 and pPAK immunoactivity in the invasive fronts versus the tumor center. In conclusion, PAK isoforms are expressed in human thyroid tissues and cell lines. PAK1 regulates thyroid cancer cell motility, and PAK1 and pPAK levels are increased in PTC invasive fronts. These data implicate PAKs as regulators of thyroid cancer invasion.

  15. Analysis of p21-Activated Kinase Function in Neurofibromatosis Type 2

    DTIC Science & Technology

    2009-01-01

    SchΔ(39-121) Pak1+/+ mice developed NF2-related pathologies (schwannomatosis, nerve sheath tumors, sarcomas ), whereas 2/34 P0- SchΔ(39-121) Pak1...Pak3 and Pak5 HEK293 cells were transfected with 0.5 mg Pak3 or myc-Pak5 DNA using lipo - some-mediated transfection (Lipofectamine 2000, Invitrogen

  16. Signaling, Regulation, and Specificity of the Type II p21-activated Kinases*

    PubMed Central

    Ha, Byung Hak; Morse, Elizabeth M.; Turk, Benjamin E.; Boggon, Titus J.

    2015-01-01

    The p21-activated kinases (PAKs) are a family of six serine/threonine kinases that act as key effectors of RHO family GTPases in mammalian cells. PAKs are subdivided into two groups: type I PAKs (PAK1, PAK2, and PAK3) and type II PAKs (PAK4, PAK5, and PAK6). Although these groups are involved in common signaling pathways, recent work indicates that the two groups have distinct modes of regulation and have both unique and common substrates. Here, we review recent insights into the molecular level details that govern regulation of type II PAK signaling. We also consider mechanisms by which signal transduction is regulated at the level of substrate specificity. Finally, we discuss the implications of these studies for clinical targeting of these kinases. PMID:25855792

  17. Multidrug resistant gram-negative bacteria in clinical isolates from Karachi.

    PubMed

    Saeed, Asma; Khatoon, Hajra; Ansari, Fasihuddin Ahmed

    2009-01-01

    A total of 54 gram-negative bacteria obtained from various pathological labs and hospitals of Karachi were screened for their resistance to ampicillin, chloramphenicol, gentamycin, kanamycin, neomycin, streptomycin and tetracycline antibiotics. Of the 54 bacteria, 50 were resistant to one or more antibiotics. Among the resistant bacteria, 13 out of 28 were found to transfer their resistances by conjugation. This indicates that at least 46% of clinical gram-negative bacteria in Karachi possess various types of transferable R plasmids, such as pAK5, pAK9, pAK10, pAK11, pAK12, pAK13, pAK14, pAK15, pAK16, pAK17, pAK18, pAK19, pAK20 and pAK21. The non-conjugative R plasmids included pMT14 and pZ26. Only pAK15 showed 26% segregation even after 20 consecutive transfers in plain broth (spontaneous segregation) whereas only pAK15 and pAK16 showed any significant loss of their markers in curing by acridine orange. The stability of R plasmids is more dangerous from clinical point of view.

  18. Myungshin Im, Won-Kee Park, Changsu Choi, Yiseul Jeon, Ji-Hoon Kim (CEOU/SNU), Giseon Baek, Young-Seok Oh, Soojong Pak (Kyunghee Univ.)

    NASA Astrophysics Data System (ADS)

    Im, Myungshin; Park, Won-Kee; Choi, Changsu; Jeon, Yiseul; Kim, Ji-Hoon; Baek, Giseon; Oh, Young-Seok; Pak, Soojong

    2011-08-01

    On August 30 02:05 UT, we started observations of SN2011fe=PTF11kly (ATel#3581) in grizY and is/iz-bands using CQUEAN on the 2.1m telescope at McDonald observatory, and in zYJHK-bands using UKIRT. SN2011fe is found to have J~11.58 and K~11.55 mag (error~0.05 mag), still brightening in NIR with respect to ATel#3605.

  19. The Botrytis cinerea PAK kinase BcCla4 mediates morphogenesis, growth and cell cycle regulating processes downstream of BcRac.

    PubMed

    Minz-Dub, Anna; Sharon, Amir

    2017-02-06

    Rac proteins are involved in a variety of cellular processes. Effector proteins that interact with active Rac convey the GTPase-generated signal to downstream developmental cascades and processes. Here we report on the analysis of the main effector and signal cascade downstream of BcRac, the Rac homolog of the grey mold fungus Botrytis cinerea. Several lines of evidence highlighted the p21-activated kinase Cla4 as an important effector of Rac in fungi. Analysis of Δbccla4 strains revealed that the BcCla4 protein was sufficient to mediate all of the examined BcRac-driven processes, including hyphal growth and morphogenesis, conidia production and pathogenicity. In addition, the Δbccla4 strains had altered nuclei content, a phenomenon that was previously observed in Δbcrac isolates, thus connecting the BcRac/BcCla4 module with cell cycle control. Further analyses revealed that BcRac/BcCla4 control mitotic entry through changes in phosphorylation status of the cyclin dependent kinase BcCdk1. The complete cascade includes the kinase BcWee1, which is downstream of BcCla4 and upstream of BcCdk1. These results provide a mechanistic insight on the connection of cell cycle, morphogenesis and pathogenicity in fungi, and position BcCla4 as the most essential effector and central regulator of all of these processes downstream of BcRac.

  20. Small molecules that allosterically inhibit p21-activated kinase activity by binding to the regulatory p21-binding domain.

    PubMed

    Kim, Duk-Joong; Choi, Chang-Ki; Lee, Chan-Soo; Park, Mee-Hee; Tian, Xizhe; Kim, Nam Doo; Lee, Kee-In; Choi, Joong-Kwon; Ahn, Jin Hee; Shin, Eun-Young; Shin, Injae; Kim, Eung-Gook

    2016-04-29

    p21-activated kinases (PAKs) are key regulators of actin dynamics, cell proliferation and cell survival. Deregulation of PAK activity contributes to the pathogenesis of various human diseases, including cancer and neurological disorders. Using an ELISA-based screening protocol, we identified naphtho(hydro)quinone-based small molecules that allosterically inhibit PAK activity. These molecules interfere with the interactions between the p21-binding domain (PBD) of PAK1 and Rho GTPases by binding to the PBD. Importantly, they inhibit the activity of full-length PAKs and are selective for PAK1 and PAK3 in vitro and in living cells. These compounds may potentially be useful for determining the details of the PAK signaling pathway and may also be used as lead molecules in the development of more selective and potent PAK inhibitors.

  1. p21-activated kinase 2 regulates HSPC cytoskeleton, migration, and homing via CDC42 activation and interaction with β-Pix

    PubMed Central

    Reddy, Pavankumar N. G.; Radu, Maria; Xu, Ke; Wood, Jenna; Harris, Chad E.; Chernoff, Jonathan

    2016-01-01

    Cytoskeletal remodeling of hematopoietic stem and progenitor cells (HSPCs) is essential for homing to the bone marrow (BM). The Ras-related C3 botulinum toxin substrate (Rac)/cell division control protein 42 homolog (CDC42) effector p21-activated kinase (Pak2) has been implicated in HSPC homing and engraftment. However, the molecular pathways mediating Pak2 functions in HSPCs are unknown. Here, we demonstrate that both Pak2 kinase activity and its interaction with the PAK-interacting exchange factor-β (β-Pix) are required to reconstitute defective Pak2Δ/Δ HSPC homing to the BM. Pak2 serine/threonine kinase activity is required for stromal-derived factor-1 (SDF1α) chemokine-induced HSPC directional migration, whereas Pak2 interaction with β-Pix is required to regulate the velocity of HSPC migration and precise F-actin assembly. Lack of SDF1α-induced filopodia and associated abnormal cell protrusions seen in Pak2Δ/Δ HSPCs were rescued by wild-type (WT) Pak2 but not by a Pak2-kinase dead mutant (KD). Expression of a β-Pix interaction-defective mutant of Pak2 rescued filopodia formation but led to abnormal F-actin bundles. Although CDC42 has previously been considered an upstream regulator of Pak2, we found a paradoxical decrease in baseline activation of CDC42 in Pak2Δ/Δ HSPCs, which was rescued by expression of Pak2-WT but not by Pak2-KD; defective homing of Pak2-deleted HSPCs was rescued by constitutive active CDC42. These data demonstrate that both Pak2 kinase activity and its interaction with β-Pix are essential for HSPC filopodia formation, cytoskeletal integrity, and homing via activation of CDC42. Taken together, we provide mechanistic insights into the role of Pak2 in HSPC migration and homing. PMID:26932803

  2. Heat sinking for printed circuitry

    DOEpatents

    Wilson, S.K.; Richardson, G.; Pinkerton, A.L.

    1984-09-11

    A flat pak or other solid-state device mounted on a printed circuit board directly over a hole extends therethrough so that the bottom of the pak or device extends beyond the bottom of the circuit board. A heat sink disposed beneath the circuit board contacts the bottom of the pak or device and provides direct heat sinking thereto. Pressure may be applied to the top of the pak or device to assure good mechanical and thermal contact with the heat sink.

  3. Caveolin-1 Modulates Androgen Receptor Signaling in Advanced Prostate Cancer

    DTIC Science & Technology

    2005-02-01

    that additional priming events are needed. Alternatively, center of the activation loop within the catalytic domain. Crys- the TPY motif in the group...proline-tyrosine, TPY ) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was also blocked by...mediated activation. PAK4 and PAK5 were similarly activated by MKK6, consistent with a conserved TPY motif in their activation domains. The activation of

  4. Redundant Canonical and Noncanonical Caenorhabditis elegans p21-Activated Kinase Signaling Governs Distal Tip Cell Migrations

    PubMed Central

    Peters, Eldon C.; Gossett, Andrea J.; Goldstein, Bob; Der, Channing J.; Reiner, David J.

    2013-01-01

    p21-activated kinases (Paks) are prominent mediators of Rac/Cdc42-dependent and -independent signaling and regulate signal transduction and cytoskeletal-based cell movements. We used the reproducible migrations of the Caenorhabditis elegans gonadal distal tip cells to show that two of the three nematode Pak proteins, MAX-2 and PAK-1, function redundantly in regulation of cell migration but are regulated by very different mechanisms. First, we suggest that MAX-2 requires CED-10/Rac function and thus functions canonically. Second, PIX-1 and GIT-1 function in the same role as PAK-1, and PAK-1 interaction with PIX-1 is required for PAK-1 activity; thus, PAK-1 functions noncanonically. The human Pak-Pix-Git complex is central to noncanonical Pak signaling and requires only modest Rac/CDC-42 input. Unlike the human complex, our results suggest that the C. elegans Pak-Pix-Git complex requires PAK-1 kinase domain activity. This study delineates signaling network relationships in this cell migration model, thus providing potential further mechanistic insights and an assessment of total Pak contribution to cell migration events. PMID:23390595

  5. Inhibition of group 1 p21-activated kinases suppresses pancreatic stellate cell activation and increases survival of mice with pancreatic cancer.

    PubMed

    Yeo, Dannel; Phillips, Phoebe; Baldwin, Graham S; He, Hong; Nikfarjam, Mehrdad

    2017-05-01

    Pancreatic cancer remains one of the most lethal of all solid tumors. Pancreatic stellate cells (PSCs) are primarily responsible for the fibrosis that constitutes the stroma and p21-activated kinase 1 (PAK1) may have a role in signalling pathways involving PSCs. This study aimed to examine the role of PAK1 in PSCs and in the interaction of PSCs with pancreatic cancer cells. Human PSCs were isolated using the modified outgrowth method. The effect of inhibiting PAK1 with group 1 PAK inhibitor, FRAX597, on cell proliferation and apoptosis in vitro was measured by thymidine incorporation and annexin V assays, respectively. The effect of depleting host PAK1 on the survival of mice with pancreatic Pan02 cell tumors was evaluated using PAK1 knockout (KO) mice. PAK1 was expressed in isolated PSCs. FRAX597 reduced the activation of PSCs, inhibited PSC proliferation, and increased PSC apoptosis at least in partial by inhibiting PAK1 activity. The decreased expression and activity of PAK1 in PAK1 KO mice tumors was associated with an increased mouse survival. These results implicate PAK1 as a regulator of PSC activation, proliferation and apoptosis. Targeting stromal PAK1 could increase therapeutic response and survival of patients with pancreatic cancer.

  6. Sorting Nexin 27 Protein Regulates Trafficking of a p21-activated Kinase (PAK) Interacting Exchange Factor (β-Pix)-G Protein-coupled Receptor Kinase Interacting Protein (GIT) Complex via a PDZ Domain Interaction*

    PubMed Central

    Valdes, Julie L.; Tang, Jingrong; McDermott, Mark I.; Kuo, Jean-Cheng; Zimmerman, Seth P.; Wincovitch, Stephen M.; Waterman, Clare M.; Milgram, Sharon L.; Playford, Martin P.

    2011-01-01

    Sorting nexin 27 (SNX27) is a 62-kDa protein localized to early endosomes and known to regulate the intracellular trafficking of ion channels and receptors. In addition to a PX domain, SNX27 is the only sorting family member that contains a PDZ domain. To identify novel SNX27-PDZ binding partners, we performed a proteomic screen in mouse principal kidney cortical collecting duct cells using a GST-SNX27 fusion construct as bait. We found that β-Pix (p21-activated kinase-interactive exchange factor), a guanine nucleotide exchange factor for the Rho family of small GTPases known to regulate cell motility directly interacted with SNX27. The association of β-Pix and SNX27 is specific for β-Pix isoforms terminating in the type-1 PDZ binding motif (ETNL). In the same screen we also identified Git1/2 as a potential SNX27 interacting protein. The interaction between SNX27 and Git1/2 is indirect and mediated by β-Pix. Furthermore, we show recruitment of the β-Pix·Git complex to endosomal sites in a SNX27-dependent manner. Finally, migration assays revealed that depletion of SNX27 from HeLa and mouse principal kidney cortical collecting duct cells significantly decreases cell motility. We propose a model by which SNX27 regulates trafficking of β-Pix to focal adhesions and thereby influences cell motility. PMID:21926430

  7. Sorting nexin 27 protein regulates trafficking of a p21-activated kinase (PAK) interacting exchange factor (β-Pix)-G protein-coupled receptor kinase interacting protein (GIT) complex via a PDZ domain interaction.

    PubMed

    Valdes, Julie L; Tang, Jingrong; McDermott, Mark I; Kuo, Jean-Cheng; Zimmerman, Seth P; Wincovitch, Stephen M; Waterman, Clare M; Milgram, Sharon L; Playford, Martin P

    2011-11-11

    Sorting nexin 27 (SNX27) is a 62-kDa protein localized to early endosomes and known to regulate the intracellular trafficking of ion channels and receptors. In addition to a PX domain, SNX27 is the only sorting family member that contains a PDZ domain. To identify novel SNX27-PDZ binding partners, we performed a proteomic screen in mouse principal kidney cortical collecting duct cells using a GST-SNX27 fusion construct as bait. We found that β-Pix (p21-activated kinase-interactive exchange factor), a guanine nucleotide exchange factor for the Rho family of small GTPases known to regulate cell motility directly interacted with SNX27. The association of β-Pix and SNX27 is specific for β-Pix isoforms terminating in the type-1 PDZ binding motif (ETNL). In the same screen we also identified Git1/2 as a potential SNX27 interacting protein. The interaction between SNX27 and Git1/2 is indirect and mediated by β-Pix. Furthermore, we show recruitment of the β-Pix·Git complex to endosomal sites in a SNX27-dependent manner. Finally, migration assays revealed that depletion of SNX27 from HeLa and mouse principal kidney cortical collecting duct cells significantly decreases cell motility. We propose a model by which SNX27 regulates trafficking of β-Pix to focal adhesions and thereby influences cell motility.

  8. Interdisciplinary Educational Collaborations: Chemistry and Computer Science 967 Ronald S. Haines, Daniel T. Woo, Benjamin T. Hudson, Joji C. Mori, Evey S. M. Ngan, and Wing-Yee Pak

    NASA Astrophysics Data System (ADS)

    Goedhart, Martin J.

    2007-06-01

    While chemists are usually aware of the possibilities of interdisciplinary collaboration in chemical research they may be less aware of the possibilities of such collaboration in education. This article documents an ongoing collaboration between a chemist and computer scientist to co-supervise computer science students engaged in developing software for chemical education, highlights the benefits to both the chemistry and computer science students, notes some unexpected outcomes, and provides guidance to those planning such collaborations. The experiences described in this work should motivate chemistry educators to approach their colleagues in other disciplines with proposals for joint research projects. The collaboration described here initially resulted in the development of student-friendly software for operating a spectrophotometer. Recent co-supervised students have begun developing other software for chemical education.

  9. Survival Escherichia coli O157:H7 transformed with either the pAK1-lux or pXEN-13 plasmids in in vitro bovine ruminal and fecal microbial fermentations

    USDA-ARS?s Scientific Manuscript database

    The use of luminescent technology may serve as a viable model for the real-time validation of various pre-harvest interventions on the colonization or shedding of Escherichia coli O157:H7 within cattle. The objective of this study was to determine if the growth of E. coli O157:H7 (ATCC 43888) in ru...

  10. Regulation of macropinocytosis by p21-activated kinase-1.

    PubMed

    Dharmawardhane, S; Schürmann, A; Sells, M A; Chernoff, J; Schmid, S L; Bokoch, G M

    2000-10-01

    The process of macropinocytosis is an essential aspect of normal cell function, contributing to both growth and motile processes of cells. p21-activated kinases (PAKs) are targets for activated Rac and Cdc42 guanosine 5'-triphosphatases and have been shown to regulate the actin-myosin cytoskeleton. In fibroblasts PAK1 localizes to areas of membrane ruffling, as well as to amiloride-sensitive pinocytic vesicles. Expression of a PAK1 kinase autoinhibitory domain blocked both platelet-derived growth factor- and RacQ61L-stimulated uptake of 70-kDa dextran particles, whereas an inactive version of this domain did not, indicating that PAK kinase activity is required for normal growth factor-induced macropinocytosis. The mechanisms by which PAK modulate macropinocytosis were examined in NIH3T3 cell lines expressing various PAK1 constructs under the control of a tetracycline-responsive transactivator. Cells expressing PAK1 (H83,86L), a mutant that dramatically stimulates formation of dorsal membrane ruffles, exhibited increased macropinocytic uptake of 70-kDa dextran particles in the absence of additional stimulation. This effect was not antagonized by coexpression of dominant-negative Rac1-T17N. In the presence of platelet-derived growth factor, both PAK1 (H83,86L) and a highly kinase active PAK1 (T423E) mutant dramatically enhanced the uptake of 70-kDa dextran. Neither wild-type PAK1 nor vector controls exhibited enhanced macropinocytosis, nor did PAK1 (H83,86L) affect clathrin-dependent endocytic mechanisms. Active versions of PAK1 enhanced both growth factor-stimulated 70-kDa dextran uptake and efflux, suggesting that PAK1 activity modulated pinocytic vesicle cycling. These data indicate that PAK1 plays an important regulatory role in the process of macropinocytosis, perhaps related to the requirement for PAK in directed cell motility.

  11. The role of p21-activated kinases in hepatocellular carcinoma metastasis

    PubMed Central

    2014-01-01

    The p21-activated kinases (PAKs) are downstream effectors of the Rho family small GTPases as well as a wide variety of mitogenic factors and have been implicated in cancer formation, development and metastasis. PAKs phosphorylate a wide spectrum of substrates to mediate extracellular signals and regulate cytoskeletal remodeling, cell motility and survival. In this review, we aim to summarize the findings regarding the oncogenic role and the underlying mechanisms of PAKs signaling in various cancers, and in particular highlight the prime importance of PAKs in hepatocellular carcinoma (HCC) progression and metastasis. Recent studies exploring the potential therapeutic application of PAK inhibitors will also be discussed. PMID:25093037

  12. New system enables fleets to tailor maintenance programs

    SciTech Connect

    Fiscor, S.

    2007-08-15

    The Lubrizol Corporation has developed FluiPak and FluiPak Lite systems to pull engine oils and sample them continuously, helping to maintain mining equipment between oil change intervals. The FluiPak system has a control box, a sensor box and a reserve system. FluiPak Lite is simply a cut-down version of FluiPak. The article explains how the system works to detect in real time oil quality and contaminants and hence optimize drain intervals and also prolong engine component life and reduce waste oil disposal costs. 100 units are currently installed in coal operations. 5 figs., 5 photos.

  13. The P21-activated kinase expression pattern is different in non-small cell lung cancer and affects lung cancer cell sensitivity to epidermal growth factor receptor tyrosine kinase inhibitors.

    PubMed

    Liu, Yang; Wang, Si; Dong, Qian-Ze; Jiang, Gui-Yang; Han, Yong; Wang, Liang; Wang, En-Hua

    2016-03-01

    Exploring methods for increasing epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) sensitivity has become a major focus in non-small cell lung cancer (NSCLC). Major downstream effectors of the Rho family small guanosine triphosphatases, P21-activated kinases (PAKs) activate the main signaling pathways downstream of EGFR and thus promote tumor cell proliferation. In this study, we explored the expression pattern of phosphorylated PAKs in NSCLC and their potential value as drug targets for treating cancer. The expression and prognostic significance of phosphorylated group I and II PAKs were evaluated in 182 patients with NSCLC. Immunohistochemical analysis revealed low group I PAK expression in normal lung tissues and increased expressed in the cytoplasm, particularly in lung squamous cell carcinoma. Abnormal group I PAK expression was associated with lymph node metastases and high tumor-node-metastases (TNM) stage in NSCLC patients and correlated with poor prognosis. We used group I PAK inhibitor (IPA3) to specifically decrease group I PAK activity in human lung cancer cell lines. Decreased group I PAK activity inhibited cell proliferation and combined IPA3 and EGFR-TKI (gefitinib) treatment inhibited cell proliferation in an obvious manner. Together, our results revealed the PAK expression pattern in NSCLC, and a role for group I PAK in cell proliferation, which provides evidence that decreased PAK activity may have a potential application as a molecular targeted therapy in advanced NSCLC.

  14. P21 activated kinases

    PubMed Central

    Rane, Chetan K; Minden, Audrey

    2014-01-01

    The p21 activated kinases (Paks) are well known effector proteins for the Rho GTPases Cdc42 and Rac. The Paks contain 6 members, which fall into 2 families of proteins. The first family consists of Paks 1, 2, and 3, and the second consists of Paks 4, 5, and 6. While some of the Paks are ubiquitously expressed, others have more restrictive tissue specificity. All of them are found in the nervous system. Studies using cell culture, transgenic mice, and knockout mice, have revealed important roles for the Paks in cytoskeletal organization and in many aspects of cell growth and development. This review discusses the basic structures of the Paks, and their roles in cell growth, development, and in cancer. PMID:24658305

  15. p21-activated kinase 2 regulates HSPC cytoskeleton, migration, and homing via CDC42 activation and interaction with β-Pix.

    PubMed

    Reddy, Pavankumar N G; Radu, Maria; Xu, Ke; Wood, Jenna; Harris, Chad E; Chernoff, Jonathan; Williams, David A

    2016-04-21

    Cytoskeletal remodeling of hematopoietic stem and progenitor cells (HSPCs) is essential for homing to the bone marrow (BM). The Ras-related C3 botulinum toxin substrate (Rac)/cell division control protein 42 homolog (CDC42) effector p21-activated kinase (Pak2) has been implicated in HSPC homing and engraftment. However, the molecular pathways mediating Pak2 functions in HSPCs are unknown. Here, we demonstrate that both Pak2 kinase activity and its interaction with the PAK-interacting exchange factor-β (β-Pix) are required to reconstitute defective ITALIC! Pak2 (ITALIC! Δ/Δ)HSPC homing to the BM. Pak2 serine/threonine kinase activity is required for stromal-derived factor-1 (SDF1α) chemokine-induced HSPC directional migration, whereas Pak2 interaction with β-Pix is required to regulate the velocity of HSPC migration and precise F-actin assembly. Lack of SDF1α-induced filopodia and associated abnormal cell protrusions seen in ITALIC! Pak2 (ITALIC! Δ/Δ)HSPCs were rescued by wild-type (WT) Pak2 but not by a Pak2-kinase dead mutant (KD). Expression of a β-Pix interaction-defective mutant of Pak2 rescued filopodia formation but led to abnormal F-actin bundles. Although CDC42 has previously been considered an upstream regulator of Pak2, we found a paradoxical decrease in baseline activation of CDC42 in ITALIC! Pak2 (ITALIC! Δ/Δ)HSPCs, which was rescued by expression of Pak2-WT but not by Pak2-KD; defective homing of ITALIC! Pak2-deleted HSPCs was rescued by constitutive active CDC42. These data demonstrate that both Pak2 kinase activity and its interaction with β-Pix are essential for HSPC filopodia formation, cytoskeletal integrity, and homing via activation of CDC42. Taken together, we provide mechanistic insights into the role of Pak2 in HSPC migration and homing. © 2016 by The American Society of Hematology.

  16. p21-activated kinase group II small compound inhibitor GNE-2861 perturbs estrogen receptor alpha signaling and restores tamoxifen-sensitivity in breast cancer cells

    PubMed Central

    Li, Zhilun; Lorent, Julie; Zhao, Chunyan; Dahlman-Wright, Karin; Strömblad, Staffan

    2015-01-01

    Estrogen receptor alpha (ERα) is highly expressed in most breast cancers. Consequently, ERα modulators, such as tamoxifen, are successful in breast cancer treatment, although tamoxifen resistance is commonly observed. While tamoxifen resistance may be caused by altered ERα signaling, the molecular mechanisms regulating ERα signaling and tamoxifen resistance are not entirely clear. Here, we found that PAK4 expression was consistently correlated to poor patient outcome in endocrine treated and tamoxifen-only treated breast cancer patients. Importantly, while PAK4 overexpression promoted tamoxifen resistance in MCF-7 human breast cancer cells, pharmacological treatment with a group II PAK (PAK4, 5, 6) inhibitor, GNE-2861, sensitized tamoxifen resistant MCF-7/LCC2 breast cancer cells to tamoxifen. Mechanistically, we identified a regulatory positive feedback loop, where ERα bound to the PAK4 gene, thereby promoting PAK4 expression, while PAK4 in turn stabilized the ERα protein, activated ERα transcriptional activity and ERα target gene expression. Further, PAK4 phosphorylated ERα-Ser305, a phosphorylation event needed for the PAK4 activation of ERα-dependent transcription. In conclusion, PAK4 may be a suitable target for perturbing ERα signaling and tamoxifen resistance in breast cancer patients. PMID:26554417

  17. Identification of Novel Gene Targets and Functions of p21-Activated Kinase 1 during DNA Damage by Gene Expression Profiling

    PubMed Central

    Motwani, Mona; Li, Da-Qiang; Horvath, Anelia; Kumar, Rakesh

    2013-01-01

    P21-activated kinase 1 (PAK1), a serine/threonine protein kinase, modulates many cellular processes by phosphorylating its downstream substrates. In addition to its role in the cytoplasm, PAK1 also affects gene transcription due to its nuclear localization and association with chromatin. It is now recognized that PAK1 kinase activity and its nuclear translocation are rapidly stimulated by ionizing radiation (IR), and that PAK1 activation is a component of the DNA damage response. Owing to the role of PAK1 in the cell survival, its association with the chromatin, and now, stimulation by ionizing radiation, we hypothesize that PAK1 may be contributing to modulation of genes with roles in cellular processes that might be important in the DNA damage response. The purpose of this study was to identify new PAK1 targets in response to ionizing radiation with putative role in the DNA damage response. We examined the effect of IR on the gene expression patterns in the murine embryonic fibroblasts with or without Pak1 using microarray technology. Differentially expressed transcripts were identified using Gene Spring GX 10.0.2. Pathway, network, functional analyses and gene family classification were carried out using Kyoto Encyclopedia of Genes and Genomes (KEGG), Ingenuity Pathway, Gene Ontology and PANTHER respectively. Selective targets of PAK1 were validated by RT-qPCR. For the first time, we provide a genome-wide analysis of PAK1 and identify its targets with potential roles in the DNA damage response. Gene Ontology analysis identified genes in the IR-stimulated cells that were involved in cell cycle arrest and cell death. Pathway analysis revealed p53 pathway being most influenced by IR responsive, PAK1 targets. Gene family of transcription factors was over represented and gene networks involved in DNA replication, repair and cellular signaling were identified. In brief, this study identifies novel PAK1 dependent IR responsive genes which reveal new aspects of PAK1

  18. Chemie

    NASA Astrophysics Data System (ADS)

    Kurzweil, Peter

    Das Kapitel "Chemie″ gibt eine kompakte Übersicht über die für den Maschinebau notwendigen chemischen Grundlagen und ihre technischen Anwendungen. Auf Atombau, Radioaktivität und Periodensystem folgen chemische Bindung, Stöchiometrie und chemische Reaktionen, Säuren und Basen, Fällungsreaktionen und Elektrochemie, bis hin zu den organischen Stoffklassen. Praktische Hinweise beleuchten die chemische Thermodynamik und Katalyse, Wasserhärte und Wasseraufbereitung, Luftverunreinigungen und Bauchemie, bis hin zu Batterien und Galvanotechnik.

  19. Coordinating structural and functional synapse development: postsynaptic p21-activated kinase independently specifies glutamate receptor abundance and postsynaptic morphology.

    PubMed

    Albin, Stephanie D; Davis, Graeme W

    2004-08-04

    Here, we show that postsynaptic p21-activated kinase (Pak) signaling diverges into two genetically separable pathways at the Drosophila neuromuscular junction. One pathway controls glutamate receptor abundance. Pak signaling within this pathway is specified by a required interaction with the adaptor protein Dreadlocks (Dock). We demonstrate that Dock is localized to the synapse via an Src homology 2-mediated protein interaction. Dock is not necessary for Pak localization but is necessary to restrict Pak signaling to control glutamate receptor abundance. A second genetically separable function of Pak kinase signaling controls muscle membrane specialization through the regulation of synaptic Discs-large. In this pathway, Dock is dispensable. We present a model in which divergent Pak signaling is able to coordinate two different features of postsynaptic maturation, receptor abundance, and muscle membrane specialization.

  20. Activation of cAMP signaling attenuates impaired hepatic glucose disposal in aged male p21-activated protein kinase-1 knockout mice.

    PubMed

    Chiang, Yu-Ting Alex; Ip, Wilfred; Shao, Weijuan; Song, Zhuolun Eric; Chernoff, Jonathan; Jin, Tianru

    2014-06-01

    p21-activated protein kinase-1 (Pak1) plays a role in insulin secretion and glucagon-like peptide-1 (GLP-1) production. Pak1(-/-) mice were found to carry a defect in ip pyruvate tolerance test (IPPTT), leading us to speculate whether Pak1 represses hepatic gluconeogenesis. We show here that the defect in IPPTT became more severe in aged Pak1(-/-) mice. In primary hepatocytes, 2,2'-dihydroxy-1,1'-dinaphthyldisulfide, a potent inhibitor of group I Paks, reduced basal glucose production (GP), attenuated forskolin- or glucagon-stimulated GP, and attenuated the stimulation of forskolin on the expression of Pck1 and G6pc. In addition, the capacity of primary hepatocytes isolated from Pak1(-/-) mice in GP at the basal level is significantly lower than that of the control littermates. These in vitro observations imply that the direct effect of Paks in hepatocytes is the stimulation of gluconeogenesis and that the impairment in IPPTT in Pak1(-/-) mice is due to the lack of Pak1 elsewhere. Consecutive ip injection of forskolin for 2 weeks increased gut proglucagon expression, associated with improved IPPTT in aged Pak1(-/-) mice and wild-type controls. In addition, administration of the DPP-IV (dipeptidyl peptidase-4) inhibitor sitagliptin for 1 week reversed the defect in IPPTT in aged Pak1(-/-) mice, associated with increased plasma GLP-1 levels. Our observations indicate a potential role of Pak1 in the gut/pancreas/liver axis in controlling glucose disposal and affirmed the therapeutic application of GLP-1 and DPP-IV inhibitors in attenuating hepatic gluconeogenesis.

  1. Chinese Policy Toward South Asia: Implications and Prospects for Nepal

    DTIC Science & Technology

    2013-03-01

    India, Pakistan, Nepal and Bhutan), while the other three ( Bangladesh , Sri Lanka and Maldives) do not share borders with China (Map 1). Nepal and...India; contingent cooperation with Pakistan, Bangladesh and Sri Lanka; and secretive cooptation with Nepal and Maldives.16 Pursuing this strategy at...Pak War of 1965, China sided with Pakistan. Again during the Indo-Pak War in 1971, which led to the birth of Bangladesh , Sino-Pak cooperation was

  2. Identifying Breast Cancer Oncogenes

    DTIC Science & Technology

    2011-10-01

    which is a study of 3131 human tumor samples and cancer cell lines including 243 breast samples. Tumorscape showed that PAK1 is located in an...chromosome 11q of human tumor samples and cancer cell lines that exhibit highest level of PAK1 amplification divided according to cancer type...breast, non-small cell (NSC) lung, ovarian (Ov), small cell lung (SCL), melanoma (Mel) and esophageal squamous (Esq). PAK1 and CCND1 1oci are marked . B

  3. Transcriptional regulation of ataxia-telangiectasia and Rad3-related protein by activated p21-activated kinase-1 protects keratinocytes in UV-B-induced premalignant skin lesions.

    PubMed

    Beesetti, S; Mavuluri, J; Surabhi, R P; Oberyszyn, T M; Tober, K; Pitani, R S; Joseph, L D; Venkatraman, G; Rayala, S K

    2017-07-10

    Sun-induced skin lesions, in particular actinic keratosis, are generally considered as premalignant skin lesions that can progress into squamous cell carcinoma (SCC) and invasive SCC if left untreated. Therefore, understanding the molecular mechanisms by which the ultraviolet-B (UV-B)-exposed cells are being protected and the signaling pathways that promote the progression of certain premalignant skin lesions to malignant lesions will permit us to prevent or cure skin cancers. In the current study, we found that phospho-p21-activated kinase-1 (Pak1) and Pak1 expression was high in clinical samples of sunlight-induced premalignant skin lesions assessed by immunohistochemistry. Further, we observed that phospho-Pak1 and Pak1 levels are high in UV-B-exposed hairless SKH mouse model skin samples as compared with unexposed skin tissue. Our results from cell line and animal models showed that Pak1 is activated in response to UV-B radiation, and this activated Pak1 translocates from the cytoplasm to the nucleus. Inside the nucleus, Pak1 via C-Fos binds to a specific promoter region of DNA repair kinase ATR (ataxia-telangiectasia and Rad3-related protein) and acts as a transcriptional regulator of ATR. Results from our analysis showed that Pak1 overexpression, knockdown and Pak1 knockout cell line models showed that Pak1 confers protection to keratinocytes from UV-B-induced apoptosis and DNA damage via ATR. To our knowledge, this is the first study that evaluates the functional and clinical significance of a signaling molecule, Pak1, in sun-induced premalignant skin lesions and indicates that increased Pak1 activation and expression could serve as an early warning sign of progression toward non-melanoma skin cancer, if ignored.Oncogene advance online publication, 10 July 2017; doi:10.1038/onc.2017.218.

  4. [Nature of cancer explored from the perspective of the functional evolution of proto-oncogenes].

    PubMed

    Watari, Akihiro

    2012-01-01

    The products of proto-oncogene play critical roles in the development or maintenance of multicellular societies in animals via strict regulatory systems. When these regulatory systems are disrupted, proto-oncogenes can become oncogenes, and thereby induce cell transformation and carcinogenesis. To understand the molecular basis for development of the regulatory system of proto-oncogenes during evolution, we screened for ancestral proto-oncogenes from the unicellular choanoflagellate Monosiga ovata (M. ovata) by monitoring their transforming ability in mammalian cells; consequently, we isolated a Pak gene ortholog, which encodes a serine/threonine kinase as a 'primitive oncogene'. We also cloned Pak orthologs from fungi and the multicellular sponge Ephydatia fluviatilis, and compared their regulatory features with that of M. ovata Pak (MoPak). MoPak is constitutively active and induces cell transformation in mammalian cells. In contrast, Pak orthologs from multicellular animals are strictly regulated. Analyses of Pak mutants revealed that structural alterations in the auto-inhibitory domain (AID) are responsible for the enhanced kinase activity and the oncogenic activity of MoPak. Furthermore, we show that Rho family GTPases-mediated regulatory system of Pak kinase is conserved throughout the evolution from unicellular to multicellular animals, but the MoPak is more sensitive to the Rho family GTPases-mediated activation than multicellular Pak. These results show that maturation of AID function was required for the development of the strict regulatory system of the Pak proto-oncogene, and support the potential link between the development of the regulatory system of proto-oncogenes and the evolution of multicellularity. Further analysis of oncogenic functions of proto-oncogene orthologs in the unicellular genes would provide some insights into the mechanisms of the destruction of multicellular society in cancer.

  5. Biphasic regulation of myosin light chain phosphorylation by p21-activated kinase modulates intestinal smooth muscle contractility.

    PubMed

    Chu, Ji; Pham, Ngoc T; Olate, Nicole; Kislitsyna, Karina; Day, Mary-Clare; LeTourneau, Phillip A; Kots, Alexander; Stewart, Randolph H; Laine, Glen A; Cox, Charles S; Uray, Karen

    2013-01-11

    Supraphysiological mechanical stretching in smooth muscle results in decreased contractile activity. However, the mechanism is unclear. Previous studies indicated that intestinal motility dysfunction after edema development is associated with increased smooth muscle stress and decreased myosin light chain (MLC) phosphorylation in vivo, providing an ideal model for studying mechanical stress-mediated decrease in smooth muscle contraction. Primary human intestinal smooth muscle cells (hISMCs) were subjected to either control cyclical stretch (CCS) or edema (increasing) cyclical stretch (ECS), mimicking the biophysical forces in non-edematous and edematous intestinal smooth muscle in vivo. ECS induced significant decreases in phosphorylation of MLC and MLC phosphatase targeting subunit (MYPT1) and a significant increase in p21-activated kinase (PAK) activity compared with CCS. PAK regulated MLC phosphorylation in an activity-dependent biphasic manner. PAK activation increased MLC and MYPT1 phosphorylation in CCS but decreased MLC and MYPT1 phosphorylation in hISMCs subjected to ECS. PAK inhibition had the opposite results. siRNA studies showed that PAK1 plays a critical role in regulating MLC phosphorylation in hISMCs. PAK1 enhanced MLC phosphorylation via phosphorylating MYPT1 on Thr-696, whereas PAK1 inhibited MLC phosphorylation via decreasing MYPT1 on both Thr-696 and Thr-853. Importantly, in vivo data indicated that PAK activity increased in edematous tissue, and inhibition of PAK in edematous intestine improved intestinal motility. We conclude that PAK1 positively regulates MLC phosphorylation in intestinal smooth muscle through increasing inhibitory phosphorylation of MYPT1 under physiologic conditions, whereas PAK1 negatively regulates MLC phosphorylation via inhibiting MYPT1 phosphorylation when PAK activity is increased under pathologic conditions.

  6. Cytotaxonomy of Eurypyga helias (Gruiformes, Eurypygidae): First Karyotypic Description and Phylogenetic Proximity with Rynochetidae

    PubMed Central

    dos Santos, Michelly da Silva; Tagliarini, Marcella Mergulhão; O´Brien, Patricia C. M.; Ferguson-Smith, Malcolm A.; de Oliveira, Edivaldo H. C.

    2015-01-01

    The sunbittern (Eurypyga helias) is a South American Gruiformes, the only member of Family Eurypigidae. In most phylogenetic proposals, it is placed in a more distant position than other families of the so-called “core Gruiformes”. Different studies based on molecular, morphological and biogeographical data suggest that the Eurypigidae is closely related to the kagu (Rhynochetos jubatus), the only species in Rynochetidae, another family not included in the core Gruiformes. Here, the karyotype of the sunbittern is described for the first time, by classical and molecular cytogenetics, using whole chromosome probes derived from Gallus gallus and Leucopternis albicollis. We found a diploid number of 80, with only one pair of biarmed autosomal macrochromosomes, similar to that observed in the kagu. Chromosome painting revealed that most syntenies found in the avian putative ancestral karyotype (PAK) were conserved in the sunbittern. However, PAK1, PAK2, and PAK5 corresponded to two chromosome pairs each. Probes derived from L. albicollis confirm that fissions in PAK1 and PAK2 were centric, whereas in PAK5 the fission is interstitial. In addition, there is fusion of segments homologous to PAK2q and PAK5. From a phylogenetic point of view, comparisons of our results with two other Gruiformes belonging to family Rallidae suggest that the PAK5q fission might be a synapomorphy for Gruiformes. Fissions in PAK1 and PAK2 are found only in Eurypigidae, and might also occur in Rynochetidae, in view of the similar chromosomal morphology between the sunbittern and the kagu. This suggests a close phylogenetic relationship between Eurypigidae and Rynochetidae, whose common ancestor was separated by the Gondwana vicariancy in South America and New Caledonia, respectively. PMID:26624624

  7. Ablation of p21-activated kinase-1 in mice promotes isoproterenol-induced cardiac hypertrophy in association with activation of Erk1/2 and inhibition of protein phosphatase 2A.

    PubMed

    Taglieri, Domenico M; Monasky, Michelle M; Knezevic, Ivana; Sheehan, Katherine A; Lei, Ming; Wang, Xin; Chernoff, Jonathan; Wolska, Beata M; Ke, Yunbo; Solaro, R John

    2011-12-01

    Earlier investigations in our lab indicated an anti-adrenergic effect induced by activation of p21-activated kinase (Pak-1) and protein phosphatase 2A (PP2A). Our objective was to test the hypothesis that Pak-1/PP2A is a signaling cascade controlling stress-induced cardiac growth. We determined the effects of ablation of the Pak-1 gene on the response of the myocardium to chronic stress of isoproterenol (ISO) administration. Wild-type (WT) and Pak-1-knockout (Pak-1-KO) mice were randomized into six groups to receive either ISO, saline (CTRL), or ISO and FR180204, a selective inhibitor of Erk1/2. Echocardiography revealed that hearts of the Pak-1-KO/ISO group had increased LV fractional shortening, reduced LV chamber volume in diastole and systole, increased cardiac hypertrophy, and enhanced transmitral early filling deceleration time, compared to all other groups. The changes were associated with an increase in relative Erk1/2 activation in Pak-1-KO/ISO mice versus all other groups. ISO-induced cardiac hypertrophy and Erk1/2 activation in Pak-1-KO/ISO were attenuated when the selective Erk1/2 inhibitor FR180204 was administered. Immunoprecipitation showed an association between Pak-1, PP2A, and Erk1/2. Cardiac myocytes infected with an adenoviral vector expressing constitutively active Pak-1 showed a repression of Erk1/2 activation. p38 MAPK phosphorylation was decreased in Pak-1-KO/ISO and Pak-1-KO/CTRL mice compared to WT. Levels of phosphorylated PP2A were increased in ISO-treated Pak-1-KO mice, indicating reduced phosphatase activity. Maximum Ca(2+)-activated tension in detergent-extracted bundles of papillary fibers from ISO-treated Pak-1-KO mice was higher than in all other groups. Analysis of cTnI phosphorylation indicated that compared to WT, ISO-induced phosphorylation of cTnI was blunted in Pak-1-KO mice. Active Pak-1 is a natural inhibitor of Erk1/2 and a novel anti-hypertrophic signaling molecule upstream of PP2A. 2011 Elsevier Ltd. All rights reserved.

  8. Basic fibroblast growth factor-induced translocation of p21-activated kinase to the membrane is independent of phospholipase C-gamma1 in the differentiation of PC12 cells.

    PubMed

    Shin, Kyung-Sun; Shin, Eun-Young; Lee, Chan-Soo; Quan, Song-Hua; Woo, Kyung-Nam; Soung, Nak-Kyun; Kwak, Sahng-June; Kim, Seung Ryul; Kim, Eung-Gook

    2002-05-31

    p21-activated kinase (PAK) targeting to the plasma membrane is essential for PC12 cell neurite outgrowth. Phospholipase C-gamma1 (PLC-gamma1) can mediate the PAK translocation in response to growth factors, since PLC-gamma1 binds to both tyrosine-phosphorylated receptor tyrosine kinases and PAK through its SH2 and SH3 domain, respectively. In the present study, we examined a potential role for PLC-gamma1 in the basic fibroblast growth factor (bFGF)-induced PAK translocation using stable PC12 cell lines that overexpress in a tetracycline-inducible manner either the wild-type FGFR-1 or the Y766F FGFR-1 mutant. Phosphatidylinositol hydrolysis was increased 6.5-fold in response to bFGF in the wild type cells but negligible in the mutant cells. The recombinant GST-PLC-gamma1 SH3 was able to bind to PAK1 but not GST alone. However, examination of PLC-gamma1 as an adaptor for translocation of PAK1 in cells showed that both cells transfected with pEGFP-PAK1 was able to differentiate for 24 h, as visualized by laser confocal microscopy. Translocation of PAK1 to growth cones occurs at similar levels in both wild and mutant cells. These results suggest that a protein(s) other than PLC-gamma1 is functionally relevant for PAK targeting.

  9. P21-activated kinase 1 regulates resistance to BRAF inhibition in human cancer cells.

    PubMed

    Babagana, Mahamat; Johnson, Sydney; Slabodkin, Hannah; Bshara, Wiam; Morrison, Carl; Kandel, Eugene S

    2017-01-04

    BRAF is a commonly mutated oncogene in various human malignancies and a target of a new class of anti-cancer agents, BRAF-inhibitors (BRAFi). The initial enthusiasm for these agents, based on the early successes in the management of metastatic melanoma, is now challenged by the mounting evidence of intrinsic BRAFi-insensitivity in many BRAF-mutated tumors, by the scarcity of complete responses, and by the inevitable emergence of drug resistance in initially responsive cases. These setbacks put an emphasis on discovering the means to increase the efficacy of BRAFi and to prevent or overcome BRAFi-resistance. We explored the role of p21-activated kinases (PAKs), in particular PAK1, in BRAFi response. BRAFi lowered the levels of active PAK1 in treated cells. An activated form of PAK1 conferred BRAFi-resistance on otherwise sensitive cells, while genetic or pharmacologic suppression of PAK1 had a sensitizing effect. While activation of AKT1 and RAC1 proto-oncogenes increased BRAFi-tolerance, the protective effect was negated in the presence of PAK inhibitors. Furthermore, combining otherwise ineffective doses of PAK- and BRAF-inhibitors synergistically affected intrinsically BRAFi-resistant cells. Considering the high incidence of PAK1 activation in cancers, our findings suggests PAK inhibition as a strategy to augment BRAFi therapy and overcome some of the well-known resistance mechanisms.

  10. p-21-Activated kinase 1 mediates gastrin-stimulated proliferation in the colorectal mucosa via multiple signaling pathways.

    PubMed

    Huynh, Nhi; Yim, Mildred; Chernoff, Jonathan; Shulkes, Arthur; Baldwin, Graham S; He, Hong

    2013-03-15

    Gastrins, including amidated (Gamide) and glycine-extended (Ggly) forms, function as growth factors for the gastrointestinal mucosa. The p-21-activated kinase 1 (PAK1) plays important roles in growth factor signaling networks that control cell motility, proliferation, differentiation, and transformation. PAK1, activated by both Gamide and Ggly, mediates gastrin-stimulated proliferation and migration, and activation of β-catenin, in gastric epithelial cells. The aim of this study was to investigate the role of PAK1 in the regulation by gastrin of proliferation in the normal colorectal mucosa in vivo. Mucosal proliferation was measured in PAK1 knockout (PAK1 KO) mice by immunohistochemistry. The expression of phosphorylated and unphosphorylated forms of the signaling molecules PAK1, extracellular signal-regulated kinase (ERK), and protein kinase B (AKT), and the expression of β-catenin and its downstream targets c-Myc and cyclin D1, were measured in gastrin knockout (Gas KO) and PAK1 KO mice by Western blotting. The expression and activation of PAK1 are decreased in Gas KO mice, and these decreases are associated with reduced activation of ERK, AKT, and β-catenin. Proliferation in the colorectal mucosa of PAK1 KO mice is reduced, and the reduction is associated with reduced activation of ERK, AKT, and β-catenin. In compensation, antral gastrin mRNA and serum gastrin concentrations are increased in PAK1 KO mice. These results indicate that PAK1 mediates the stimulation of colorectal proliferation by gastrins via multiple signaling pathways involving activation of ERK, AKT, and β-catenin.

  11. Group I p21-activated kinases facilitate Tax-mediated transcriptional activation of the human T-cell leukemia virus type 1 long terminal repeats

    PubMed Central

    2013-01-01

    Background Human T-cell leukemia virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia and tropical spastic paraparesis. HTLV-1 encodes transactivator protein Tax that interacts with various cellular factors to modulate transcription and other biological functions. Additional cellular mediators of Tax-mediated transcriptional activation of HTLV-1 long terminal repeats (LTR) remain to be identified and characterized. Results In this study, we investigated the regulatory role of group I p21-activated kinases (Paks) in Tax-induced LTR activation. Both wild-type and kinase-dead mutants of Pak3 were capable of potentiating the activity of Tax to activate LTR transcription. The effect of Paks on the LTR was attributed to the N-terminal regulatory domain and required the action of CREB, CREB-regulating transcriptional coactivators (CRTCs) and p300/CREB-binding protein. Paks physically associated with Tax and CRTCs. Paks were recruited to the LTR in the presence of Tax. siRNAs against either Pak1 or Pak3 prevented the interaction of Tax with CRTC1 and the recruitment of Tax to the LTR. These siRNAs also inhibited LTR-dependent transcription in HTLV-1-transformed MT4 cells and in cells transfected with an infectious clone of HTLV-1. Conclusion Group I Paks augment Tax-mediated transcriptional activation of HTLV-1 LTR in a kinase-independent manner. PMID:23622267

  12. Phosphorylation-dependent regulation of nuclear localization and functions of integrin-linked kinase

    PubMed Central

    Acconcia, Filippo; Barnes, Christopher J.; Singh, Rajesh R.; Talukder, Amjad H.; Kumar, Rakesh

    2007-01-01

    Integrin-linked kinase (ILK) is a phosphorylated protein that regulates physiological processes that overlap with those regulated by p21-activated kinase 1 (PAK1). Here we report the possible role of ILK phosphorylation by PAK1 in ILK-mediated signaling and intracellular translocation. We found that PAK1 phosphorylates ILK at threonine-173 and serine-246 in vitro and in vivo. Depletion of PAK1 decreased the levels of endogenous ILK phosphorylation in vivo. Mutation of PAK1 phosphorylation sites on ILK to alanine reduced cell motility and cell proliferation. Biochemical fractionation, confocal microscopy, and chromatin-interaction analyses of human cells revealed that ILK localizes predominantly in the cytoplasm but also resides in the nucleus. Transfection of MCF-7 cells with point mutants ILK-T173A, ILK-S246A, or ILK-T173A; S246A (ILK-DM) altered ILK localization. Selective depletion of PAK1 dramatically increased the nuclear and focal point accumulation of ILK, further demonstrating a role for PAK1 in ILK translocation. We also identified functional nuclear localization sequence and nuclear export sequence motifs in ILK, delineated an apparently integral role for ILK in maintaining normal nuclear integrity, and established that ILK interacts with the regulatory region of the CNKSR3 gene chromatin to negatively modulate its expression. Together, these results suggest that ILK is a PAK1 substrate, undergoes phosphorylation-dependent shuttling between the cell nucleus and cytoplasm, and interacts with gene-regulatory chromatin. PMID:17420447

  13. Infrared emission features: probing the interstellar PAH population and circumstellar environment of Herbig Ae/Be stars

    NASA Astrophysics Data System (ADS)

    Boersma, Christiaan

    2009-12-01

    AKs zijn alom vertegenwoordigd en bieden een uitstekend middel om de veelheid aan objecten verspreid over het heelal te bestuderen. Met name in gebieden waar zich sterren en planeten vormen, helpen ze bij het ontwarren van de grootschalige structuur. PAKs staat voor polycyclische aromatische koolwaterstoffen en ze vormen een familie van grote kippengaasvormige moleculen opgebouwd uit koolstof en waterstof. Op aarde worden ze onderander aangetroffen in de verbrandingsproducten van fossiele brandstoffen. PAKs vormen het overgangsgebied van stofdeeltjes ter grote van een micron naar moleculair "gas". PAKs zijn uniek op twee manieren. Allereerst, PAKs fluoresceren na de absorptie van een enkel ultraviolet foton, waardoor ze te zien zijn in zeer koude gebieden, ver weg van de aanstralende bron. In de tweede plaats, gegeven de complexiteit van deze moleculen, kunnen ze een belangrijke rol hebben gespeeld in het ontstaan van leven. Met behulp van topklasse ruimte- en grond gestationeerde observatoria, zoals bijvoorbeeld Spitzer en de 10-meter klasse telescopen in Chili, is de PAK-emissie afkomstig van middelzware, zich vormende, sterren onderzocht. Door gebruik van zowel beeldvorming als spectroscopie, zijn morfologische en evolutionaire aspecten van de PAK-emissie vastgesteld. De NASA Ames PAK IR Spectroscopische Database is een verzameling van meer dan 600 berekend en ongeveer 200 experimenteel bepaalde spectra. Deze unieke database gaat eind 2009 publiek. Gebruikmakend van deze database is een systematische zoektocht gedaan naar kandidaten die verantwoordelijk kunnen worden gehouden voor de emissie in twee, tot op heden, niet goed bestudeerde regio's van het PAKs-spectra.

  14. 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

  15. Regulation of the Cool/Pix proteins: key binding partners of the Cdc42/Rac targets, the p21-activated kinases.

    PubMed

    Feng, Qiyu; Albeck, John G; Cerione, Richard A; Yang, Wannian

    2002-02-15

    The Cool (cloned-out of library)/Pix (for PAK-interactive exchange factor) proteins directly bind to members of the PAK family of serine/threonine kinases and regulate their activity. Three members of the Cool/Pix family have shown distinct regulatory activities: (i) p50(Cool-1) inhibits Cdc42/Rac-stimulated PAK activity, (ii) p85(Cool-1)/beta-Pix has a permissive effect on Cdc42/Rac-stimulated activity, and (iii) p90(Cool-2)/alpha-Pix strongly activates PAK. We initially suspected that these different functional effects were due to a binding interaction that occurs at the carboxyl-terminal ends of the larger Cool/Pix proteins, thus enabling them to stimulate (or at least permit) rather than inhibit PAK activity. This led to the identification of the Cat proteins (for Cool-associated tyrosine phosphosubstrates). However, here we show that the Cat proteins bind to the carboxyl-terminal ends of p85(Cool-1) (residues 523-546) and Cool-2 (residues 647-670), and that the binding of Cat to Cool-2 in fact is not necessary for the Cool-2-mediated activation of PAK. Rather, an 18-amino acid region, designated T1, that is present in the Cool-1 proteins, but missing in Cool-2, is essential for controlling the regulation of PAK activity by Cool-1/beta-Pix in vivo. Deletion of T1 yielded a p85(Cool-1) molecule that mimicked the Cool-2 protein and was capable of strongly stimulating PAK activity. However, when T1 was added to Cool-2, the ability of Cool-2 to directly activate PAK was lost. We conclude that T1 represents a novel regulatory domain that accounts for the specific functional effects on PAK activity exhibited by the different members of the Cool/Pix family.

  16. Functional integrity of the t-tubular system in cardiomyocytes depends on p21-activated kinase 1

    PubMed Central

    DeSantiago, Jaime; Bare, Dan J; Ke, Yunbo; Sheehan, Katherine A.; Solaro, R. John; Banach, Kathrin

    2013-01-01

    p21-activated kinase (Pak1), a serine-threonine protein kinase, regulates cytoskeletal dynamics and cell motility. Recent experiments further demonstrate that loss of Pak1 results in exaggerated hypertrophic growth in response to pathophysiological stimuli. Calcium (Ca) signaling plays an important role in the regulation of transcription factors involved in hypertrophic remodeling. Here we aimed to determine the role of Pak1 in cardiac excitation-contraction coupling (ECC). Ca transients were recorded in isolated, ventricular myocytes (VMs) from WT and Pak1−/− mice. Pak1−/− Ca transients had a decreased amplitude, prolonged rise time and delayed recovery time. Di-8-ANNEPS staining revealed a decreased t-tubular density in Pak1−/− VMs that coincided with decreased cell capacitance and increased dis-synchrony of Ca induced Ca release (CICR) at individual release units. These changes were not observed in atrial myocytes of Pak1−/− mice where the t-tubular system is only sparsely developed. Experiments in cultured rabbit VMs supported a role of Pak1 in the maintenance of the t-tubular structure. T-tubular density in rabbit VMs significantly decreased within 24h of culture. This was accompanied by a decrease of the Ca transient amplitude and a prolongation of its rise time. However, overexpression of constitutively active Pak1 in VMs attenuated the structural remodeling as well as changes in ECC. The results provide significant support for a prominent role of Pak1 activity not only in the functional regulation of ECC but for the structural maintenance of the t-tubular system whose remodeling is an integral feature of hypertrophic remodeling. PMID:23612118

  17. Leucine-rich repeat kinase 2 interacts with p21-activated kinase 6 to control neurite complexity in mammalian brain.

    PubMed

    Civiero, Laura; Cirnaru, Maria Daniela; Beilina, Alexandra; Rodella, Umberto; Russo, Isabella; Belluzzi, Elisa; Lobbestael, Evy; Reyniers, Lauran; Hondhamuni, Geshanthi; Lewis, Patrick A; Van den Haute, Chris; Baekelandt, Veerle; Bandopadhyay, Rina; Bubacco, Luigi; Piccoli, Giovanni; Cookson, Mark R; Taymans, Jean-Marc; Greggio, Elisa

    2015-12-01

    Leucine-rich repeat kinase 2 (LRRK2) is a causative gene for Parkinson's disease, but the physiological function and the mechanism(s) by which the cellular activity of LRRK2 is regulated are poorly understood. Here, we identified p21-activated kinase 6 (PAK6) as a novel interactor of the GTPase/ROC domain of LRRK2. p21-activated kinases are serine-threonine kinases that serve as targets for the small GTP binding proteins Cdc42 and Rac1 and have been implicated in different morphogenetic processes through remodeling of the actin cytoskeleton such as synapse formation and neuritogenesis. Using an in vivo neuromorphology assay, we show that PAK6 is a positive regulator of neurite outgrowth and that LRRK2 is required for this function. Analyses of post-mortem brain tissue from idiopathic and LRRK2 G2019S carriers reveal an increase in PAK6 activation state, whereas knock-out LRRK2 mice display reduced PAK6 activation and phosphorylation of PAK6 substrates. Taken together, these results support a critical role of LRRK2 GTPase domain in cytoskeletal dynamics in vivo through the novel interactor PAK6, and provide a valuable platform to unravel the mechanism underlying LRRK2-mediated pathophysiology. We propose p21-activated kinase 6 (PAK6) as a novel interactor of leucine-rich repeat kinase 2 (LRRK2), a kinase involved in Parkinson's disease (PD). In health, PAK6 regulates neurite complexity in the brain and LRRK2 is required for its function, (a) whereas PAK6 is aberrantly activated in LRRK2-linked PD brain (b) suggesting that LRRK2 toxicity is mediated by PAK6.

  18. The case for pancreas after kidney transplantation.

    PubMed

    Fridell, Jonathan A; Mangus, Richard S; Hollinger, Edward F; Taber, Tim E; Goble, Michelle L; Mohler, Elaine; Milgrom, Martin L; Powelson, John A

    2009-01-01

    Pancreas after kidney (PAK) transplantation has historically demonstrated inferior pancreas allograft survival compared to simultaneous pancreas and kidney (SPK) transplantation. Under our current immunosuppression protocol, we have noted excellent outcomes and rare immunological graft loss. The goal of this study was to compare pancreas allograft survival in PAK and SPK recipients using this regimen. This was a single center retrospective review of all SPK and PAK transplants performed between January 2003 and November 2007. All transplants were performed with systemic venous drainage and enteric exocrine drainage. Immunosuppression included induction with rabbit anti-thymocyte globulin (thymoglobulin), early steroid withdrawal, and maintenance with tacrolimus and sirolimus or mycophenolate mofetil. Study end points included graft and patient survival and immunosuppression related complications. Transplants included PAK 61 (30%) and SPK 142 (70%). One-yr patient survival was PAK 98% and SPK 95% (p = 0.44) and pancreas graft survival was PAK 95% and SPK 90% (p = 0.28). Acute cellular rejection was uncommon with 2% requiring treatment in each group. Survival for PAK using thymoglobulin induction, early steroid withdrawal and tacrolimus-based immunosuppression is at least comparable to SPK and should be pursued in the recipient with a potential living donor.

  19. Dermoscopy of Pigmented Actinic Keratosis of the Face: A Study of 232 Cases.

    PubMed

    Kelati, A; Baybay, H; Moscarella, E; Argenziano, G; Gallouj, S; Mernissi, F Z

    2017-07-11

    The diagnosis of pigmented actinic keratosis (PAK) is often challenging because of overlapping features with lentigo maligna. To investigate dermoscopic patterns of PAK according to their different evolutionary stages, and to correlate the pattern with clinical characteristics of the patients. Descriptive and analytical study of 232 PAK. Dermoscopic patterns were divided into two categories: the follicule surroundings' abnormalities (FSA) and follicular keratosis' abnormalities (FKA). FSA and FKA dermoscopic patterns were related to male gender, except for star-like appearance, double white clods and dermoscopic horn (p≤0.04). Rhomboidal, annular granular pattern, gray halo, white circle and double clods were dermoscopic pattern significantly related to xeroderma pigmentosum's type of skin. Based on the evolutionary stages of PAK, the jelly sign was significantly related to thin patches of PAK. Central crusts and scales were related to thick plaques and the star-like appearance to hypertrophic PAK. The presence of 2 or more dermoscopic signs in both FSA and FKA was noticed in 99.1% of lesions. The dermoscopic diagnosis of PAK vary according to the evolutionary stages of the disease, this will increase the diagnosis accuracy, with therapeutic implications. Copyright © 2017 AEDV. Publicado por Elsevier España, S.L.U. All rights reserved.

  20. LeftyA decreases Actin Polymerization and Stiffness in Human Endometrial Cancer Cells

    PubMed Central

    Salker, Madhuri S.; Schierbaum, Nicolas; Alowayed, Nour; Singh, Yogesh; Mack, Andreas F.; Stournaras, Christos; Schäffer, Tilman E.; Lang, Florian

    2016-01-01

    LeftyA, a cytokine regulating stemness and embryonic differentiation, down-regulates cell proliferation and migration. Cell proliferation and motility require actin reorganization, which is under control of ras-related C3 botulinum toxin substrate 1 (Rac1) and p21 protein-activated kinase 1 (PAK1). The present study explored whether LeftyA modifies actin cytoskeleton, shape and stiffness of Ishikawa cells, a well differentiated endometrial carcinoma cell line. The effect of LeftyA on globular over filamentous actin ratio was determined utilizing Western blotting and flow cytometry. Rac1 and PAK1 transcript levels were measured by qRT-PCR as well as active Rac1 and PAK1 by immunoblotting. Cell stiffness (quantified by the elastic modulus), cell surface area and cell volume were studied by atomic force microscopy (AFM). As a result, 2 hours treatment with LeftyA (25 ng/ml) significantly decreased Rac1 and PAK1 transcript levels and activity, depolymerized actin, and decreased cell stiffness, surface area and volume. The effect of LeftyA on actin polymerization was mimicked by pharmacological inhibition of Rac1 and PAK1. In the presence of the Rac1 or PAK1 inhibitor LeftyA did not lead to significant further actin depolymerization. In conclusion, LeftyA leads to disruption of Rac1 and Pak1 activity with subsequent actin depolymerization, cell softening and cell shrinkage. PMID:27404958

  1. Entamoeba histolytica RacC Selectively Engages p21-Activated Kinase Effectors

    PubMed Central

    2015-01-01

    Rho family GTPases modulate actin cytoskeleton dynamics by signaling through multiple effectors, including the p21-activated kinases (PAKs). The intestinal parasite Entamoeba histolytica expresses ∼20 Rho family GTPases and seven isoforms of PAK, two of which have been implicated in pathogenesis-related processes such as amoebic motility and invasion and host cell phagocytosis. Here, we describe two previously unstudied PAK isoforms, EhPAK4 and EhPAK5, as highly specific effectors of EhRacC. A structural model based on 2.35 Å X-ray crystallographic data of a complex between EhRacCQ65L·GTP and the EhPAK4 p21 binding domain (PBD) reveals a fairly well-conserved Rho/effector interface despite deviation of the PBD α-helix. A structural comparison with EhRho1 in complex with EhFormin1 suggests likely determinants of Rho family GTPase signaling specificity in E. histolytica. These findings suggest a high degree of Rho family GTPase diversity and specificity in the single-cell parasite E. histolytica. Because PAKs regulate pathogenesis-related processes in E. histolytica, they may be valid pharmacologic targets for anti-amoebiasis drugs. PMID:25529118

  2. A new role for cofilin in retinal neovascularization

    PubMed Central

    Kumar, Raj; Janjanam, Jagadeesh; Singh, Nikhlesh K.; Rao, Gadiparthi N.

    2016-01-01

    ABSTRACT Pak1 plays an important role in several cellular processes, including cell migration, but its role in pathological angiogenesis is not known. Here, we have determined its role in pathological retinal angiogenesis using an oxygen-induced retinopathy (OIR) model. VEGFA induced phosphorylation of Pak1 and its effector cofilin in a manner that was dependent on time as well as p38MAPKβ (also known as MAPK11) in human retinal microvascular endothelial cells (HRMVECs). Depletion of the levels of any of these molecules inhibited VEGFA-induced HRMVEC F-actin stress fiber formation, migration, proliferation, sprouting and tube formation. In accordance with these observations, hypoxia induced Pak1 and cofilin phosphorylation with p38MAPKβ being downstream to Pak1 and upstream to cofilin in mouse retina. Furthermore, Pak1 deficiency abolished hypoxia-induced p38MAPKβ and cofilin phosphorylation and abrogated retinal endothelial cell proliferation, tip cell formation and neovascularization. In addition, small interfering RNA (siRNA)-mediated downregulation of p38MAPKβ or cofilin levels in the wild-type mouse retina also diminished endothelial cell proliferation, tip cell formation and neovascularization. Taken together, these observations suggest that, although the p38MAPKβ–Pak1–cofilin axis is required for HRMVEC migration, proliferation, sprouting and tubulogenesis, Pak1–p38MAPKβ–cofilin signaling is also essential for hypoxia-induced mouse retinal endothelial cell proliferation, tip cell formation and neovascularization. PMID:26857814

  3. p21-activated kinase 1 restricts tonic endocannabinoid signaling in the hippocampus

    PubMed Central

    Xia, Shuting; Zhou, Zikai; Leung, Celeste; Zhu, Yuehua; Pan, Xingxiu; Qi, Junxia; Morena, Maria; Hill, Matthew N; Xie, Wei; Jia, Zhengping

    2016-01-01

    PAK1 inhibitors are known to markedly improve social and cognitive function in several animal models of brain disorders, including autism, but the underlying mechanisms remain elusive. We show here that disruption of PAK1 in mice suppresses inhibitory neurotransmission through an increase in tonic, but not phasic, secretion of endocannabinoids (eCB). Consistently, we found elevated levels of anandamide (AEA), but not 2-arachidonoylglycerol (2-AG) following PAK1 disruption. This increased tonic AEA signaling is mediated by reduced cyclooxygenase-2 (COX-2), and COX-2 inhibitors recapitulate the effect of PAK1 deletion on GABAergic transmission in a CB1 receptor-dependent manner. These results establish a novel signaling process whereby PAK1 upregulates COX-2, reduces AEA and restricts tonic eCB-mediated processes. Because PAK1 and eCB are both critically involved in many other organ systems in addition to the brain, our findings may provide a unified mechanism by which PAK1 regulates these systems and their dysfunctions including cancers, inflammations and allergies. DOI: http://dx.doi.org/10.7554/eLife.14653.001 PMID:27296803

  4. The Cossidae (Lepidoptera) of Afghanistan with description of three new species and special notes on the fauna of Bande-Amir National Park.

    PubMed

    Yakovlev, Roman V; Pljustch, Igor G; Skrylnik, Yuriy; Pak, Oleg; Witt, Thomas J

    2015-07-23

    The annotated list of Cossidae of Afghanistan consists of 44 species in 17 genera from the four subfamilies Catoptinae, Cossinae, Zeuzerinae, and Mehariinae. Three new species are described: Cossulus habibae Yakovlev, Pljustch, Skrylnik & Pak, sp. nov., Semagystia bamiani Yakovlev, Pljustch, Skrylnik & Pak, sp. nov., Phragmacossia bandeamiri Yakovlev, Pljustch, Skrylnik & Pak, sp. nov.; all from Band-e-Amir National Park in Bamian Province. Three species (Dervishiya cadambae (Moore, 1865), Semagystia cossoides (Graeser, 1892), Phragmacossia territa (Staudinger, 1879)) are reported for the first time from Afghanistan. A brief biogeographical analysis of the Cossidae of Afghanistan is given.

  5. Genetics Home Reference: Dandy-Walker malformation

    MedlinePlus

    ... from mild to severe, and those with normal intelligence may have learning disabilities. Children with Dandy-Walker ... Dandy-Walker Malformation: A Clinical and Surgical Outcome Analysis. J Coll Physicians Surg Pak. 2015 Jun;25( ...

  6. The Cbl proteins are binding partners for the Cool/Pix family of p21-activated kinase-binding proteins.

    PubMed

    Flanders, James A; Feng, Qiyu; Bagrodia, Shubha; Laux, Maria T; Singavarapu, Avinash; Cerione, Richard A

    2003-08-28

    Members of the Cool protein family contain SH3, Dbl, and pleckstrin homology domains and are binding partners for the p21-activated kinase (PAK). Using the yeast two-hybrid screen, we identified Cbl-b as a Cool family binding partner. We co-immunoprecipitated endogenous Cool and Cbl-b from a variety of breast cancer cell lines. The Cool-Cbl-b interaction requires the SH3 domain of Cool and competes with the binding of PAK to Cool proteins. Expression of Cbl-b effectively blocks the ability of Cool-2 to stimulate PAK, thus providing an additional mechanism, aside from catalyzing receptor ubiquitination, by which Cbl-b acts as a negative regulator for signaling activities requiring PAK activation.

  7. Precision Nanoparticles

    ScienceCinema

    John Hemminger

    2016-07-12

    A revolutionary technology that efficiently produces nanoparticles in uniform and prescribed sizes (1-100 nanometers) using supercritical fluids. INL researcher Robert Fox was joined by Idaho State University researchers Rene Rodriquez and Joshua Pak in d

  8. Light exposure before learning improves memory consolidation at night

    PubMed Central

    Shan, Li-Li; Guo, Hao; Song, Ning-Ning; Jia, Zheng-Ping; Hu, Xin-Tian; Huang, Jing-Fei; Ding, Yu-Qiang; Richter-Levine, Gal; Zhou, Qi-Xin; Xu, Lin

    2015-01-01

    Light is recently recognized as a modulator able to activate the hippocampus and modulate memory processing, but little is known about the molecular mechanisms. Here, we report that in mice, a short pulse of white light before learning dramatically improves consolidation of contextual fear memory during the night. The light exposure increases hippocampal active p21-activated kinase 1 (PAK1) and CA1 long-term potentiation (LTP). These light effects are abolished in PAK1 knockout and dominant-negative transgenic mice, but preserved by expression of constitutively active PAK1 in the hippocampus. Our results indicate that light can act as a switch of PAK1 activity that modulate CA1 LTP and thereby memory consolidation without affecting learning and short-term memory. PMID:26493375

  9. Precision Nanoparticles

    SciTech Connect

    John Hemminger

    2009-07-21

    A revolutionary technology that efficiently produces nanoparticles in uniform and prescribed sizes (1-100 nanometers) using supercritical fluids. INL researcher Robert Fox was joined by Idaho State University researchers Rene Rodriquez and Joshua Pak in d

  10. Further Adventures of Dr. Sony's Mad Monster Machine, or Cutting the Studio Umbilical Cord

    ERIC Educational Resources Information Center

    Steffin, Sherwin

    1977-01-01

    We must reexamine our often total reliance on the studio as the only source for instructional television production. The porta-pak provides an alternative. Tips are provided for successful field production. (Author/STS)

  11. Ombitasvir, Paritaprevir, Ritonavir, and Dasabuvir

    MedlinePlus

    Viekira Pak® (as a combination product containing Ombitasvir, Paritaprevir, Ritonavir, Dasabuvir) ... C infection (swelling of the liver caused by a virus). Ombitasvir is a hepatitis C virus (HCV) ...

  12. Small-molecule p21-activated kinase inhibitor PF-3758309 is a potent inhibitor of oncogenic signaling and tumor growth

    PubMed Central

    Murray, Brion W.; Guo, Chuangxing; Piraino, Joseph; Westwick, John K.; Zhang, Cathy; Lamerdin, Jane; Dagostino, Eleanor; Knighton, Daniel; Loi, Cho-Ming; Zager, Michael; Kraynov, Eugenia; Popoff, Ian; Christensen, James G.; Martinez, Ricardo; Kephart, Susan E.; Marakovits, Joseph; Karlicek, Shannon; Bergqvist, Simon; Smeal, Tod

    2010-01-01

    Despite abundant evidence that aberrant Rho-family GTPase activation contributes to most steps of cancer initiation and progression, there is a dearth of inhibitors of their effectors (e.g., p21-activated kinases). Through high-throughput screening and structure-based design, we identify PF-3758309, a potent (Kd = 2.7 nM), ATP-competitive, pyrrolopyrazole inhibitor of PAK4. In cells, PF-3758309 inhibits phosphorylation of the PAK4 substrate GEF-H1 (IC50 = 1.3 nM) and anchorage-independent growth of a panel of tumor cell lines (IC50 = 4.7 ± 3 nM). The molecular underpinnings of PF-3758309 biological effects were characterized using an integration of traditional and emerging technologies. Crystallographic characterization of the PF-3758309/PAK4 complex defined determinants of potency and kinase selectivity. Global high-content cellular analysis confirms that PF-3758309 modulates known PAK4-dependent signaling nodes and identifies unexpected links to additional pathways (e.g., p53). In tumor models, PF-3758309 inhibits PAK4-dependent pathways in proteomic studies and regulates functional activities related to cell proliferation and survival. PF-3758309 blocks the growth of multiple human tumor xenografts, with a plasma EC50 value of 0.4 nM in the most sensitive model. This study defines PAK4-related pathways, provides additional support for PAK4 as a therapeutic target with a unique combination of functions (apoptotic, cytoskeletal, cell-cycle), and identifies a potent, orally available small-molecule PAK inhibitor with significant promise for the treatment of human cancers. PMID:20439741

  13. Pancreas-after-kidney versus synchronous pancreas-kidney transplantation: comparison of intermediate-term results.

    PubMed

    Bazerbachi, Fateh; Selzner, Markus; Marquez, Max A; Norgate, Andrea; McGilvray, Ian D; Schiff, Jeffrey; Cattral, Mark S

    2013-02-15

    Controversy persists over the safety and efficacy of pancreas transplantation in patients with insulin-dependent diabetes mellitus who have received a prior kidney transplant. We compared the outcomes of recipients who received either Synchronous-Pancreas Kidney-Transplantation (SPK, n=123) or Pancreas-After-Kidney-Transplants(n=49)at our institution between August 2002 to January 2010. Donor and recipient demographics were similar. Time interval between kidney and pancreas transplantation was 5.9 ± 3.8 (4.8 [1.6-12.2]) years. The majority of kidney-recipients in PAK group were transplanted at outside institutions and referred to us for PAK. Most patients received thymoglobulin induction and were maintained on tacrolimus, MMF, and prednisone. For SPK versus PAK recipients, there was no difference in median of length of hospital stay or incidence of overall complications. All PAK recipients are alive with functioning kidney grafts, whereas the 1-, 3-, and 5-year SPK patient survival rates were 98%,96%,and 94%, P=0.09. The 1-,3-, and 5-yr uncensored pancreas survival rates for SPK versus PAK were 93% vs. 90%, 90% vs. 90%, and 82% versus 85%, respectively (P=0.4). Glycemic control and intermediate survival outcomes were similar in both groups. Pancreas-graft outcomes in SPK and PAK were equivalent in our study, but our specific population entailed among other factors a long K to PAK time interval; PAK could be a comparable option to SPK for patients with access to kidney grafts.

  14. Long-term outcomes of pancreas after kidney transplantation in small centers: is it justified?

    PubMed

    Laftavi, M R; Pankewycz, O; Gruessner, A; Brian, Murray; Kohli, R; Feng, L; Said, M; Sharma, R; Patel, S

    2014-01-01

    Currently, the long-term advantages of having a pancreas transplantation (PT) are debated, particularly in patients receiving pancreas after kidney (PAK) allografts. The United Network for Organ Sharing (UNOS) requires that a transplant center perform a minimum number of PT per year to remain an active PT center. The long-term outcomes and challenges of PAK in small pancreas transplant centers are not well studied. In this retrospective analysis, we report short- and long-term outcomes in a small center performing 2-9 PT annually. Forty-eight PT (25 simultaneous pancreas and kidney transplantation [SPK], 23 PAK) were performed in our center. Donor and recipient demographics were similar in both groups. All suitable local donors were used for SPK. All organs for PAK transplantation were imported from other UNOS regions. Mean follow-up was 61 ± 46 and 74 ± 46 months for SPK and PAK, respectively. Patient and graft survival rates were similar in SPK and PAK groups and better than the reported national average. Four patients (11%) died (1 due to trauma, 1 brain lymphoma, 1 ruptured aneurysm; and 1 unknown cause). Two patients (4%; 1 SPK, 1 PAK) lost their grafts because of thrombosis on postoperative days 3 and 5 in 2002. No graft thrombosis occurred since 2002. Seven patients (15%) required reoperation (4 for bleeding, 2 anastomotic leaks, 1 small bowel perforation). Two patients (4%) developed post-transplantation lymphoproliferative disease. Five patients (11%) experienced cytomegalovirus antigenemia which responded well to antiviral therapy. Compared with outcomes for diabetic patients on dialysis, current SPK and PAK short- and long-term results are favorable even in a small PT center. Therefore, unless there is a contraindication, PT should be offered to all type 1 diabetic patients with end-stage renal disease at the time of kidney transplantation or afterward. Copyright © 2014 Elsevier Inc. All rights reserved.

  15. PAK–PIX interactions regulate adhesion dynamics and membrane protrusion to control neurite outgrowth

    PubMed Central

    Santiago-Medina, Miguel; Gregus, Kelly A.; Gomez, Timothy M.

    2013-01-01

    Summary The roles of P21-activated kinase (PAK) in the regulation of axon outgrowth downstream of extracellular matrix (ECM) proteins are poorly understood. Here we show that PAK1–3 and PIX are expressed in the developing spinal cord and differentially localize to point contacts and filopodial tips within motile growth cones. Using a specific interfering peptide called PAK18, we found that axon outgrowth is robustly stimulated on laminin by partial inhibition of PAK–PIX interactions and PAK function, whereas complete inhibition of PAK function stalls axon outgrowth. Furthermore, modest inhibition of PAK–PIX stimulates the assembly and turnover of growth cone point contacts, whereas strong inhibition over-stabilizes adhesions. Point mutations within PAK confirm the importance of PIX binding. Together our data suggest that regulation of PAK–PIX interactions in growth cones controls neurite outgrowth by influencing the activity of several important mediators of actin filament polymerization and retrograde flow, as well as integrin-dependent adhesion to laminin. PMID:23321640

  16. Disruption of p21-activated kinase 1 gene diminishes atherosclerosis in apolipoprotein E-deficient mice

    PubMed Central

    Singh, Nikhlesh K.; Kotla, Sivareddy; Dyukova, Elena; Traylor Jr., James G.; Orr, A. Wayne; Chernoff, Jonathan; Marion, Tony N.; Rao, Gadiparthi N.

    2015-01-01

    Pak1 plays an important role in various cellular processes, including cell motility, polarity, survival and proliferation. To date, its role in atherogenesis has not been explored. Here we report the effect of Pak1 on atherogenesis using atherosclerosis-prone apolipoprotein E-deficient (ApoE−/−) mice as a model. Disruption of Pak1 in ApoE−/− mice results in reduced plaque burden, significantly attenuates circulating IL-6 and MCP-1 levels, limits the expression of adhesion molecules and diminishes the macrophage content in the aortic root of ApoE−/− mice. We also observed reduced oxidized LDL uptake and increased cholesterol efflux by macrophages and smooth muscle cells of ApoE−/−:Pak1−/− mice as compared with ApoE−/− mice. In addition, we detect increased Pak1 phosphorylation in human atherosclerotic arteries, suggesting its role in human atherogenesis. Altogether, these results identify Pak1 as an important factor in the initiation and progression of atherogenesis. PMID:26104863

  17. Role of pili in adhesion of Pseudomonas aeruginosa to human respiratory epithelial cells.

    PubMed Central

    Doig, P; Todd, T; Sastry, P A; Lee, K K; Hodges, R S; Paranchych, W; Irvin, R T

    1988-01-01

    The ability of pili from Pseudomonas aeruginosa K (PAK) to act as an adhesin to human respiratory epithelial cells was examined using an in vitro adhesion assay. Equilibrium analysis of PAK binding to human buccal epithelial cells (BECs) and tracheal epithelial cells (TECs) by means of a Langmuir adsorption isotherm revealed that the maximum numbers of binding sites per epithelial cell (N) were 255 for BECs and 236 for TECs, with apparent association constants (Ka) of 2.8 x 10(-9) and 5.8 x 10(-9) ml/CFU, respectively. Trypsinization of the BECs before the binding assay increased N to 605 and decreased the Ka to 1.7 x 10(-9) ml/CFU. Addition of homologous pili to the binding assay with BECs or TECs or the addition of anti-pilus Fab fragments inhibited PAK adherence. Binding of purified pili to BECs was shown to reach saturation. Purified pili and PAK competed for the same receptor on the BEC surface. Further, by using peptide fragments of PAK pilin (derived from the native pili or produced synthetically) in the binding assay for PAK to BECs, we have presumptively identified the pilus binding domain in the C-terminal region of the pilin and shown that the C-terminal disulfide bridge is important in maintaining the functionality of the binding domain. PMID:2897336

  18. The Search for Therapeutic Bacteriophages Uncovers One New Subfamily and Two New Genera of Pseudomonas-Infecting Myoviridae

    PubMed Central

    Henry, Marine; Bobay, Louis-Marie; Chevallereau, Anne; Saussereau, Emilie; Ceyssens, Pieter-Jan; Debarbieux, Laurent

    2015-01-01

    In a previous study, six virulent bacteriophages PAK_P1, PAK_P2, PAK_P3, PAK_P4, PAK_P5 and CHA_P1 were evaluated for their in vivo efficacy in treating Pseudomonas aeruginosa infections using a mouse model of lung infection. Here, we show that their genomes are closely related to five other Pseudomonas phages and allow a subdivision into two clades, PAK_P1-like and KPP10-like viruses, based on differences in genome size, %GC and genomic contents, as well as number of tRNAs. These two clades are well delineated, with a mean of 86% and 92% of proteins considered homologous within individual clades, and 25% proteins considered homologous between the two clades. By ESI-MS/MS analysis we determined that their virions are composed of at least 25 different proteins and electron microscopy revealed a morphology identical to the hallmark Salmonella phage Felix O1. A search for additional bacteriophage homologs, using profiles of protein families defined from the analysis of the 11 genomes, identified 10 additional candidates infecting hosts from different species. By carrying out a phylogenetic analysis using these 21 genomes we were able to define a new subfamily of viruses, the Felixounavirinae within the Myoviridae family. The new Felixounavirinae subfamily includes three genera: Felixounalikevirus, PAK_P1likevirus and KPP10likevirus. Sequencing genomes of bacteriophages with therapeutic potential increases the quantity of genomic data on closely related bacteriophages, leading to establishment of new taxonomic clades and the development of strategies for analyzing viral genomes as presented in this article. PMID:25629728

  19. Pancreas survival in simultaneous pancreas-kidney and pancreas-after-kidney transplantations: a five-year follow-up report.

    PubMed

    Dinckan, Ayhan; Aliosmanoglu, Ibrahim; Kocak, Huseyin; Sari, Ramazan; Erdogan, Okan; Ertug, Zeki; Suleymanlar, Gultekin; Gurkan, Alihan

    2012-01-01

    Pancreas transplantation methods, such as simultaneous pancreas-kidney (SPK) transplantation and pancreas-after-kidney (PAK) transplantation, have become the most important treatments for patients with type-1 diabetes mellitus (DM)-related end-stage renal diseases (ESRD). The purpose of the study was to compare the clinical results of the pancreas graft in patients after SPK and PAK transplantations and to present the findings of our 5-year follow-up. A total of 55 patients who had kidney and pancreas transplantation between February 2003 and December 2010 were included in the study. The patients were divided into 2 groups based on the timing of the pancreas transplantation: SPK (n=21) and PAK (n=34). The patients in the SPK group consisted of 13 males and 8 females, with a mean age of 33.6±6.8 years; whereas 25 males and 9 females formed the PAK group, with a mean age of 32.0±6.0 years. In the early postoperative period, the SPK group had 3 patients with vascular thrombosis (2 venous, 1 arterial) and the PAK group had 7 patients with thrombosis (4 venous, 3 arterial) (p=0.319). At the end of the 5-year follow-up, the patient, kidney and pancreas survival rates in the SPK group were 95.2%, 95.2%, and 61.9% respectively, and the corresponding values in the PAK group were 97%, 91.2%, 61.8% (p=0.382, p=0.504, p=0.927). We concluded that PAK is just as effective as SPK to prevent the destructive effects of DM when the waiting time for SPK is long and a potential live donor is present.

  20. Mesalamine modulates intercellular adhesion through inhibition of p-21 activated kinase-1

    PubMed Central

    Khare, Vineeta; Lyakhovich, Alex; Dammann, Kyle; Lang, Michaela; Borgmann, Melanie; Tichy, Boris; Pospisilova, Sarka; Luciani, Gloria; Campregher, Christoph; Evstatiev, Rayko; Pflueger, Maren; Hundsberger, Harald; Gasche, Christoph

    2013-01-01

    Mesalamine (5-ASA) is widely used for the treatment of ulcerative colitis, a remitting condition characterized by chronic inflammation of the colon. Knowledge about the molecular and cellular targets of 5-ASA is limited and a clear understanding of its activity in intestinal homeostasis and interference with neoplastic progression is lacking. We sought to identify molecular pathways interfered by 5-ASA, using CRC cell lines with different genetic background. Microarray was performed for gene expression profile of 5-ASA-treated and untreated cells (HCT116 and HT29). Filtering and analysis of data identified three oncogenic pathways interfered by 5-ASA: MAPK/ERK pathway, cell adhesion and β-catenin/Wnt signaling. PAK1 emerged as a consensus target of 5-ASA, orchestrating these pathways. We further investigated the effect of 5-ASA on cell adhesion. 5-ASA increased cell adhesion which was measured by cell adhesion assay and transcellular-resistance measurement. Moreover, 5-ASA treatment restored membranous expression of adhesion molecules E-cadherin and β-catenin. Role of PAK1 as a mediator of mesalamine activity was validated in vitro and in vivo. Inhibition of PAK1 by RNA interference also increased cell adhesion. PAK1 expression was elevated in APCmin polyps and 5-ASA treatment reduced its expression. Our data demonstrates novel pharmacological mechanism of mesalamine in modulation of cell adhesion and role of PAK1 in APCmin polyposis. We propose that inhibition of PAK1 expression by 5-ASA can impede with neoplastic progression in colorectal carcinogenesis. The mechanism of PAK1 inhibition and induction of membranous translocation of adhesion proteins by 5-ASA might be independent of its known anti-inflammatory action. PMID:23146664

  1. Pseudomonas aeruginosa selective adherence to and entry into human endothelial cells.

    PubMed Central

    Plotkowski, M C; Saliba, A M; Pereira, S H; Cervante, M P; Bajolet-Laudinat, O

    1994-01-01

    The pathogenesis of Pseudomonas aeruginosa disseminated infections depends on bacterial interaction with blood vessels. We have hypothesized that in order to traverse the endothelial barrier, bacteria would have to adhere to and damage endothelial cells. To test this hypothesis, we studied the adherence to human endothelial cells in primary culture of the piliated P. aeruginosa strain PAK and of two isogenic nonpiliated strains: PAK/p-, which carries a mutation in the pilin structural gene, and PAK-N1, a mutant defective in the regulatory rpoN gene. PAK adhered significantly more than did the pilus-lacking strains. P. aeruginosa was also taken up by endothelial cells, as determined by quantitative bacteriologic assays and by transmission electron microscopy. This internalization of P. aeruginosa seems to be a selective process, since the piliated strain was taken up significantly more than the nonpiliated bacteria and the avirulent Escherichia coli DH5 alpha, even following bacterial centrifugation onto the cell monolayers. A significant fraction of the internalized P. aeruginosa PAK was recovered in a viable form after 6 h of residence within endothelial cells. Progressive endothelial cell damage resulted from PAK intracellular harboring, as indicated by the release of lactate dehydrogenase. An increasing concentration of PAK cells was recovered from the extracellular medium with time, suggesting that ingested bacteria were released from endothelial cells and multiplied freely. We speculate that in vivo the ability of some P. aeruginosa strains to resist intracellular residence would afford protection from host defenses and antibiotics and that the release of viable bacteria into bloodstream may represent a central feature of the pathogenesis of bacteremia in compromised patients. Images PMID:7960126

  2. Nuclear factor of activated T cells c1 mediates p21-activated kinase 1 activation in the modulation of chemokine-induced human aortic smooth muscle cell F-actin stress fiber formation, migration, and proliferation and injury-induced vascular wall remodeling.

    PubMed

    Kundumani-Sridharan, Venkatesh; Singh, Nikhlesh K; Kumar, Sanjay; Gadepalli, Ravisekhar; Rao, Gadiparthi N

    2013-07-26

    Recent literature suggests that cyclin-dependent kinases (CDKs) mediate cell migration. However, the mechanisms were not known. Therefore, the objective of this study is to test whether cyclin/CDKs activate Pak1, an effector of Rac1, whose involvement in the modulation of cell migration and proliferation is well established. Monocyte chemotactic protein 1 (MCP1) induced Pak1 phosphorylation/activation in human aortic smooth muscle cells (HASMCs) in a delayed time-dependent manner. MCP1 also stimulated F-actin stress fiber formation in a delayed manner in HASMCs, as well as the migration and proliferation of these cells. Inhibition of Pak1 suppressed MCP1-induced HASMC F-actin stress fiber formation, migration, and proliferation. MCP1 induced cyclin D1 expression as well as CDK6 and CDK4 activities, and these effects were dependent on activation of NFATc1. Depletion of NFATc1, cyclin D1, CDK6, or CDK4 levels attenuated MCP1-induced Pak1 phosphorylation/activation and resulted in decreased HASMC F-actin stress fiber formation, migration, and proliferation. CDK4, which appeared to be activated downstream of CDK6, formed a complex with Pak1 in response to MCP1. MCP1 also activated Rac1 in a time-dependent manner, and depletion/inhibition of its levels/activation abrogated MCP1-induced NFATc1-cyclin D1-CDK6-CDK4-Pak1 signaling and, thereby, decreased HASMC F-actin stress fiber formation, migration, and proliferation. In addition, smooth muscle-specific deletion of NFATc1 led to decreased cyclin D1 expression and CDK6, CDK4, and Pak1 activities, resulting in reduced neointima formation in response to injury. Thus, these observations reveal that Pak1 is a downstream effector of CDK4 and Rac1-dependent, NFATc1-mediated cyclin D1 expression and CDK6 activity mediate this effect. In addition, smooth muscle-specific deletion of NFATc1 prevented the capacity of vascular smooth muscle cells for MCP-1-induced activation of the cyclin D1-CDK6-CDK4-Pak1 signaling axis, affecting

  3. Subverting Host Cell P21-Activated Kinase: A Case of Convergent Evolution across Pathogens.

    PubMed

    John Von Freyend, Simona; Kwok-Schuelein, Terry; Netter, Hans J; Haqshenas, Gholamreza; Semblat, Jean-Philippe; Doerig, Christian

    2017-04-21

    Intracellular pathogens have evolved a wide range of strategies to not only escape from the immune systems of their hosts, but also to directly exploit a variety of host factors to facilitate the infection process. One such strategy is to subvert host cell signalling pathways to the advantage of the pathogen. Recent research has highlighted that the human serine/threonine kinase PAK, or p21-activated kinase, is a central component of host-pathogen interactions in many infection systems involving viruses, bacteria, and eukaryotic pathogens. PAK paralogues are found in most mammalian tissues, where they play vital roles in a wide range of functions. The role of PAKs in cell proliferation and survival, and their involvement in a number of cancers, is of great interest in the context of drug discovery. In this review we discuss the latest insights into the surprisingly central role human PAK1 plays for the infection by such different infectious disease agents as viruses, bacteria, and parasitic protists. It is our intention to open serious discussion on the applicability of PAK inhibitors for the treatment, not only of neoplastic diseases, which is currently the primary objective of drug discovery research targeting these enzymes, but also of a wide range of infectious diseases.

  4. Subverting Host Cell P21-Activated Kinase: A Case of Convergent Evolution across Pathogens

    PubMed Central

    John von Freyend, Simona; Kwok-Schuelein, Terry; Netter, Hans J.; Haqshenas, Gholamreza; Semblat, Jean-Philippe; Doerig, Christian

    2017-01-01

    Intracellular pathogens have evolved a wide range of strategies to not only escape from the immune systems of their hosts, but also to directly exploit a variety of host factors to facilitate the infection process. One such strategy is to subvert host cell signalling pathways to the advantage of the pathogen. Recent research has highlighted that the human serine/threonine kinase PAK, or p21-activated kinase, is a central component of host-pathogen interactions in many infection systems involving viruses, bacteria, and eukaryotic pathogens. PAK paralogues are found in most mammalian tissues, where they play vital roles in a wide range of functions. The role of PAKs in cell proliferation and survival, and their involvement in a number of cancers, is of great interest in the context of drug discovery. In this review we discuss the latest insights into the surprisingly central role human PAK1 plays for the infection by such different infectious disease agents as viruses, bacteria, and parasitic protists. It is our intention to open serious discussion on the applicability of PAK inhibitors for the treatment, not only of neoplastic diseases, which is currently the primary objective of drug discovery research targeting these enzymes, but also of a wide range of infectious diseases. PMID:28430160

  5. Endogenous, hyperactive Rac3 controls proliferation of breast cancer cells by a p21-activated kinase-dependent pathway

    PubMed Central

    Mira, Jean-Paul; Benard, Valerie; Groffen, John; Sanders, Luraynne C.; Knaus, Ulla G.

    2000-01-01

    Uncontrolled cell proliferation is a major feature of cancer. Experimental cellular models have implicated some members of the Rho GTPase family in this process. However, direct evidence for active Rho GTPases in tumors or cancer cell lines has never been provided. In this paper, we show that endogenous, hyperactive Rac3 is present in highly proliferative human breast cancer-derived cell lines and tumor tissues. Rac3 activity results from both its distinct subcellular localization at the membrane and altered regulatory factors affecting the guanine nucleotide state of Rac3. Associated with active Rac3 was deregulated, persistent kinase activity of two isoforms of the Rac effector p21-activated kinase (Pak) and of c-Jun N-terminal kinase (JNK). Introducing dominant-negative Rac3 and Pak1 fragments into a breast cancer cell line revealed that active Rac3 drives Pak and JNK kinase activities by two separate pathways. Only the Rac3–Pak pathway was critical for DNA synthesis, independently of JNK. These findings identify Rac3 as a consistently active Rho GTPase in human cancer cells and suggest an important role for Rac3 and Pak in tumor growth. PMID:10618392

  6. Evaluation of Serodiagnostic Assays for Mycobacterium bovis Infection in Elk, White-Tailed Deer, and Reindeer in the United States

    PubMed Central

    Nelson, Jeffrey T.; Orloski, Kathleen A.; Lloyd, Audra L.; Camacho, Mark; Schoenbaum, Mark A.; Robbe-Austerman, Suelee; Thomsen, Bruce V.; Hall, S. Mark

    2012-01-01

    In 2011, the United States Department of Agriculture conducted a project in which elk (Cervus elaphus spp.), white-tailed deer (WTD) (Odocoileus virginianus), and reindeer (Rangifer tarandus) were evaluated by the single cervical tuberculin test (SCT), comparative cervical tuberculin test (CCT), and serologic tests. The rapid antibody detection tests evaluated were the CervidTB Stat-Pak (Stat-Pak), and the Dual Path Platform VetTB (DPP). Blood was collected from presumably uninfected animals prior to tuberculin injection for the SCT. A total of 1,783 animals were enrolled in the project. Of these, 1,752 (98.3%) were classified as presumably uninfected, based on originating from a captive cervid herd with no history of exposure to TB. Stat-Pak specificity estimates were 92.4% in reindeer, 96.7% in WTD, and 98.3% in elk and were not significantly different from SCT specificity estimates. Using the DPP in series on Stat-Pak antibody-positive samples improved specificity in the three species. Thirty one animals were classified as confirmed infected, based on necropsy and laboratory results, and 27/31 were antibody positive on Stat-Pak for an estimated sensitivity of 87.1%. The study findings indicate that rapid serologic tests used in series are comparable to the SCT and CCT and may have a greater ability to detect TB-infected cervids. PMID:22792512

  7. Predicting In Vivo Efficacy of Therapeutic Bacteriophages Used To Treat Pulmonary Infections

    PubMed Central

    Henry, Marine; Lavigne, Rob

    2013-01-01

    The potential of bacteriophage therapy to treat infections caused by antibiotic-resistant bacteria has now been well established using various animal models. While numerous newly isolated bacteriophages have been claimed to be potential therapeutic candidates on the basis of in vitro observations, the parameters used to guide their choice among billions of available bacteriophages are still not clearly defined. We made use of a mouse lung infection model and a bioluminescent strain of Pseudomonas aeruginosa to compare the activities in vitro and in vivo of a set of nine different bacteriophages (PAK_P1, PAK_P2, PAK_P3, PAK_P4, PAK_P5, CHA_P1, LBL3, LUZ19, and PhiKZ). For seven bacteriophages, a good correlation was found between in vitro and in vivo activity. While the remaining two bacteriophages were active in vitro, they were not sufficiently active in vivo under similar conditions to rescue infected animals. Based on the bioluminescence recorded at 2 and 8 h postinfection, we also define for the first time a reliable index to predict treatment efficacy. Our results showed that the bacteriophages isolated directly on the targeted host were the most efficient in vivo, supporting a personalized approach favoring an optimal treatment. PMID:24041900

  8. Complete nucleotide sequence and taxonomy of Sugarcane streak mosaic virus, member of a novel genus in the family Potyviridae.

    PubMed

    Xu, D-L; Zhou, G-H; Xie, Y-J; Mock, R; Li, R

    2010-06-01

    The complete genomic sequence of a Pakistani isolate of Sugarcane streak mosaic virus (SCSMV-PAK) is determined to be 9782 nucleotides in length, excluding the 3' poly(A) tail, and it comprises a large open reading frame encoding a polyprotein of 3130 amino acid residues. The deduced polyprotein is likely to be cleaved at nine putative protease sites by three viral proteases to ten mature proteins. Conserved motifs of orthologous proteins of other potyviruses are identified in corresponding positions of SCSMV-PAK. The genomic organization is virtually identical to the genera Ipomovirus, Potyvirus, Rymovirus, and Tritimovirus in the family Potyviridae. Sequence analyses indicate that the SCSMV-PAK genomic sequence is different from those of Sugarcane mosaic virus and Sorghum mosaic virus, two viruses with very similar symptoms and host range to SCSMV-PAK. SCSMV-PAK shares 52.7% identity with Triticum mosaic virus (TriMV) and 26.4-31.5% identities with species of the existing genera and unassigned viruses in the Potyviridae at the polyprotein sequence level. Phylogenetic analyses of the polyprotein and deduced mature protein amino acid sequences reveal that SCSMV, together with TriMV, forms a distinct group in the family at the genus level. Therefore, SCSMV should represent a new genus, Susmovirus, in the Potyviridae.

  9. Ethnic Differences in Appointment-Keeping and Implications for the Patient-Centered Medical Home—Findings from the Diabetes Study of Northern California (DISTANCE)

    PubMed Central

    Parker, Melissa M; Moffet, Howard H; Schillinger, Dean; Adler, Nancy; Fernandez, Alicia; Ciechanowski, Paul; Karter, Andrew J

    2012-01-01

    Objective To examine ethnic differences in appointment-keeping in a managed care setting. Data Sources/Study Setting Kaiser Permanente Diabetes Study of Northern California (DISTANCE), 2005–2007, n = 12,957. Study Design Cohort study. Poor appointment-keeping (PAK) was defined as missing >1/3 of planned, primary care appointments. Poisson regression models were used to estimate ethnic-specific relative risks of PAK (adjusting for demographic, socio-economic, health status, and facility effects). Data Collection/Extraction Methods Administrative/electronic health records and survey responses. Principal Findings Poor appointment-keeping rates differed >2-fold across ethnicities: Latinos (12 percent), African Americans (10 percent), Filipinos (7 percent), Caucasians (6 percent), and Asians (5 percent), but also varied by medical center. Receiving >50 percent of outpatient care via same-day appointments was associated with a 4-fold greater PAK rate. PAK was associated with 20, 30, and 40 percent increased risk of elevated HbA1c (>7 percent), low-density lipoprotein (>100 mm/dl), and systolic blood pressure (>130 mmHg), respectively. Conclusions Latinos and African Americans were at highest risk of missing planned primary care appointments. PAK was associated with a greater reliance on same-day visits and substantively poorer clinical outcomes. These results have important implications for public health and health plan policy, as primary care rapidly expands toward open access to care supported by the patient-centered medical home model. PMID:22091785

  10. The impact of method on kidney graft and patient survival in kidney-pancreas transplantations for type I diabetes mellitus.

    PubMed

    Dinckan, Ayhan; Aliosmanoglu, Ibrahim; Kocak, Huseyin; Mesci, Ayhan; Altunbas, Hasan; Gurkan, Alihan

    2015-01-01

    Patients who develop end-stage renal disease (ESRD) associated with Type I Diabetes Mellitus may receive kidney alone (KA) transplantation, simultaneous pancreas-kidney (SPK) transplantation, or a pancreas after kidney (PAK) transplantation. The goal of this study is to examine the long-term impact of pancreas transplantation on kidney graft and patient survival rates. A total of 85 transplantation cases, consisting of 30 that received living donor KA, 21 that received SPK, and 34 that received PAK, from 2003-2010 at Akdeniz University Organ Transplantation Institute were retrospectively screened. There was a graft loss in 4 cases from the KA group, and in 1 case from each of the SPK and PAK groups. The five-year kidney graft survival rates were 86.7% in KA, 95.2% in SPK, and 97.1% in PAK. There was a single patient loss in both KA and SPK. The kidney survival percentages were higher in SPK and PAK groups compared to the KA group. Therefore, SPK should be the primary preference in these patients; however, for the cases that have a living donor, pancreas transplantation should be considered after kidney transplantation, or the patients can be followed-up on with close blood sugar control.

  11. p21-activated kinase regulates mast cell degranulation via effects on calcium mobilization and cytoskeletal dynamics

    PubMed Central

    Allen, Jayme D.; Jaffer, Zahara M.; Park, Su-Jung; Burgin, Sarah; Hofmann, Clemens; Sells, Mary Ann; Chen, Shi; Derr-Yellin, Ethel; Michels, Elizabeth G.; McDaniel, Andrew; Bessler, Waylan K.; Ingram, David A.; Atkinson, Simon J.; Travers, Jeffrey B.

    2009-01-01

    Mast cells are key participants in allergic diseases via activation of high-affinity IgE receptors (FcϵRI) resulting in release of proinflammatory mediators. The biochemical pathways linking IgE activation to calcium influx and cytoskeletal changes required for intracellular granule release are incompletely understood. We demonstrate, genetically, that Pak1 is required for this process. In a passive cutaneous anaphylaxis experiment, Wsh/Wsh mast cell–deficient mice locally reconstituted with Pak1−/− bone marrow–derived mast cells (BMMCs) experienced strikingly decreased allergen-induced vascular permeability compared with controls. Consistent with the in vivo phenotype, Pak1−/− BMMCs exhibited a reduction in FcϵRI-induced degranulation. Further, Pak1−/− BMMCs demonstrated diminished calcium mobilization and altered depolymerization of cortical filamentous actin (F-actin) in response to FcϵRI stimulation. These data implicate Pak1 as an essential molecular target for modulating acute mast cell responses that contribute to allergic diseases. PMID:19124833

  12. Effect of plant species on nitrogen recovery in aquaponics.

    PubMed

    Hu, Zhen; Lee, Jae Woo; Chandran, Kartik; Kim, Sungpyo; Brotto, Ariane Coelho; Khanal, Samir Kumar

    2015-01-01

    Nitrogen transformations in aquaponics with different edible plant species, i.e., tomato (Lycopersicon esculentum) and pak choi (Brassica campestris L. subsp. chinensis) were systematically examined and compared. Results showed that nitrogen utilization efficiencies (NUE) of tomato- and pak choi-based aquaponic systems were 41.3% and 34.4%, respectively. The abundance of nitrifying bacteria in tomato-based aquaponics was 4.2-folds higher than that in pak choi-based aquaponics, primarily due to its higher root surface area. In addition, tomato-based aquaponics had better water quality than that of pak choi-based aquaponics. About 1.5-1.9% of nitrogen input were emitted to atmosphere as nitrous oxide (N2O) in tomato- and pak choi-based aquaponic systems, respectively, suggesting that aquaponics is a potential anthropogenic source of N2O emission. Overall, this is the first intensive study that examined the role plant species played in aquaponics, which could provide new strategy in designing and operating an aquaponic system.

  13. Glucose-regulated protein 78 (GRP78) binds directly to PIP3 phosphatase SKIP and determines its localization.

    PubMed

    Ijuin, Takeshi; Hatano, Naoya; Takenawa, Tadaomi

    2016-05-01

    Skeletal muscle and kidney-enriched inositol polyphosphate phosphatase (SKIP), a PIP3 phosphatase, has been implicated in the regulation of insulin signaling in skeletal muscle. SKIP interacts with Pak1 and glucose-regulated protein 78 (GRP78), both of which are necessary for the regulation of insulin signaling. In this study, we showed that GRP78 directly binds to the SKIP C-terminal homology (SKICH) domain of SKIP and that this binding is necessary for the localization of SKIP at the ER. In addition, in vitro binding analysis showed that GRP78 and Pak1 competitively bind to SKIP. Taken together, these findings suggest a model by which GRP78 regulates intracellular localization of SKIP and how SKIP binds to Pak1 on insulin stimulation.

  14. Activity-dependent regulation of genes implicated in X-linked non-specific mental retardation.

    PubMed

    Boda, B; Mas, C; Muller, D

    2002-01-01

    X-linked forms of non-specific mental retardation are complex disorders, for which mutations in several genes have recently been identified. These include OPHN1, GDI1, PAK3, IL1RAPL, TM4SF2, FMR2 and RSK2. To investigate the mechanisms through which alterations of these gene products could result in cognitive impairment, we analyzed their expression using quantitative PCR technique in two in vitro models of activity-dependent gene regulation: kainate-induced seizures and long-term synaptic potentiation (LTP). We found that the level of expression of four genes, PAK3, IL1RAPL, RSK2 and TM4SF2, was significantly up-regulated following kainate treatment. Furthermore we observed a significant increase in mRNA levels of PAK3 and IL1RAPL following LTP induction. These results suggest a possible role for these four genes in activity-dependent brain plasticity.

  15. Glioblastoma following treatment with fingolimod for relapsing-remitting multiple sclerosis.

    PubMed

    Sharim, Justin; Tashjian, Randy; Golzy, Nima; Pouratian, Nader

    2016-08-01

    Glioblastoma is an uncommon and aggressive primary brain tumor with incidence of 3 per 100,000 annually. We report a 50-year-old woman diagnosed with glioblastoma within threeyears of induction of fingolimod therapy for relapsing-remitting multiple sclerosis. Fingolimod, an immunomodulating agent used in the treatment of relapsing-remitting multiple sclerosis, has also been suggested to impart a cardioprotective role in heart failure and arrhythmia via activation of P21-activated kinase-1 (Pak1). In the brain, Pak1 activation has been shown to correlate with decreased survival time amongst patients with glioblastoma. A molecular mechanism underlying a link between fingolimod use and glioblastoma development may involve activation of Pak1. To our knowledge, this is the first report of a potential association between fingolimod use and glioblastoma development.

  16. Macroencapsulation of mixed waste debris at the Hanford Nuclear Reservation -- Final project report by AST Environmental Services, LLC

    SciTech Connect

    Baker, T.L.

    1998-02-25

    This report summarizes the results of a full-scale demonstration of a high density polyethylene (HDPE) package, manufactured by Arrow Construction, Inc. of Montgomery, Alabama. The HDPE package, called ARROW-PAK, was designed and patented by Arrow as both a method to macroencapsulation of radioactively contaminated lead and as an improved form of waste package for treatment and interim and final storage and/or disposal of drums of mixed waste. Mixed waste is waste that is radioactive, and meets the criteria established by the United States Environmental Protection Agency (US EPA) for a hazardous material. Results from previous testing conducted for the Department of Energy (DOE) at the Idaho National Engineering Laboratory in 1994 found that the ARROW-PAK fabrication process produces an HDPE package that passes all helium leak tests and drop tests, and is fabricated with materials impervious to the types of environmental factors encountered during the lifetime of the ARROW-PAK, estimated to be from 100 to 300 years. Arrow Construction, Inc. has successfully completed full-scale demonstration of its ARROW-PAK mixed waste macroencapsulation treatment unit at the DOE Hanford Site. This testing was conducted in accordance with Radiological Work Permit No. T-860, applicable project plans and procedures, and in close consultation with Waste Management Federal Services of Hanford, Inc.`s project management, health and safety, and quality assurance representatives. The ARROW-PAK field demonstration successfully treated 880 drums of mixed waste debris feedstock which were compacted and placed in 149 70-gallon overpack drums prior to macroencapsulation in accordance with the US EPA Alternate Debris Treatment Standards, 40 CFR 268.45. Based on all of the results, the ARROW-PAK process provides an effective treatment, storage and/or disposal option that compares favorably with current mixed waste management practices.

  17. pix-1 controls early elongation in parallel with mel-11 and let-502 in Caenorhabditis elegans.

    PubMed

    Martin, Emmanuel; Harel, Sharon; Nkengfac, Bernard; Hamiche, Karim; Neault, Mathieu; Jenna, Sarah

    2014-01-01

    Cell shape changes are crucial for metazoan development. During Caenorhabditis elegans embryogenesis, epidermal cell shape changes transform ovoid embryos into vermiform larvae. This process is divided into two phases: early and late elongation. Early elongation involves the contraction of filamentous actin bundles by phosphorylated non-muscle myosin in a subset of epidermal (hypodermal) cells. The genes controlling early elongation are associated with two parallel pathways. The first one involves the rho-1/RHOA-specific effector let-502/Rho-kinase and mel-11/myosin phosphatase regulatory subunit. The second pathway involves the CDC42/RAC-specific effector pak-1. Late elongation is driven by mechanotransduction in ventral and dorsal hypodermal cells in response to body-wall muscle contractions, and involves the CDC42/RAC-specific Guanine-nucleotide Exchange Factor (GEF) pix-1, the GTPase ced-10/RAC and pak-1. In this study, pix-1 is shown to control early elongation in parallel with let-502/mel-11, as previously shown for pak-1. We show that pix-1, pak-1 and let-502 control the rate of elongation, and the antero-posterior morphology of the embryos. In particular, pix-1 and pak-1 are shown to control head, but not tail width, while let-502 controls both head and tail width. This suggests that let-502 function is required throughout the antero-posterior axis of the embryo during early elongation, while pix-1/pak-1 function may be mostly required in the anterior part of the embryo. Supporting this hypothesis we show that low pix-1 expression level in the dorsal-posterior hypodermal cells is required to ensure high elongation rate during early elongation.

  18. pix-1 Controls Early Elongation in Parallel with mel-11 and let-502 in Caenorhabditis elegans

    PubMed Central

    Nkengfac, Bernard; Hamiche, Karim; Neault, Mathieu; Jenna, Sarah

    2014-01-01

    Cell shape changes are crucial for metazoan development. During Caenorhabditis elegans embryogenesis, epidermal cell shape changes transform ovoid embryos into vermiform larvae. This process is divided into two phases: early and late elongation. Early elongation involves the contraction of filamentous actin bundles by phosphorylated non-muscle myosin in a subset of epidermal (hypodermal) cells. The genes controlling early elongation are associated with two parallel pathways. The first one involves the rho-1/RHOA-specific effector let-502/Rho-kinase and mel-11/myosin phosphatase regulatory subunit. The second pathway involves the CDC42/RAC-specific effector pak-1. Late elongation is driven by mechanotransduction in ventral and dorsal hypodermal cells in response to body-wall muscle contractions, and involves the CDC42/RAC-specific Guanine-nucleotide Exchange Factor (GEF) pix-1, the GTPase ced-10/RAC and pak-1. In this study, pix-1 is shown to control early elongation in parallel with let-502/mel-11, as previously shown for pak-1. We show that pix-1, pak-1 and let-502 control the rate of elongation, and the antero-posterior morphology of the embryos. In particular, pix-1 and pak-1 are shown to control head, but not tail width, while let-502 controls both head and tail width. This suggests that let-502 function is required throughout the antero-posterior axis of the embryo during early elongation, while pix-1/pak-1 function may be mostly required in the anterior part of the embryo. Supporting this hypothesis we show that low pix-1 expression level in the dorsal-posterior hypodermal cells is required to ensure high elongation rate during early elongation. PMID:24732978

  19. P21-Activated Kinase Inhibitors FRAX486 and IPA3: Inhibition of Prostate Stromal Cell Growth and Effects on Smooth Muscle Contraction in the Human Prostate

    PubMed Central

    Wang, Yiming; Gratzke, Christian; Tamalunas, Alexander; Wiemer, Nicolas; Ciotkowska, Anna; Rutz, Beata; Waidelich, Raphaela; Strittmatter, Frank; Liu, Chunxiao; Stief, Christian G.; Hennenberg, Martin

    2016-01-01

    Prostate smooth muscle tone and hyperplastic growth are involved in the pathophysiology and treatment of male lower urinary tract symptoms (LUTS). Available drugs are characterized by limited efficacy. Patients’ adherence is particularly low to combination therapies of 5α-reductase inhibitors and α1-adrenoceptor antagonists, which are supposed to target contraction and growth simultaneously. Consequently, molecular etiology of benign prostatic hyperplasia (BPH) and new compounds interfering with smooth muscle contraction or growth in the prostate are of high interest. Here, we studied effects of p21-activated kinase (PAK) inhibitors (FRAX486, IPA3) in hyperplastic human prostate tissues, and in stromal cells (WPMY-1). In hyperplastic prostate tissues, PAK1, -2, -4, and -6 may be constitutively expressed in catecholaminergic neurons, while PAK1 was detected in smooth muscle and WPMY-1 cells. Neurogenic contractions of prostate strips by electric field stimulation were significantly inhibited by high concentrations of FRAX486 (30 μM) or IPA3 (300 μM), while noradrenaline- and phenylephrine-induced contractions were not affected. FRAX486 (30 μM) inhibited endothelin-1- and -2-induced contractions. In WPMY-1 cells, FRAX486 or IPA3 (24 h) induced concentration-dependent (1–10 μM) degeneration of actin filaments. This was paralleled by attenuation of proliferation rate, being observed from 1 to 10 μM FRAX486 or IPA3. Cytotoxicity of FRAX486 and IPA3 in WPMY-1 cells was time- and concentration-dependent. Stimulation of WPMY-1 cells with endothelin-1 or dihydrotestosterone, but not noradrenaline induced PAK phosphorylation, indicating PAK activation by endothelin-1. Thus, PAK inhibitors may inhibit neurogenic and endothelin-induced smooth muscle contractions in the hyperplastic human prostate, and growth of stromal cells. Targeting prostate smooth muscle contraction and stromal growth at once by a single compound is principally possible, at least under

  20. Field Evaluation of a Rapid Immunochromatographic Assay for Detection of Trypanosoma cruzi Infection by Use of Whole Blood▿

    PubMed Central

    Roddy, Paul; Goiri, Javier; Flevaud, Laurence; Palma, Pedro Pablo; Morote, Silvia; Lima, Nines; Villa, Luis; Torrico, Faustino; Albajar-Viñas, Pedro

    2008-01-01

    Laboratory and clinical diagnostic classification of seropositive individuals, followed by treatment and supportive therapy, is an established component of Chagas' disease control in areas where this disease is endemic. However, most Chagas' disease patients live in remote areas where neither equipped laboratories nor skilled human resources are widely available. Employing a rapid diagnostic test (RDT), when using whole blood samples, is the best option for Chagas' disease control. A high sensitivity and specificity for the Chagas Stat-Pak RDT (Chembio Diagnostic Systems, Inc., Medford, NY) has been reported for assays using serum and plasma, but its validity for the detection of antibodies to Trypanosoma cruzi infection in whole blood is unknown. This cross-sectional study measured the sensitivity and specificity of the Chagas Stat-Pak with whole blood, using conventional serological assays for comparison. The interobserver reliability in the interpretation of the Chagas Stat-Pak results and “ease-of-use” criterion needed to perform the Chagas Stat-Pak and conventional assays were also measured. The Chagas Stat-Pak yielded a high specificity (99.0%, 95% confidence interval [CI] = 98.4 to 99.4%) but a relatively low sensitivity (93.4%, 95% CI = 87.4 to 97.1%). The interobserver reliability was excellent (kappa [n = 1,913] = 0.999, P < 0.0001), and the quantified ease-of-use criterion suggested that the RDT is simple to perform. Despite the attributes of the Chagas Stat-Pak, it is not an ideal diagnostic test for the population investigated in the present study due to its relatively low sensitivity and high cost. The RDT manufacturer is called upon to improve the test if the international community hopes to make progress in controlling Chagas infections in areas where this disease is endemic. PMID:18400910

  1. Prevailing PA Mutation K356R in Avian Influenza H9N2 Virus Increases Mammalian Replication and Pathogenicity

    PubMed Central

    Xu, Guanlong; Zhang, Xuxiao; Gao, Weihua; Wang, Chenxi; Wang, Jinliang; Sun, Honglei; Sun, Yipeng; Guo, Lu; Zhang, Rui; Chang, Kin-Chow; Liu, Jinhua

    2016-01-01

    ABSTRACT Adaptation of the viral polymerase complex comprising PB1, PB2, and PA is necessary for efficient influenza A virus replication in new host species. We found that PA mutation K356R (PA-K356R) has become predominant since 2014 in avian H9N2 viruses in China as with seasonal human H1N1 viruses. The same mutation is also found in most human isolates of emergent avian H7N9 and H10N8 viruses whose six internal gene segments are derived from the H9N2 virus. We further demonstrated the mammalian adaptive functionality of the PA-K356R mutation. Avian H9N2 virus with the PA-K356R mutation in human A549 cells showed increased nuclear accumulation of PA and increased viral polymerase activity that resulted in elevated levels of viral transcription and virus output. The same mutant virus in mice also enhanced virus replication and caused lethal infection. In addition, combined mutation of PA-K356R and PB2-E627K, a well-known mammalian adaptive marker, in the H9N2 virus showed further cooperative increases in virus production and severity of infection in vitro and in vivo. In summary, PA-K356R behaves as a novel mammalian tropism mutation, which, along with other mutations such as PB2-E627K, might render avian H9N2 viruses adapted for human infection. IMPORTANCE Mutations of the polymerase complex (PB1, PB2, and PA) of influenza A virus are necessary for viral adaptation to new hosts. This study reports a novel and predominant mammalian adaptive mutation, PA-K356R, in avian H9N2 viruses and human isolates of emergent H7N9 and H10N8 viruses. We found that PA-356R in H9N2 viruses causes significant increases in virus replication and severity of infection in human cells and mice and that PA-K356R cooperates with the PB2-E627K mutation, a well-characterized human adaptive marker, to exacerbate mammalian infection in vitro and in vivo. Therefore, the PA-K356R mutation is a significant adaptation in H9N2 viruses and related H7N9 and H10N8 reassortants toward human

  2. Tamoxifen Dependent Interaction Between in Estrogen Receptor and a Novel p21 Activated Kinase

    DTIC Science & Technology

    2004-06-01

    Moreover, PAK6 was directly activated by MKK6, and mutation of tyrosine 566 in a consensus MKK6 site (threonine- proline-tyrosine, TPY ) in the activation...similarly activated by MKK6, consistent with a conserved TPY motif in their activation domains. The activation of PAK6 by both p38 MAP kinase and MKK6...the other catalytic domain and blocks its function. Binding of GTP-Cdc42 or -Rac causes the AID to dissociate from the catalytic domain and activates

  3. Solid-phase extraction of sugar cane soot extract for analysis by gas chromatography with flame ionisation and mass spectrometric detection.

    PubMed

    Zamperlini, G C; Santiago-Silva, M; Vilegas, W

    2000-08-11

    The incomplete combustion of biomass is one of the most important sources of emissions of organic compounds into the atmosphere, like polycyclic aromatic hydrocarbons (PAHs) which show genotoxic activity. Since environmental samples generally contain interferents and trace amounts of PAHs of interest, concentration and clean-up procedures are usually required prior to the final chromatographic analysis. This paper discusses the performance of Sep-Pak cartridges (silica gel and RP18) on clean-up of sugar cane soot extract. The best results were obtained with a silica Sep-Pak cartridge. The recoveries ranged from 79% (benzo[b]fluoranthene) to 113% (benzo[e]pyrene).

  4. Structural and Functional Studies Indicate That the EPEC Effector, EspG, Directly Binds p21-Activated Kinase

    SciTech Connect

    Germane, Katherine L.; Spiller, Benjamin W.

    2011-09-20

    Bacterial pathogens secrete effectors into their hosts that subvert host defenses and redirect host processes. EspG is a type three secretion effector with a disputed function that is found in enteropathogenic Escherichia coli. Here we show that EspG is structurally similar to VirA, a Shigella virulence factor; EspG has a large, conserved pocket on its surface; EspG binds directly to the amino-terminal inhibitory domain of human p21-activated kinase (PAK); and mutations to conserved residues in the surface pocket disrupt the interaction with PAK.

  5. Out of Area or Out of Reach? European Military Support for Operations in Southwest Asia.

    DTIC Science & Technology

    1995-01-01

    Watson, Bruce W. Watson, Jr., David Dunphy, Richard Dejong, Brian Gagne, Michael Kirsch , and Yong Pak, "The Effects of the War on Other Nations," in...1994-95 (London: Butler and Tanner Ltd., 1994), p. 219. 12Washington Times, 13 June 1994, p. 14. 13The narrow margin of victory for Helmut Kohl’s CDU

  6. The summertime "heat" low over Pakistan/northwestern India: evolution and origin

    NASA Astrophysics Data System (ADS)

    Bollasina, Massimo; Nigam, Sumant

    2011-09-01

    A deep low in sea-level pressure is present from May to September over Pakistan and northwestern India (hereafter, the Pak-India low). It is often referred as the "heat" low to convey the significance of surface thermal effects reckoned to be important for its origin. The present analysis, rooted in observations and diagnostic modeling, suggests that the Pak-India low is influenced both by regional and remote forcing. Regionally, the influence of Hindu Kush mountains is found to be stronger than the impact of land-surface heating and attendant sensible heating of the planetary boundary layer, questioning the suitability of the "heat" label in canonical references to this circulation feature. Observational analysis indicates that the notable May-to-June deepening of the Pak-India low and its further deepening in July, however, arises from remote forcing—the development of monsoon deep-convection over the Bay of Bengal and eastern India in June and July. It is hypothesized that the associated upstream descent over Iran-Turkmenistan-Afghanistan (i.e. east of the Caspian Sea) and related low-level northerlies over the Elburz-Zagros-Hindu Kush mountains contribute to the strengthening of the Pak-India low in June (and July) from interaction with regional orography.

  7. Genomic structure of human alpha-pix, and variable deletions in a poly (T) tract in gastric cancer tissue.

    PubMed

    Wang, Y J; Oba, S M; Yoshii, S; Song, J P; Wang, Y; Kanamori, M; Ota, S; Tanaka, M; Sugimura, H

    2001-03-10

    PAK-interacting exchange factor (PIX) has been reported to mediate the recruitment of PAK into focal adhesions and activate Rac, thus creating a feedback loop that stimulate PAK and other targets. This pathway is thought to be related to cellular changes, such as transformation and migration, that are often encountered in cancer cells. Here, we report the genomic structure of alpha-PIX, one of the PAK- interacting exchange factors, including the identification of the promoter region, which consisted 772 amino acids in 22 exons, spanning about 100 kb on genome of X chromosome. All splice sites conformed to the GT-AT rule. To investigate the role of alpha-PIX in carcinogenesis, we screened 60 cases of gastric cancer for mutations and polymorphisms using an intron-primer that covered all the exons, but no mutations or polymorphisms were found in the coding region. However an 18 bp repeat of thymidine tract was present in 50 bp downstream from exon 12 and the deletion of variable numbers of mononucleotide repeats was observed in seven out of the 60 gastric cancer tissue specimens that were examined. These seven cases all exhibited a mutator phenotype, suggesting that the deletions are passenger mutations. Thus our results revealed that alpha-PIX probably does not play any primary role in human gastric carcinogenesis.

  8. The Shank family of postsynaptic density proteins interacts with and promotes synaptic accumulation of the beta PIX guanine nucleotide exchange factor for Rac1 and Cdc42.

    PubMed

    Park, Eunhye; Na, Moonseok; Choi, Jeonghoon; Kim, Seho; Lee, Jae-Ran; Yoon, Jiyoung; Park, Dongeun; Sheng, Morgan; Kim, Eunjoon

    2003-05-23

    The Shank/ProSAP family of multidomain proteins is known to play an important role in organizing synaptic multiprotein complexes. Here we report a novel interaction between Shank and beta PIX, a guanine nucleotide exchange factor for the Rac1 and Cdc42 small GTPases. This interaction is mediated by the PDZ domain of Shank and the C-terminal leucine zipper domain and the PDZ domain-binding motif at the extreme C terminus of beta PIX. Shank colocalizes with beta PIX at excitatory synaptic sites in cultured neurons. In brain, Shank forms a complex with beta PIX and beta PIX-associated signaling molecules including p21-associated kinase (PAK), an effector kinase of Rac1/Cdc42. Importantly, overexpression of Shank in cultured neurons promotes synaptic accumulation of beta PIX and PAK. Considering the involvement of Rac1 and PAK in spine dynamics, these results suggest that Shank recruits beta PIX and PAK to spines for the regulation of postsynaptic structure.

  9. A Robot to Help Make the Rounds

    NASA Technical Reports Server (NTRS)

    2003-01-01

    This paper presents a discussion on the Pyxis HelpMate SecurePak (SP) trackless robotic courier designed by Transitions Research Corporation, to navigate autonomously throughout medical facilities, transporting pharmaceuticals, laboratory specimens, equipment, supplies, meals, medical records, and radiology films between support departments and nursing floors.

  10. 9 CFR 77.34 - Official tuberculosis tests.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... used in individual captive elk, red deer, white-tailed deer, fallow deer, and reindeer, and in herds of... CervidTB Stat-Pak® test is a primary test that may be used in affected herds of captive elk, red deer...

  11. How Does Culture Shape Students' Perceptions of Scientists? Cross-National Comparative Study of American and Chinese Elementary Students

    ERIC Educational Resources Information Center

    Farland-Smith, Donna

    2009-01-01

    For decades, researchers have been convinced that one stereotypic image of scientists existed among children worldwide (Chambers, 1983; Chiang & Guo, 1996; Fung, 2002; Maoldomhnaigh & Hunt, 1988; Newton & Newton, 1992, 1998; She, 1998; Song, Pak, & Jang, 1992). This study, however, moves beyond that stereotypic image and examines…

  12. Drug Induced Pneumonitis Secondary to Treatment with Paritaprevir/Ritonavir/Ombitasvir and Dasabuvir (VIEKIRA PAK®) for Chronic Hepatitis C: Case Report of an Unexpected Life-Threatening Adverse Reaction

    PubMed Central

    Faire, Bridget; Gane, Edward

    2017-01-01

    VIEKIRA PAK (ritonavir-boosted paritaprevir/ombitasvir and dasabuvir) is an approved treatment for compensated patients with genotype 1 (GT1) chronic hepatitis C virus (HCV) infection. This oral regimen has minimal adverse effects and is well tolerated. Cure rates are 97% in patients infected with HCV GT 1a and 99% in those with HCV GT 1b. We report the first case of life-threatening allergic pneumonitis associated with VIEKIRA PAK. This unexpected serious adverse event occurred in a 68-year-old Chinese female with genotype 1b chronic hepatitis C and Child-Pugh A cirrhosis. One week into treatment with VIEKIRA PAK without ribavirin, she was admitted to hospital with respiratory distress and acute kidney injury requiring intensive care input. She was initially diagnosed with community acquired pneumonia and improved promptly with intravenous antibiotics and supported care. No bacterial or viral pathogens were cultured. Following complete recovery, she recommenced VIEKIRA PAK but represented 5 days later with more rapidly progressive respiratory failure, requiring intubation and ventilation, inotropic support, and haemodialysis. The final diagnosis was drug induced pneumonitis. PMID:28408931

  13. The Effect of an Instructional Unit Incorporating Live Animals on Knowledge of Nutrition for Different Age Levels.

    ERIC Educational Resources Information Center

    Roth, Anne I.; Wunderlich, Kenneth W.

    A nutrition education unit, Rat Pak, developed by Dairy Council, Inc., is an attempt to influence students to make wise food choices. It consists of eleven lessons in an instructional sequence which incorporates the use of white rats as a means of illustrating the effect of improper diet while teaching proper diet. The purpose of this…

  14. 77 FR 5028 - Withdrawal of Approval of New Animal Drug Applications

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-01

    ...., Box 209, Harding Hwy., Buena, NJ 08310. NADA 049-890 NORCO T-2 Pre-Pak (tylosin phosphate). Norco... procaine). Putnam, Porterville, CA 93257. ] NADA 095-953 MOORMABOOST TY 4000 Medicated (tylosin ADM..., Inc., Box 209, Harding Hwy., Buena, NJ 08310. NADA 107-957 TYLAN 20 Sulfa-G (tylosin phosphate ADM...

  15. 76 FR 16417 - Product Cancellation Order for Certain Pesticide Registrations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-23

    ....... Ammonium thiosulfate 010806-00105 Pro/Pak Shure Shot Phenothrin Wasp & Hornet Tetramethrin Spray. 013283-00013 Rainbow Wasp & Ant Bioallethrin Spray. 033660-00003 Trifluralin Trifluralin Technical. 040849-00052 Enforcer Wasp & Phenothrin Hornet Killer XI. Tetramethrin 062719-00619 Oxyfluorfen Oxyfluorfen...

  16. 76 FR 4692 - Notice of Receipt of Requests To Voluntarily Cancel Certain Pesticide Registrations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-01-26

    .... 009499-00001 Oxalis/Spurge X...... Ammonium thiosulfate. 010806-00105 Pro/Pak Shure Shot Phenothrin. Wasp & Hornet Spray. Tetramethrin. 013283-00013 Rainbow Wasp & Ant Bioallethrin. Spray. 033660-00003 Trifluralin Technical Trifluralin. 040849-00052 Enforcer Wasp & Phenothrin. Hornet Killer XI. Tetramethrin. 062719-00619...

  17. Use of GC/MS Analysis to Distinguish Between Vapor Intrusion and Indoor Sources of VOCs - Standardized Protocol for On-Site Evaluation of Vapor Intrusion

    DTIC Science & Technology

    2014-07-01

    Engineered Fluid Toluene Some paints and adhesives SprayPAK Enamel , Minwax Wood Finish Xylenes Adhesives , paints, gasoline Bonide Tree Sprays and... adhesives . However, carbon tetrachloride and chloroform are also associated with household cleaning products containing chlorine bleach (Odabasi, 2008...Radio Shack Anti Static Foaming Cleaner Chloroform Dry cleaned clothes, fire extinguishers, adhesive remover, chlorinated drinking water Time Mist

  18. Pseudomonas pili. Studies on antigenic determinants and mammalian cell receptors.

    PubMed

    Paranchych, W; Sastry, P A; Drake, D; Pearlstone, J R; Smillie, L B

    1985-01-01

    P. aeruginosa PAK pili are thin 5.2 nm diameter filaments containing a single 15-kd polypeptide subunit which is 144 amino acid residues in length. Studies on pili binding to a variety of synthetic sugars representing many di- tri- and tetra-saccharide structures found in mammalian glycoproteins and glycolipids failed to reveal any significant binding activity. On the other hand, a wide spectrum of binding activities was observed when a variety of structural proteins and enzymes were used as binding substrates. Of 30 proteins tested, phosphorylase b, pyruvate kinase and aldolase showed highest pilus binding activity. It was concluded that the PAK pilus receptor is probably a polypeptide rather than an oligosaccharide. Using arginine-specific cleavage to produce four large peptides, several proteases to produce subfragments of the large peptides, and antipilus rabbit antiserum, PAK pilin was found to contain four antigenic determinants. Epitopes near the NH2- and COOH-termini were only weakly immunogenic, whereas two epitopes near the center of the pilus protein titrated about 85% of the antipilus antibodies. Cleavage of the pilus protein into smaller peptides resulted in marked decreases in the affinity of antigenic peptides for their specific antibodies, suggesting that the immunodominant epitopes of PAK pilin are conformation-specific.

  19. Working with Children to Protect Our Future

    ERIC Educational Resources Information Center

    Williams, Nick

    2011-01-01

    The author has been teaching primary school children for 20 years, and has always been passionate about teaching young people about the environment. In this article, he describes his work with Tetra Pak and WWF-UK to develop a national, school-based competition and teaching programme to help children understand the importance of using renewable…

  20. Narrow-Band Imaging and Spatially Resolved Spectra of Nova Shells

    NASA Astrophysics Data System (ADS)

    Hillwig, T. C.; Honeycutt, R. K.; Shore, S. N.

    2000-12-01

    Observations of nova shells were made at the WIYN Observatory using the WIYN Imager, the ``naked'' DensePak fiber array, and a Barlow 4x magnifying assembly used with DensePak. DensePak was used to obtain spatially resolved spectra of several nova shells at wavelengths including the Hα , Hβ , [OIII], and [NII] emission lines. The purpose is to derive true shapes and sizes of the nova shells, velocity structure, and abundance structure. The ability to spatially resolve the shell with spectroscopy, with the accuracy and resolution available to DensePak is a useful and unique tool. The velocity structure of the shell provides data which can be compared to models of expected shell structure. Measuring abundances in different, spatially resolved portions of the shell can give indications of the cause of the structure. For example, in shaping by a fast wind, we may expect to see different abundances in the slowly moving ejected material than in the material comprising the fast wind (which becomes apparent in planetary nebulae with wind-blown bubbles). Imaging also provides, along with comparison to velocity structure, an additional constraint on the determination of parallax distances, and the narrow-band imaging can supply estimates of excitation levels in various regions of the shells. All of these are important contributors to the determination of the physical mechanism responsible for the nova shell structure. The first phase of this research is presented here.

  1. Indole-3-Acetic Acid Produced by Burkholderia heleia Acts as a Phenylacetic Acid Antagonist to Disrupt Tropolone Biosynthesis in Burkholderia plantarii

    PubMed Central

    Wang, Mengcen; Tachibana, Seiji; Murai, Yuta; Li, Li; Lau, Sharon Yu Ling; Cao, Mengchao; Zhu, Guonian; Hashimoto, Makoto; Hashidoko, Yasuyuki

    2016-01-01

    Burkholderia heleia PAK1-2 is a potent biocontrol agent isolated from rice rhizosphere, as it prevents bacterial rice seedling blight disease caused by Burkholderia plantarii. Here, we isolated a non-antibacterial metabolite from the culture fluid of B. heleia PAK1-2 that was able to suppress B. plantarii virulence and subsequently identified as indole-3-acetic acid (IAA). IAA suppressed the production of tropolone in B. plantarii in a dose-dependent manner without any antibacterial and quorum quenching activity, suggesting that IAA inhibited steps of tropolone biosynthesis. Consistent with this, supplementing cultures of B. plantarii with either L-[ring-2H5]phenylalanine or [ring-2H2~5]phenylacetic acid revealed that phenylacetic acid (PAA), which is the dominant metabolite during the early growth stage, is a direct precursor of tropolone. Exposure of B. plantarii to IAA suppressed production of both PAA and tropolone. These data particularly showed that IAA produced by B. heleia PAK1-2 disrupts tropolone production during bioconversion of PAA to tropolone via the ring-rearrangement on the phenyl group of the precursor to attenuate the virulence of B. plantarii. B. heleia PAK1-2 is thus likely a microbial community coordinating bacterium in rhizosphere ecosystems, which never eliminates phytopathogens but only represses production of phytotoxins or bacteriocidal substances. PMID:26935539

  2. ENVIRONMENTAL TECHNOLOGY VERIFICATON: TEST REPORT OF CONTROL OF BIOAEROSOLS IN HVAC SYSTEMS AAF INTERNATIONAL DRIPAK 90/95%

    EPA Science Inventory

    The Environmental Technology Verification report discusses the technology and performance of the DriPak 90/95% air filter for dust and bioaerosol filtration manufactured by AAF International. The pressure drop across the filter was 104 Pa clean and 348 Pa dust loaded, and the fil...

  3. Field application of serodiagnostics to identify elephants with Tuberculosis prior to case confirmation by culture

    USDA-ARS?s Scientific Manuscript database

    Three serologic methods for antibody detection in elephant tuberculosis (TB), multiantigen print immunoassay (MAPIA), ElephantTB STAT-PAK kit, and DPP VetTB test, were validated prospectively using serial serum samples from 14 captive elephants in 5 countries which were diagnosed with TB by positive...

  4. A hippocampal Cdk5 pathway regulates extinction of contextual fear

    PubMed Central

    Sananbenesi, Farahnaz; Fischer, Andre; Wang, Xinyu; Schrick, Christina; Neve, Rachael; Radulovic, Jelena; Tsai, Li-Huei

    2008-01-01

    Treatment of emotional disorders involves the promotion of extinction processes, which are defined as the learned reduction of fear. The molecular mechanisms underlying extinction have only begun to be elucidated. By employing genetic and pharmacological approaches in mice, we show here that extinction requires downregulation of Rac-1 and cyclin-dependent kinase 5 (Cdk5), and upregulation of p21 activated kinase-1 (PAK-1) activity. This is physiologically achieved by a Rac-1–dependent relocation of the Cdk5 activator p35 from the membrane to the cytosol and dissociation of p35 from PAK-1. Moreover, our data suggest that Cdk5/p35 activity prevents extinction in part by inhibition of PAK-1 activity in a Rac-1–dependent manner. We propose that extinction of contextual fear is regulated by counteracting components of a molecular pathway involving Rac-1, Cdk5 and PAK-1. Our data suggest that this pathway could provide a suitable target for therapeutic treatment of emotional disorders. PMID:17632506

  5. Inhibition of Tulane Virus replication via exposure to Lowbush Blueberry (Vaccinium angustifolium) fractional components

    USDA-ARS?s Scientific Manuscript database

    Tulane Virus (TV) is a common viral surrogate for human norovirus in lab studies. In the present study, the phenotypic response of TV when exposed to fractional components extracted from lowbush blueberries was investigated. Lowbush blueberry extract (F1) was separated using a C-18 Sep-Pak cartridge...

  6. 79 FR 43923 - Approved Tests for Bovine Tuberculosis in Cervids

    Federal Register 2010, 2011, 2012, 2013, 2014

    2014-07-29

    ... Animal and Plant Health Inspection Service 9 CFR Part 77 Approved Tests for Bovine Tuberculosis in... comments. SUMMARY: We are amending the regulations regarding official tuberculosis tests for captive cervids to remove the CervidTB Stat- Pak as an official bovine tuberculosis test for the following...

  7. The Genus Culex, Subgenus Eumelanomyia Theobald in Southeast Asia and Adjacent Areas

    DTIC Science & Technology

    1972-01-01

    tergal surface; subapical lobe short or elon or sma 1; distimere slender, sickle shaped and long; B ate; proximal division with 3 stout rods...Nok, 1 P. Tak: Huey Lan Saeng, 6d, 69, 12 p. Nakhon Ratchasima: Pak Chong; Musk Lek; Ban Tha Ma Prang; Khlong Pai; Khao Suan Horn; 7~r, 9?, 1 P, 7 p

  8. Surface Water Investigations in Afghanistan: A Summary of Activities from 1952 to 1969

    DTIC Science & Technology

    1969-03-01

    Chakhansur Registan Spin Baldak Chaman Zahedran Gizab Qala-Hazar Qadam Qala Ahangaran Deshu Sistan Basin Adraskan Shindand Kajaki (32°l6-65...Chakhansur Rigestan(region) Spin Buidak Chaman (Pak) Zahedan (Iran) Gizab Hazar Qadam Ahangaran Deh Shu Seistan(region

  9. Complete genome sequence of a recent panzootic virulent Newcastle disease virus from Pakistan

    USDA-ARS?s Scientific Manuscript database

    Complete genome sequence of a new strain of Newcastle disease virus (NDV) (chicken/Pak/Lahore-611/2013) is reported. The strain was isolated from a vaccinated chicken flock in Pakistan in 2013 and has panzootic features. The genome is 15192 nucleotides in length and is classified as sub-genotype V...

  10. [Enantioseparation behavior of chiral stationary phases AD, AS and OD].

    PubMed

    Li, Liqun; Fan, Jun; Zhang, Jing; Chen, Xiaodong; Wang, Tai; He, Jianfeng; Zhang, Weiguang

    2016-01-01

    Over the past decades, HPLC enantioseparation with chiral stationary phases (CSPs) has been widely applied in chiral analysis and preparation of new pharmaceuticals, pesticides, food, etc. Herein, enantioseparation of 20 chiral compounds have been carried out on three polysaccharide-based CSPs (EnantioPak AD, AS and OD) with normal phases by HPLC, separately. The influences of skeletal structure and the kinds of derivative groups on separation behaviors of these CSPs have been studied in detail. As results indicated, except for compound 13, the other compounds were baseline separated on EnantioPak AD, with most of resolution over 2. 0; in addition, better separation for acidic or basic compounds was achieved through adding acidic/basic additives into the mobile phase of hexane-alcohol. For four aromatic alcohols (compounds 13-16), their retention in the EnantioPak AD column showed a weakening tendency with increase of carbon number in side chain group, and the reverse trend of their resolution was observed. Furthermore, EnantioPak AD showed much better separation performance for eight compounds (13-20) than the others. In short, these results have provided some references for further investigation of separation behavior and applications of polysaccharide-based CSPs.

  11. UnPAKing RUNX3 functions-Both sides of the coin.

    PubMed

    Kumar, Arun; Sundaram, Sandhya; Rayala, Suresh K; Venkatraman, Ganesh

    2017-06-19

    Post translational modifications of RUNX3 have been shown to play an important role in directing RUNX3 functions. In this review we highlight the phosphorylation dependent functions of RUNX3 as regulated by PAK1 and its implications on tumorigenesis.

  12. Area Handbook Series: Thailand. A Country Study

    DTIC Science & Technology

    1987-09-01

    Paleolithic culture in the region and continuous human habitation for at least 20,000 years. The pace of economic and social development was uneven and...Pakistanis, 107 National Security Council, 192, 255 Pak Mai (New Party), 222 National Socialist Party, 37 Paleolithic culture, 4 National Union (Sahachat

  13. The Impact of Chinese Development of Nuclear Weapons on the Pakistan-Indian Dispute

    DTIC Science & Technology

    1966-04-08

    Conference of Dec. 10, 1962." Dept of State Bulletin, Vol. 47, 31 Dec. 1962, p. 998. (Statement of recent US efforts on Kashmir.) 65. Salam , Abdus ...34 RoyaJ Central Asaln Journal, Viii. 51, Oct. 1964, p. 223. 39 Abdus Sal am, !Pak i sLa I- -The Case for Tecict I Cal leve I plmielt , BHlleLtln of

  14. Actions of Rho family small G proteins and p21-activated protein kinases on mitogen-activated protein kinase family members.

    PubMed Central

    Frost, J A; Xu, S; Hutchison, M R; Marcus, S; Cobb, M H

    1996-01-01

    The mitogen-activated protein (MAP) kinases are a family of serine/threonine kinases that are regulated by distinct extracellular stimuli. The currently known members include extracellular signal-regulated protein kinase 1 (ERK1), ERK2, the c-Jun N-terminal kinase/stress-activated protein kinases (JNK/SAPKs), and p38 MAP kinases. We find that overexpression of the Ste20-related enzymes p21-activated kinase 1 (PAK1) and PAK2 in 293 cells is sufficient to activate JNK/SAPK and to a lesser extent p38 MAP kinase but not ERK2. Rat MAP/ERK kinase kinase 1 can stimulate the activity of each of these MAP kinases. Although neither activated Rac nor the PAKs stimulate ERK2 activity, overexpression of either dominant negative Rac2 or the N-terminal regulatory domain of PAK1 inhibits Ras-mediated activation of ERK2, suggesting a permissive role for Rac in the control of the ERK pathway. Furthermore, constitutively active Rac2, Cdc42hs, and RhoA synergize with an activated form of Raf to increase ERK2 activity. These findings reveal a previously unrecognized connection between Rho family small G proteins and the ERK pathway. PMID:8668187

  15. Wetlands systems in southern Thailand: The essential resources for sustainable regional development

    Treesearch

    Rotchanatch Darnsawasdi; Prassert Chitpong

    2000-01-01

    Parts of Southern Thailand are inundated by water for months annually resulting in various wetlands including, among others, Tapi River Basin, Pak Panang River Basin, Songkhla Lake Basin, Pangnga Bay, Pattani River Basin, and Narathiwas Peat Swamp. Most wetlands perform functions such as flood retention, water filtration, bird and wildlife habitat,and tree growth....

  16. Over 500 pancreas transplants by a single team in São Paulo, Brazil.

    PubMed

    Perosa, Marcelo; Crescentini, Fabio; Noujaim, Huda; Mota, Leonardo T; Branez, Juan Rafael; Ianhez, Luiz Estevam; Ferreira, Gustavo; de Oliveira, Rodrigo Azevedo; Genzini, Tércio

    2011-01-01

    Pancreas transplantation (PT) remains a developing practice in Latin America. From 1996 to 2009, 506 PTs were performed by our team in the following categories: simultaneous pancreas-kidney (SPK), simultaneous deceased donor pancreas and living-donor kidney (SPLK), pancreas after kidney (PAK), and pancreas transplant alone (PTA). Enteric drainage was preferred for SPK and bladder drainage for solitary PT or SPLK. Immunosuppression was with tacrolimus, mycophenolate mofetil, and steroids, and anti-lymphocytic drugs were used to induce solitary PT and SPLK. The series includes 254 SPK, 60 SPLK, 94 PAK, and 98 PTA. The one-yr patient survivals were 82% for SPK, 90% for SPLK, 95% for PTA, and 93% for PAK. The one-yr pancreas graft survivals were 70% for SPK, 86% for SPLK, 86% for PAK, and 77% for PTA. The one-yr kidney graft survivals were 77.5% for SPK and 89% for SPLK. This represents the largest reported PT series in Latin America. Results comparable to those of developed countries were achieved, with the exception of the SPK category. This has led our program to prioritize solitary PT and SPLK. © 2011 John Wiley & Sons A/S.

  17. Simple method for determination of patulin production by Penicillium griseofulvum Dierckx.

    PubMed Central

    Torres, M; Sanchis, V; Riba, M; Canela, R

    1986-01-01

    Patulin production by Penicillium griseofulvum was monitored with Sep-Pak cartridges and high-pressure liquid chromatography. Determination and quantification of this metabolite proved to be very simple, and our method saved time and a large amount of organic solvents. PMID:3513700

  18. Postmortem Photonic Imaging of Lux-Modified Salmonella Typhimuium Within the Gastrointestinal Tract of Swine Following Oral Inoculation In Vivo

    USDA-ARS?s Scientific Manuscript database

    The study objective was to monitor Salmonella progression by photonic detection through segments of the gastrointestinal tract after oral inoculation. Pigs (~80 kg) were inoculated orally with 3.1 or 4.1 x 1010 cfu of Salmonella Typhimurium transformed with plasmid pAK1-lux for a 6-h (n = 6) or 12-h...

  19. Postmortem photonic imaging of lux-modified Salmonella typhimurium within the gastrointestinal tract of swine following oral inoculation in vivo

    USDA-ARS?s Scientific Manuscript database

    The study objective was to monitor Salmonella progression by photonic detection through segments of the gastrointestinal tract following oral inoculation. Pigs (~ 80 kg) were inoculated orally with 3.1 or 4.1×10*10 colony forming units (cfu) of Salmonella typhimurium transformed with plasmid pAK1-lu...

  20. The A. Q. Khan Network: Causes and Implications

    DTIC Science & Technology

    2005-12-01

    Reprocessing Plant,” [Maulana Kausar Niazi , Aur Line Kat Gayee, trans. by Samuel Baid (Pak.: 1987), chapter 9,] in Pakistan’s Bomb, 358-362. 58 At...official,” Dawn (Karachi), February 5, 2004, http://www.dawn.com/2004/02/05/top5.htm. 128 See Niazi , “Unknown Facts about the Reprocessing Plant

  1. The effects of control release fertilizer (CRF) on palm growth

    USDA-ARS?s Scientific Manuscript database

    Nutri-Pak is a slow release fertilizer in a micro-pore polyethylene packet where moisture enters the packet through micro-pores located on both sides of the packet. Water dissolves the fertilizer and it slowly seeps through the same micro-pores as a vapor into the soil gradually providing nutrients ...

  2. A novel p21-activated kinase binds the actin and microtubule networks and induces microtubule stabilization

    PubMed Central

    Cau, Julien; Faure, Sandrine; Comps, Michel; Delsert, Claude; Morin, Nathalie

    2001-01-01

    Coordination of the different cytoskeleton networks in the cell is of central importance for morphogenesis, organelle transport, and motility. The Rho family proteins are well characterized for their effects on the actin cytoskeleton, but increasing evidence indicates that they may also control microtubule (MT) dynamics. Here, we demonstrate that a novel Cdc42/Rac effector, X-p21-activated kinase (PAK)5, colocalizes and binds to both the actin and MT networks and that its subcellular localization is regulated during cell cycle progression. In transfected cells, X-PAK5 promotes the formation of stabilized MTs that are associated in bundles and interferes with MTs dynamics, slowing both the elongation and shrinkage rates and inducing long paused periods. X-PAK5 subcellular localization is regulated tightly, since coexpression with active Rac or Cdc42 induces its shuttling to actin-rich structures. Thus, X-PAK5 is a novel MT-associated protein that may communicate between the actin and MT networks during cellular responses to environmental conditions. PMID:11733543

  3. Regulation of microtubule destabilizing activity of Op18/stathmin downstream of Rac1.

    PubMed

    Wittmann, Torsten; Bokoch, Gary M; Waterman-Storer, Clare M

    2004-02-13

    In the leading edge of migrating cells, a subset of microtubules exhibits net growth in a Rac1- and p21-activated kinase-dependent manner. Here, we explore the possibility of whether phosphorylation and inactivation of the microtubule-destabilizing protein Op18/stathmin could be a mechanism regulating microtubule dynamics downstream of Rac1 and p21-activated kinases. We find that, in vitro, Pak1 phosphorylates Op18/stathmin specifically at serine 16 and inactivates its catastrophe promoting activity in biochemical and time lapse microscopy microtubule assembly assays. Furthermore, phosphorylation of either serine 16 or 63 is sufficient to inhibit Op18/stathmin in vitro. In cells, the microtubule-destabilizing effect of an excess of Op18/stathmin can be partially overcome by expression of constitutively active Rac1(Q61L), which is dependent on Pak activity, suggesting that the microtubule cytoskeleton can be regulated through inactivation of Op18/stathmin downstream of Rac1 and Pak in vivo. However, in vivo, Pak1 activity alone is not sufficient to phosphorylate Op18, indicating that additional pathways downstream of Rac1 are required for Op18 regulation.

  4. From LAMP to Koha: Case Study of the Pakistan Legislative Assembly Libraries

    ERIC Educational Resources Information Center

    Shafi-Ullah, Farasat; Qutab, Saima

    2012-01-01

    Purpose: This paper aims to elaborate the library data migration process from LAMP (Library Automation Management Program) to the open source software Koha's (2.2.8 Windows based) Pakistani flavour PakLAG-Koha in six legislative assembly libraries of Pakistan. Design/methodology/approach: The paper explains different steps of the data migration…

  5. From LAMP to Koha: Case Study of the Pakistan Legislative Assembly Libraries

    ERIC Educational Resources Information Center

    Shafi-Ullah, Farasat; Qutab, Saima

    2012-01-01

    Purpose: This paper aims to elaborate the library data migration process from LAMP (Library Automation Management Program) to the open source software Koha's (2.2.8 Windows based) Pakistani flavour PakLAG-Koha in six legislative assembly libraries of Pakistan. Design/methodology/approach: The paper explains different steps of the data migration…

  6. An Analysis of the Fulton Industrial Area: A Market for Parkway’s Industrial Medicine Program.

    DTIC Science & Technology

    1983-05-30

    are not waiting for proof of the hypothesis related to the benefits of health promotion programs. Many accept the empirical evidence of employee...offered at Tee-Pak. *Interviewee did not desire to respond to the benefits of health promotion programs, because of such programs must originate from the

  7. ENVIRONMENTAL TECHNOLOGY VERIFICATON: TEST REPORT OF CONTROL OF BIOAEROSOLS IN HVAC SYSTEMS AAF INTERNATIONAL DRIPAK 90/95%

    EPA Science Inventory

    The Environmental Technology Verification report discusses the technology and performance of the DriPak 90/95% air filter for dust and bioaerosol filtration manufactured by AAF International. The pressure drop across the filter was 104 Pa clean and 348 Pa dust loaded, and the fil...

  8. 78 FR 13139 - Additional Designation of A North Korean Entity and Two North Korean Individuals Pursuant to...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-26

    ....A. DPRK Committee for Space Technology A.K.A. Department of Space Technology of North Korea A.K.A. Committee for Space Technology A.K.A. KCST Location: Pyongyang, North Korea Paek Chang-Ho A.K.A. Pak Chang... Additional Designation of A North Korean Entity and Two North Korean Individuals Pursuant to Executive Order...

  9. Disruption of PKB signaling restores polarity to cells lacking tumor suppressor PTEN

    PubMed Central

    Tang, Ming; Iijima, Miho; Kamimura, Yoichiro; Chen, Lingfeng; Long, Yu; Devreotes, Peter

    2011-01-01

    By limiting phosphotidylinositol 3,4,5-triphosphate (PIP3) levels, tumor suppressor PTEN not only controls cell growth but also maintains cell polarity required for cytokinesis and chemotaxis. To identify the critical targets of PIP3 that link it to the cytoskeleton, we deleted secondary genes to reverse the deficiencies of pten- cells in Dictyostelium. The polarity defects in pten- cells correlate with elevated phosphorylations of PKB substrates. Deletion of AKT orthologue, PkbA, or a subunit of its activator TORC2, reduced the phosphorylations and suppressed the cytokinesis and chemotaxis defects in pten- cells. In these double mutants, the excessive PIP3 levels and, presumably, activation of other PIP3-binding proteins had little or no effect on the cytoskeleton. In bands with increased phosphorylation in pten- cells, we found PKB substrates, PI5K, GefS, GacG, and PakA. Disruption of PakA in pten- cells restored a large fraction of the cells to normal behavior. Consistently, expression of phosphomimetic PakA in pten- cells exacerbated the defects but nonphosphorylatable PakA had no effect. Thus, among many putative PTEN- and PIP3-dependent events, phosphorylation of PKB substrates is the key downstream regulator of cell polarity. PMID:21169559

  10. Genetic and Functional Diversity of Human Immunodeficiency Virus Type 1 Subtype B Nef Primary Isolates

    PubMed Central

    Foster, John L.; Molina, Rene P.; Luo, Tianci; Arora, Vivek K.; Huang, Yaoxing; Ho, David D.; Garcia, J. Victor

    2001-01-01

    We have characterized the functional integrity of seven primary Nef isolates: five from a long-term nonprogressing human immunodeficiency virus (HIV)-infected individual and one each from two patients with AIDS. One of the seven Nefs was defective for CD4 downregulation, two others were defective for PAK-2 activation, and one Nef was defective for PAK-2 activation and major histocompatibility complex (MHC) class I downregulation. Five of the Nefs were tested and found to be functional for the enhancement of virus particle infectivity. The structural basis for each of the functional defects has been analyzed by constructing a consensus nef, followed by mutational analysis of the variant amino acid residues. Mutations A29V and F193I were deleterious to CD4 downregulation and PAK-2 activation, respectively, while S189R rendered Nef defective for both MHC class I downregulation and PAK-2 activation. A search of the literature identified HIVs from five patients with Nefs predominantly mutated at F193 and from one patient with Nefs predominantly mutated at A29. A29 is highly conserved in all HIV subtypes except for subtype E. F193 is conserved in subtype B (and possibly in the closely related subtype D), but none of the other HIV group M subtypes. Our results suggest that functional distinctions may exist between HIV subtypes. PMID:11160665

  11. Psychoanalysis and Transformation of Heroes in Mohsin Hamid's Novels "Moth Smoke" and "The Reluctant Fundamentalist"

    ERIC Educational Resources Information Center

    Awan, Abdul Ghafoor; Andleeb, Shaista; Yasin, Farhat

    2016-01-01

    Mohsin Hamid is an exponent of postcolonial characterization and possesses a specific touch of today's hero in local Asian context. His pen is fluent on social fiction portraying Indo-Pak culture. Few writers could rise to the height of fame right with only a couple of preliminary works. Hamid secured it through his first novel "Moth…

  12. Working with Children to Protect Our Future

    ERIC Educational Resources Information Center

    Williams, Nick

    2011-01-01

    The author has been teaching primary school children for 20 years, and has always been passionate about teaching young people about the environment. In this article, he describes his work with Tetra Pak and WWF-UK to develop a national, school-based competition and teaching programme to help children understand the importance of using renewable…

  13. A Neural Network for Estimation of Aortic Pressure from the Radial Artery Pressure Pulse

    DTIC Science & Technology

    2001-10-25

    from periphery to artery: a model based study, American Journal of Physiology, 1998,274:43, pp H1386-92 [9] C. Chen, E. Nevo , B Fetics, P Pak, F, Yin, L...36. [10] B Fetics, E Nevo , C. Chen, D Kass, Parametric model derivation of transfer function for noninvasive estimation of aortic pressure by radial

  14. Identification of the regulatory autophosphorylation site of autophosphorylation-dependent protein kinase (auto-kinase). Evidence that auto-kinase belongs to a member of the p21-activated kinase family.

    PubMed

    Yu, J S; Chen, W J; Ni, M H; Chan, W H; Yang, S D

    1998-08-15

    Autophosphorylation-dependent protein kinase (auto-kinase) was identified from pig brain and liver on the basis of its unique autophosphorylation/activation property [Yang, Fong, Yu and Liu (1987) J. Biol. Chem. 262, 7034-7040; Yang, Chang and Soderling (1987) J. Biol. Chem. 262, 9421-9427]. Its substrate consensus sequence motif was determined as being -R-X-(X)-S*/T*-X3-S/T-. To characterize auto-kinase further, we partly sequenced the kinase purified from pig liver. The N-terminal sequence (VDGGAKTSDKQKKKAXMTDE) and two internal peptide sequences (EKLRTIV and LQNPEK/ILTP/FI) of auto-kinase were obtained. These sequences identify auto-kinase as a C-terminal catalytic fragment of p21-activated protein kinase 2 (PAK2 or gamma-PAK) lacking its N-terminal regulatory region. Auto-kinase can be recognized by an antibody raised against the C-terminal peptide of human PAK2 by immunoblotting. Furthermore the autophosphorylation site sequence of auto-kinase was successfully predicted on the basis of its substrate consensus sequence motif and the known PAK2 sequence, and was further demonstrated to be RST(P)MVGTPYWMAPEVVTR by phosphoamino acid analysis, manual Edman degradation and phosphopeptide mapping via the help of phosphorylation site analysis of a synthetic peptide corresponding to the sequence of PAK2 from residues 396 to 418. During the activation process, auto-kinase autophosphorylates mainly on a single threonine residue Thr402 (according to the sequence numbering of human PAK2). In addition, a phospho-specific antibody against a synthetic phosphopeptide containing this identified sequence was generated and shown to be able to differentially recognize the activated auto-kinase autophosphorylated at Thr402 but not the non-phosphorylated/inactive auto-kinase. Immunoblot analysis with this phospho-specific antibody further revealed that the change in phosphorylation level of Thr402 of auto-kinase was well correlated with the activity change of the kinase during both

  15. p85 beta-PIX is required for cell motility through phosphorylations of focal adhesion kinase and p38 MAP kinase.

    PubMed

    Lee, Jangsoon; Jung, In Duk; Chang, Won Keun; Park, Chang Gyo; Cho, Do Yeun; Shin, Eun-Young; Seo, Dong Wan; Kim, Yong Kee; Lee, Hyang Woo; Han, Jeung-Whan; Lee, Hoi Young

    2005-07-15

    Lysophosphatidic acid (LPA) mediates diverse biological responses, including cell migration, through the activation of G-protein-coupled receptors. Recently, we have shown that LPA stimulates p21-activated kinase (PAK) that is critical for focal adhesion kinase (FAK) phosphorylation and cell motility. Here, we provide the direct evidence that p85 beta-PIX is required for cell motility of NIH-3T3 cells by LPA through FAK and p38 MAP kinase phosphorylations. LPA induced p85 beta-PIX binding to FAK in NIH-3T3 cells that was inhibited by pretreatment of the cells with phosphoinositide 3-kinase inhibitor, LY294002. Furthermore, the similar inhibition of the complex formation was also observed, when the cells were transfected with either p85 beta-PIX mutant that cannot bind GIT or dominant negative mutants of Rac1 (N17Rac1) and PAK (PAK-PID). Transfection of the cells with specific p85 beta-PIX siRNA led to drastic inhibition of LPA-induced FAK phosphorylation, peripheral redistribution of p85 beta-PIX with FAK and GIT1, and cell motility. p85 beta-PIX was also required for p38 MAP kinase phosphorylation induced by LPA. Finally, dominant negative mutant of Rho (N19Rho)-transfected cells did not affect PAK activation, while the cells stably transfected with p85 beta-PIX siRNA or N17Rac1 showed the reduction of LPA-induced PAK activation. Taken together, the present data suggest that p85 beta-PIX, located downstream of Rac1, is a key regulator for the activations of FAK or p38 MAP kinase and plays a pivotal role in focal complex formation and cell motility induced by LPA.

  16. Interaction between the Helicobacter pylori CagA and alpha-Pix in gastric epithelial AGS cells.

    PubMed

    Baek, Hye Yeon; Lim, Joo Weon; Kim, Hyeyoung

    2007-01-01

    The gastric pathogen Helicobacter pylori (H. pylori) translocates the CagA protein into epithelial cells by a type IV secretion process. Upon translocation into the host cell cytosol, CagA undergoes tyrosine phosphorylation. Phosphorylation of CagA occurs within the C terminus of the protein and is mediated by members of the Src family of tyrosine kinases. Phosphorylation of CagA induces the dephosphorylation of as yet unidentified cellular proteins, rearrangements of the host cell actin cytoskeleton, and cell scattering. This article aims to determine the cellular protein that interacts with CagA. Gastric epithelial AGS cells were stimulated with CagA-positive H. pylori (NCTC11637, at a bacteria/cell ratio of 500:1) and cultured in antibiotic-free medium. Proteins were isolated from the cells with or without H. pylori infection. CagA-interactive protein was determined by immunoprecipitation using anti-CagA antibody and proteomic analysis. We found that alpha-Pix interacts with CagA and alpha-Pix was constitutively expressed in AGS cells. Upon H. pylori stimulation, CagA was translocated into the cells and the expression of alpha-Pix (PAK-interactive exchange factor) was increased in AGS cells time dependently. The interaction of alpha-Pix with CagA was increased by H. pylori infection in AGS cells. Phosphorylation of CagA induces the dephosphorylation of alpha-Pix in AGS cells. alpha-Pix is a family of PAK-binding proteins that strongly activates PAK (p21-activated tyrosine kinase). PAK regulates changes in gene expression and mediates actin cytoskeletal and cell morphological changes. The novel finding of this study is that phosphorylation of CagA induces the dephosphorylation of alpha-Pix, which may modulate cytoskeletal changes of gastric epithelial cells through PAK.

  17. Integrated analysis of breast cancer cell lines reveals unique signaling pathways

    SciTech Connect

    Heiser, Laura M.; Wang, Nicholas J.; Talcott, Carolyn L.; Laderoute, Keith R.; Knapp, Merrill; Guan, Yinghui; Hu, Zhi; Ziyad, Safiyyah; Weber, Barbara L.; Laquerre, Sylvie; Jackson, Jeffrey R.; Wooster, Richard F.; Kuo, Wen-Lin; Gray, Joe W.; Spellman, Paul T.

    2009-03-31

    Cancer is a heterogeneous disease resulting from the accumulation of genetic defects that negatively impact control of cell division, motility, adhesion and apoptosis. Deregulation in signaling along the EGFR-MAPK pathway is common in breast cancer, though the manner in which deregulation occurs varies between both individuals and cancer subtypes. We were interested in identifying subnetworks within the EGFR-MAPK pathway that are similarly deregulated across subsets of breast cancers. To that end, we mapped genomic, transcriptional and proteomic profiles for 30 breast cancer cell lines onto a curated Pathway Logic symbolic systems model of EGFR-MEK signaling. This model was comprised of 539 molecular states and 396 rules governing signaling between active states. We analyzed these models and identified several subtype specific subnetworks, including one that suggested PAK1 is particularly important in regulating the MAPK cascade when it is over-expressed. We hypothesized that PAK1 overexpressing cell lines would have increased sensitivity to MEK inhibitors. We tested this experimentally by measuring quantitative responses of 20 breast cancer cell lines to three MEK inhibitors. We found that PAK1 over-expressing luminal breast cancer cell lines are significantly more sensitive to MEK inhibition as compared to those that express PAK1 at low levels. This indicates that PAK1 over-expression may be a useful clinical marker to identify patient populations that may be sensitive to MEK inhibitors. All together, our results support the utility of symbolic system biology models for identification of therapeutic approaches that will be effective against breast cancer subsets.

  18. Symptom Assessment in Knee Osteoarthritis Needs to Account for Physical Activity Level

    PubMed Central

    Lo, Grace H.; McAlindon, Timothy E.; Hawker, Gillian A.; Driban, Jeffrey B.; Price, Lori Lyn; Song, Jing; Eaton, Charles B.; Hochberg, Marc C.; Jackson, Rebecca D.; Kwoh, C. Kent; Nevitt, Michael C.; Dunlop, Dorothy D.

    2015-01-01

    Objective Pain is not always correlated with radiographic osteoarthritis (OA) severity possibly because people modify activities to manage symptoms. Measures of symptoms that consider pain in the context of activity level may therefore provide greater discrimination than pain alone. Our objective was to compare discrimination of a measure of pain alone with combined measures of pain relative to physical activity across radiographic OA levels. Methods This is a cross-sectional study of the Osteoarthritis Initiative accelerometer substudy, including those with and without knee OA. Two composite pain and activity knee symptom (PAKS) scores were calculated as Western Ontario and McMaster (WOMAC) Universities Osteoarthritis Pain Scale plus one divided by physical activity measures (step and activity counts). Symptom score discrimination across Kellgren and Lawrence (KL) grades were evaluated using histograms and quantile regression. Results 1806 participants, mean age 65.1 (9.1) years, mean BMI 28.4 (4.8) kg/m2, and 55.6% female, were included. WOMAC, but not PAKS scores, exhibited a floor effect. Adjusted median WOMAC by KL grades 0 – 4 were 0, 0, 1, 1, and 3 respectively. Median PAKS1 and PAKS2 were 24.9, 26.0, 32.4, 46.1, 97.9, and 7.2, 7.2, 9.2, 12.9, 23.8, respectively. PAKS scores had more statistically significant comparisons between KL grades compared with WOMAC. Conclusions Symptom assessments incorporating pain and physical activity did not exhibit a floor effect and were better able to discriminate radiographic severity than pain alone, particularly in milder disease. Pain in the context of physical activity level should be used to assess knee OA symptoms. PMID:26407008

  19. Integrated analysis of breast cancer cell lines reveals unique signaling pathways

    PubMed Central

    Heiser, Laura M; Wang, Nicholas J; Talcott, Carolyn L; Laderoute, Keith R; Knapp, Merrill; Guan, Yinghui; Hu, Zhi; Ziyad, Safiyyah; Weber, Barbara L; Laquerre, Sylvie; Jackson, Jeffrey R; Wooster, Richard F; Kuo, Wen Lin; Gray, Joe W; Spellman, Paul T

    2009-01-01

    Background Cancer is a heterogeneous disease resulting from the accumulation of genetic defects that negatively impact control of cell division, motility, adhesion and apoptosis. Deregulation in signaling along the EgfR-MAPK pathway is common in breast cancer, though the manner in which deregulation occurs varies between both individuals and cancer subtypes. Results We were interested in identifying subnetworks within the EgfR-MAPK pathway that are similarly deregulated across subsets of breast cancers. To that end, we mapped genomic, transcriptional and proteomic profiles for 30 breast cancer cell lines onto a curated Pathway Logic symbolic systems model of EgfR-MAPK signaling. This model was composed of 539 molecular states and 396 rules governing signaling between active states. We analyzed these models and identified several subtype-specific subnetworks, including one that suggested Pak1 is particularly important in regulating the MAPK cascade when it is over-expressed. We hypothesized that Pak1 over-expressing cell lines would have increased sensitivity to Mek inhibitors. We tested this experimentally by measuring quantitative responses of 20 breast cancer cell lines to three Mek inhibitors. We found that Pak1 over-expressing luminal breast cancer cell lines are significantly more sensitive to Mek inhibition compared to those that express Pak1 at low levels. This indicates that Pak1 over-expression may be a useful clinical marker to identify patient populations that may be sensitive to Mek inhibitors. Conclusions All together, our results support the utility of symbolic system biology models for identification of therapeutic approaches that will be effective against breast cancer subsets. PMID:19317917

  20. Sensitivity, specificity, and confounding factors of novel serological tests used for the rapid diagnosis of bovine tuberculosis in farmed red deer (Cervus elaphus).

    PubMed

    Buddle, Bryce M; Wilson, Tania; Denis, Michel; Greenwald, Rena; Esfandiari, Javan; Lyashchenko, Konstantin P; Liggett, Simon; Mackintosh, Colin G

    2010-04-01

    In this study, novel serological tests were used to detect tuberculosis (TB) in groups of farmed red deer (Cervus elaphus) varying in disease status or possible confounding factors. Groups of deer naturally or experimentally infected with Mycobacterium bovis and animals vaccinated against paratuberculosis were studied, as were uninfected animals and animals naturally or experimentally infected with Mycobacterium avium subsp. paratuberculosis. Sera were assayed using two rapid lateral-flow tests, Chembio's CervidTB STAT-PAK and DPP VetTB tests, and results were compared to those from tuberculin skin tests. Both serological tests had a high sensitivity, but specificity was adversely affected after animals had received a vaccine against paratuberculosis and were subsequently skin tested. The specificity of the DPP VetTB test was higher than that of the CervidTB STAT-PAK test, with natural infection with M. avium subsp. paratuberculosis adversely affecting the specificity of only the CervidTB STAT-PAK test. The sera from M. avium subsp. paratuberculosis-infected deer that produced false-positive reactions in the CervidTB STAT-PAK test were retested with a multiantigen print immunoassay (MAPIA), and some of these sera were shown to react with the MPB83 antigen. Combining the results from the serological tests and the skin tests showed only a slight increase in the sensitivity of detection of M. bovis-infected animals. It is concluded that both the CervidTB STAT-PAK and DPP VetTB tests offer rapid, convenient, and easy detection of bovine tuberculosis in deer, albeit with significant interference from paratuberculosis vaccination status and subsequent skin testing. The latter finding illustrates one of the limitations of currently available vaccines against paratuberculosis.

  1. Pancreas transplant outcomes for United States (US) and non-US cases as reported to the United Network for Organ Sharing (UNOS) and the International Pancreas Transplant Registry (IPTR) as of June 2004.

    PubMed

    Gruessner, Angelika C; Sutherland, David E R

    2005-08-01

    As of December 31, 2004, more than 23,000 pancreas transplant had been reported to the IPTR, >17,000 in the US and almost 6000 from outside the US. An analysis of US pancreas transplants performed between 1988 and 2003 showed a progressive improvement in outcome, with pancreas transplant graft survival rates (GSRs) going from 75% at 1 yr for 1988/1989 to 85% for 2002/2003 simultaneous pancreas-kidney (SPK) cases, from 55 to 78% for pancreas after kidney (PAK) cases, and from 45 to 77% for pancreas transplants alone (PTA) cases. The improvements were due both to decreases in technical failure (TF) rates (from 12 to 6% in SPK, 13-8% in PAK, and 24-7% in PTA) and immunological failure rates (going from 7 to 2% for SPK, from 28 to 7% for PAK, and from 38 to 8% for PTA cases). These results are even more impressive under the aspect that during the same time the rate of potential risk factors increased and the duct management techniques changed from bladder to enteric drainage. The improvement in outcome allowed also an increase in the number of solitary pancreas transplants from initially 12% to now 35%. Contemporary primary deceased donor pancreas transplant outcomes were calculated separately for 2000-2004 US and non-US cases. The US patient survival rates at 1 yr were >95% in each recipient category, with 1 yr primary pancreas GSRs of 85% for SPK, 78% for PAK, and 76% for PTA (p < 0.0001). The immunological graft failure rates for 2000-2004 technically successful (TS) SPK, PAK, and PTA cases were 2, 8, and 10% at 1 yr (p = 0.0001). In the majority of all transplants ED was used for duct management (81% of SPK, 67% of PAK, and 56% for PTA cases). Of the ED transplants, venous drainage via the portal system was used for 20% of SPK, 23% of PAK, and 35% of PTA cases. Duct management technique did not have a significant impact on overall pancreas graft function in the univariate or the multivariate model. The outcomes of ED and BD transplants are comparable with 85 vs. 87

  2. Die Kometenmission Rosetta

    NASA Astrophysics Data System (ADS)

    Krüger, Harald

    2016-11-01

    Die Rosetta-Mission ist ein Meilenstein in der Erforschung der Kometen und ihrer Entstehung. Eine der größten üerraschungen war die unregelmäßge hantelförmige Gestalt des Zielkometen 67P/Tschurjumow-Gerassimenko. Er besteht wahrscheinlich aus zwei Einzelkörpern, die durch ihre Schwerkraft aneinander gehalten werden. Seine Oberfläche ist sehr rau und zeigt eine sehr vielf ältige Morphologie, die auf eine Vielzahl von ablaufenden Prozessen hindeutet. Der Kometenkern ist vermutlich auf Gr ößnskalen von mehr als etwa 10 bis 100 Metern homogen, Inhomogenitäten auf kleineren Skalen k nnten f r seine Aktivä t verantwortlich sein. Diese ist auf kleine Gebiete konzentriert, und auch Oberflächenveränderungen, die sich innerhalb von einigen Tagen bis wenigen Wochen abspielen, sind lokal. Im Kometenmaterial wurde eine Vielzahl an organischen Substanzen gemessen, die zum Teil als Schlüsselmoleküle für die Synthese der Grundbausteine des Lebens gelten, wie wir es kennen.

  3. Key points of condenser refurbishment illustrated by our experience on Russian technology nuclear power stations

    SciTech Connect

    Somville, C.

    1998-07-01

    In 1990, the refurbishment of the condensers of the VVER 440 MW LOVIISA 2 Finnish power station was the first reference of GEC ALSTHOM Delas on a Russian type nuclear power station, covering the optimization studies, technical and-economical choices, manufacture and site operations. The current contract for the condenser renovation of the 4 units of the VVER 440 MW PAKS Hungarian power station goes even further through an investment of this company in a local manufacturing installation and a significant participation of the local industry. Their expertise has helped reducing site operation times from 28 days for one condenser of one Loviisa unit, to 26 days for two condensers of one Paks unit. This paper describes the various aspects and the improvements brought for both operations and highlights the technical and economical key advantages of a condenser renovation (quick return on investment, better performances, reliability and life extension of the power station).

  4. The Role of Startups

    SciTech Connect

    Babinec, Sue

    2015-02-11

    Many ARPA-E-funded universities and research institutions have created start-up companies to further catalyze their next-generation technologies. Ambri and BlackPak are two examples of ARPA-E projects that were spun out by other institutions—Massachusetts Institute of Technology and SRI International, respectively—in an effort to get their technologies out of the lab and into the market quickly. This video features remarks from ARPA-E Senior Commercialization Advisor Sue Babinec and interviews with technologists at Ambri and BlackPak, who each tell the story of how their new companies spun out of the lab and have become agile startups capable of delivering real products to the marketplace.

  5. Optimization of the treatment of wheat samples for the determination of phytic acid by HPLC with refractive index detection.

    PubMed

    Amaro, Rosa; Murillo, Miguel; González, Zurima; Escalona, Andrés; Hernández, Luís

    2009-01-01

    The treatment of wheat samples was optimized before the determination of phytic acid by high-performance liquid chromatography with refractive index detection. Drying by lyophilization and oven drying were studied; drying by lyophilization gave better results, confirming that this step is critical in preventing significant loss of analyte. In the extraction step, washing of the residue and collection of this water before retention of the phytates in the NH2 Sep-Pak cartridge were important. The retention of phytates in the NH2 Sep-Pak cartridge and elimination of the HCI did not produce significant loss (P = 0.05) in the phytic acid content of the sample. Recoveries of phytic acid averaged 91%, which is a substantial improvement with respect to values reported by others using this methodology.

  6. Standardized Antimicrobial Disc Susceptibility Testing of Anaerobic Bacteria. I. Susceptibility of Bacteroides fragilis to Tetracycline

    PubMed Central

    Sutter, Vera L.; Kwok, Yung-Yuan; Finegold, Sydney M.

    1972-01-01

    A modified Bauer-Kirby-Sherris-Turck method for disc susceptibility testing of anaerobic bacteria is presented. When tetracycline was used against 100 strains of Bacteroides fragilis as a model, reasonably reproducible results were obtained after overnight incubation in both the GasPak atmosphere and an atmosphere achieved by adding 10% CO2 to a mixture of 10% H2 and 90% N2. The minimal inhibitory concentration for the strains determined by the agar dilution technique correlated well with the results of disc tests performed in the GasPak atmosphere with 30-μg tetracycline discs. Among 63 strains isolated from 1970 to the present, only 24 (38.1%) were found to be susceptible to tetracycline. PMID:5017675

  7. The Role of Startups

    ScienceCinema

    Babinec, Sue

    2016-07-12

    Many ARPA-E-funded universities and research institutions have created start-up companies to further catalyze their next-generation technologies. Ambri and BlackPak are two examples of ARPA-E projects that were spun out by other institutions—Massachusetts Institute of Technology and SRI International, respectively—in an effort to get their technologies out of the lab and into the market quickly. This video features remarks from ARPA-E Senior Commercialization Advisor Sue Babinec and interviews with technologists at Ambri and BlackPak, who each tell the story of how their new companies spun out of the lab and have become agile startups capable of delivering real products to the marketplace.

  8. Myxoma and vaccinia viruses exploit different mechanisms to enter and infect human cancer cells

    SciTech Connect

    Villa, Nancy Y.; Bartee, Eric; Mohamed, Mohamed R.; Rahman, Masmudur M.; Barrett, John W.; McFadden, Grant

    2010-06-05

    Myxoma (MYXV) and vaccinia (VACV) viruses have recently emerged as potential oncolytic agents that can infect and kill different human cancer cells. Although both are structurally similar, it is unknown whether the pathway(s) used by these poxviruses to enter and cause oncolysis in cancer cells are mechanistically similar. Here, we compared the entry of MYXV and VACV-WR into various human cancer cells and observed significant differences: 1 - low-pH treatment accelerates fusion-mediated entry of VACV but not MYXV, 2 - the tyrosine kinase inhibitor genistein inhibits entry of VACV, but not MYXV, 3 - knockdown of PAK1 revealed that it is required for a late stage event downstream of MYXV entry into cancer cells, whereas PAK1 is required for VACV entry into the same target cells. These results suggest that VACV and MYXV exploit different mechanisms to enter into human cancer cells, thus providing some rationale for their divergent cancer cell tropisms.

  9. Myxoma and vaccinia viruses exploit different mechanisms to enter and infect human cancer cells.

    PubMed

    Villa, Nancy Y; Bartee, Eric; Mohamed, Mohamed R; Rahman, Masmudur M; Barrett, John W; McFadden, Grant

    2010-06-05

    Myxoma (MYXV) and vaccinia (VACV) viruses have recently emerged as potential oncolytic agents that can infect and kill different human cancer cells. Although both are structurally similar, it is unknown whether the pathway(s) used by these poxviruses to enter and cause oncolysis in cancer cells are mechanistically similar. Here, we compared the entry of MYXV and VACV-WR into various human cancer cells and observed significant differences: 1--low-pH treatment accelerates fusion-mediated entry of VACV but not MYXV, 2--the tyrosine kinase inhibitor genistein inhibits entry of VACV, but not MYXV, 3--knockdown of PAK1 revealed that it is required for a late stage event downstream of MYXV entry into cancer cells, whereas PAK1 is required for VACV entry into the same target cells. These results suggest that VACV and MYXV exploit different mechanisms to enter into human cancer cells, thus providing some rationale for their divergent cancer cell tropisms.

  10. Measurement of viscoelasticity of UV photoresist used for nanoimprint lithography under confinement in nanometer-sized gaps

    NASA Astrophysics Data System (ADS)

    Itoh, Shintaro; Takahashi, Kazuhiro; Fukuzawa, Kenji; Zhang, Hedong

    2017-06-01

    In UV nanoimprint lithography, the viscoelasticity of a photoresist is a dominant factor that affects defect control, throughput, and alignment accuracy, all of which are important for high-volume manufacturing. During the nanoimprint process, the photoresist is confined in nanogaps and nanocavities, between the mold and the substrate. In this study, we measure the viscoelasticity of a UV photoresist, PAK-01, confined in nanometer-sized gap widths by the sensitive shear force measurement method developed in our previous study. Experimental results show that PAK-01, a viscous liquid in the bulk state, turns into a viscoelastic liquid in the nanogaps. Qualitatively, a similar transition is observed with the typical acrylate monomer, isobornyl acrylate. These findings are significant factors that should be considered in the improvement of the accuracy and efficiency of nanoimprint lithography.

  11. Direct determination of four sulfates and seven glucuronides of 17-oxosteroids in urine by fluorescence "high-performance" liquid chromatography.

    PubMed

    Iwata, J; Suga, T

    1989-05-01

    We describe a direct method for determining four sulfates and seven glucuronides of 17-oxosteroids (17OS) in urine without hydrolysis, by use of "high-performance" liquid chromatography (HPLC) with fluorometric detection. After pretreatment of urine samples with a Sep-Pak C18 cartridge, four 17OS sulfates and seven 17OS glucuronides in the pretreated urine samples were reacted with tetrapentylammonium ions to form ion pairs. Ion-paired 17OS sulfates were extracted with benzene. By adding sodium sulfate to the remaining sample, we could then extract ion-paired 17OS glucuronides with dichloromethane. Each extract was labeled with dansyl-hydrazine in an acetic acid-acetonitrile solution. The labeled steroids were separated by HPLC on a reversed-phase Capcell-Pak C8 (silicon-polymer-coated silica gel modified with octyl groups). We monitored each effluent with a fluorometric detector (330 nmexcitation, 535 nmemission).

  12. In vivo formation of codeinone-glutathione adduct: isolation and identification of a new metabolite in the bile of codeine-treated guinea pig.

    PubMed

    Ishida, T; Yano, M; Toki, S

    1998-01-01

    Codeinone-glutathione adduct (CO-GSH) in the bile of guinea pigs given a subcutaneous injection of codeine was isolated and identified. Synthesized authentic CO-GSH was characterized by the mass and nuclear magnetic resonance spectra and used as the standard sample. The metabolite was isolated by preparative high-performance liquid chromatography on a C18 column. The fractions containing the conjugated metabolite were purified using Sep-Pak C18 cartridges. For further purification of the metabolite CO-GSH, a Radial Pak CN column was used. Structure assignment of the metabolite was then performed by fast-atom-bombardment mass spectrometry and 500 MHz Fourier-transform-NMR spectrometric analysis and identified as S-[4,5-epoxy-3-methoxy-17-methyl-6-oxomorphinan-(8S)-yl] glutathione.

  13. [RAPD-markers linked to the locus for resistance to the race 4 pathogen for black rot, Xanthomonas campestris pv. campestris (Pamm.) Dow., in Brassica rapa L].

    PubMed

    Ignatov, A N; Kuginuki, Y; Suprunova, T P; Pozmogova, G E; Seitova, A M; Dorokhov, D B; Hirai, M

    2000-03-01

    Association between the RAPD markers and the resistance to race 4 of the black rot causative agent was studied in Brassica rapa L. Experiments were carried out using doubled haploid lines, obtained via crosses between the race 4-susceptible fodder turnip and resistant pak-choi, and the F2 progeny of the crosses between the doubled haploid lines with contrasting resistance. The WE(22)980 RAPD marker inherited from the pak-choi and associated with the clubroot susceptibility was also linked to the locus responsible for the resistance to race 4 of Xanthomonas campestris pv. campestris. The two other RAPD markers were linked to susceptibility to black rot. Simultaneous association of the same DNA markers with the resistance/susceptibility to two different obligate pathogens favored the hypothesis on cluster organization of the resistance genes in plants. The markers described can be used in plant breeding and in further investigation of the genetic bases of resistance in plants.

  14. Validation of a Rapid and Reliable Test for Diagnosis of Chagas' Disease by Detection of Trypanosoma cruzi-Specific Antibodies in Blood of Donors and Patients in Central America

    PubMed Central

    Ponce, Carlos; Ponce, Elisa; Vinelli, Elizabeth; Montoya, Alberto; de Aguilar, Vilma; Gonzalez, Antonio; Zingales, Bianca; Rangel-Aldao, Rafael; Levin, Mariano J.; Esfandiari, Javan; Umezawa, Eufrosina S.; Luquetti, Alejandro O.; da Silveira, José Franco

    2005-01-01

    In this study we compared the performance of the Chagas Stat-Pak rapid immunochromatographic test with a standard enzyme-linked immunosorbent assay (ELISA) in the serodiagnosis of Chagas' disease in Central America. Out of 3,400 blood donor samples, 156 (4.6%) were positive in both assays. Three sera out of 2,084 samples from reference laboratories were negative with the rapid test but positive with the ELISA (99.8% agreement). Agreement of 100% between the two tests was observed with 339 additional sera from patients with cardiopathies and 175 sera from potential blood donors in emergency surgical cases occurring on weekends or at night. In conclusion, Chagas Stat-Pak showed 99.6% and 99.9% sensitivity and specificity, respectively, when assayed with 5,998 serum samples. It is a sensitive and specific alternative to the ELISA, as required in medical emergencies and blood screenings in Central America. PMID:16207963

  15. Evaluating the validity of the serologic test for detecting Helicobacter pylori infection in Mongolian gerbils.

    PubMed

    Kuo, Chao-Hung; Yu, Fang-Jung; Tsai, Pei-Yun; Yang, Sheau-Fang; Chang, Lin-Li; Jan, Chang-Ming; Wang, Wen-Ming; Wu, Deng-Chyang

    2007-11-01

    A strong correlation between Helicobacter pylori infection and gastric cancer has been reported. Mongolian gerbils are regarded as the most suitable animal model in which to study carcinogenesis associated with H. pylori. The aim of our study was to evaluate the accuracy of the serologic test for detecting H. pylori infection in Mongolian gerbils. The model was developed as follows: the H. pylori colony (vacuolating cytotoxin A (+)/cytotoxin-associated gene A (+)) was cultured from the mucosas of previously H. pylori-fed gerbils. These colonies were cultured in broth. Then,we fed the gerbils with 0.5-1 mL of broth (about 10(9) CFU/mL) (intragastric administration) twice within a 3-day period. After inoculation for 6 or 26 weeks, the gerbils were sacrificed and their gastric mucosas were sampled for a series of examinations. Blood samples for serologic testing (STAT-PAK) were collected. H. pylori infection was confirmed. Statistical analysis was performed using the Chi-square test. Differences were regarded as significant when the p value was less than 0.05. A total of 50 gerbils were inoculated with H. pylori and the success rate reached 88%. All 10 gerbils in the control group showed a negative result. Damage to the mucosas was more obvious following increasing periods of inoculation. The rates of sensitivity and specificity, as determined by the STAT-PAK test, were 90.9% and 100%, respectively. The positive and negative predictive values were 100% and 60%, respectively. The STAT-PAK test seemed to be more sensitive and accurate (p < 0.05) in high H. pylori densities. In conclusion, the STAT-PAK test (blood-sampling) showed acceptable results and was suitable for long-term observation of H. pylori infection.

  16. Status Report on Medical Materiel Items Tested and Evaluated for Use in the USAF Aeromedical Evacuation System.

    DTIC Science & Technology

    1986-06-01

    one end of which is connected to or terminates in the immediate vicinity of the heart (AFR 160-3). a. Type A Equipment is equipment specifically...devices when the battery charger is connected to the Amb Pak. In this mode, the unit cannot be used on electrically susceptible pacients as defined in...whose terminal end is introduced into the thorax and is conductively connected to a point accessible outside the body (such as a probe, catheter, or

  17. Pressure Vessel Calculations for VVER-440 Reactors

    NASA Astrophysics Data System (ADS)

    Hordósy, G.; Hegyi, Gy.; Keresztúri, A.; Maráczy, Cs.; Temesvári, E.; Vértes, P.; Zsolnay, É.

    2003-06-01

    Monte Carlo calculations were performed for a selected cycle of the Paks NPP Unit II to test a computational model. In the model the source term was calculated by the core design code KARATE and the neutron transport calculations were performed by the MCNP. Different forms of the source specification were examined. The calculated results were compared with measurements and in most cases fairly good agreement was found.

  18. NPP training simulators in Hungary experience in development and utilization

    SciTech Connect

    Janosy, J.S.

    1996-11-01

    The construction of the only NPP in Hungary - the Paks NPP - started in 1975. The four units of VVER-440/213 were connected to the grid in 1982, 1984, 1986 and 1987. During the construction no simulator has been delivered with the power plant. Moreover, there were no state-of-art simulators in Central and Eastern Europe and in the former Soviet Union; not for the given type, not for civil use. The only simulator for the VVER-440 existing that time was made for the Loviisa NPP in Finland. This plant is not very similar to the Paks NPP; moreover, the pressure suppression system in the hermetical part of the primary circuit, the instrumentation and control systems, the main control room and the secondary circuit are completely different. Anyway, the training of Paks operators on this simulator was out of question - regardless the similarity problems. The design of the Paks NPP was made in the Soviet Union, therefore not too much design information was available in Hungary. During the creation of simulation models the authors had to rely mostly on common theory and measured performance. Besides the efforts to create a basic principle, full-scope replica and compact simulators there was a great need to use verified codes with more detailed models for better understanding the behavior and for evaluation of the safety. Thanks to these great efforts, the simulators were expanded to evaluate the performance of the trainees, for simulation of SBLOCA and LBLOCA events; the authors are checking and validating the operational procedures; soon they start the design of the functions of a new reactor protection system and they participate in international efforts to deliver training simulators to other VVER-440 power plants. The paper gives an overview of all these activities, referring to some key publications for each of them.

  19. Molecular Innovations Toward Theranostics of Aggressive Prostate Cancer

    DTIC Science & Technology

    2014-09-01

    Milli-Q water (18 M·cm) was obtained from a Millipore Gradient Milli-Q water system (Billerica, MA). All aqueous solutions were prepared with Milli...Q water . Silica gel 60 (70-230 mesh, Merck) was used for column chromatography. Analytical thin-layer chromatography (TLC) was performed using F254...were purified by Light C-18 Sep-Pak cartridges ( Waters , Milford, MA). Bulk solvents were removed by rotary evaporator under reduced pressure, and

  20. Indicators of NGO Security in Afghanistan

    DTIC Science & Technology

    2004-12-14

    warlord in charge of the province, the aid group is less likely to come under attack. Despite their many faults , the Taliban government curbed the...province share a boundary with Afghanistan border?(Yes/No) Source: Afghanistan Information Management System shapefile 7. BorderPak- Does the...province share a boundary with Pakistan? (Yes/No) Source: Afghanistan Information Management System shapefile 8. Foodpop#- Beneficiary population of

  1. U.S. Air Force Combat Psychiatry.

    DTIC Science & Technology

    1986-01-01

    Philippines in 1942. Completing a combat tour in prior wars was not without psychological and psychosomatic cost. The prolonged tension led to progressive...in Pak Five. The theory was, a new pilot should have time to adjust to the concept of being blown away byh thoywsa e iltsol aetm real enemy bullet...be a *I roar of motors in the air, but that is the normal state over an airfield. The next minute enemy fighter planes are buzzing the field, bullets

  2. Positive skin and serologic test results of diagnostic assays for bovine tuberculosis and subsequent isolation of Mycobacterium interjectum in a pygmy hippopotamus (Hexaprotodon liberiensis).

    PubMed

    Bouts, Tim; Vordermeier, Martin; Flach, Edmund; Routh, Andrew

    2009-09-01

    A 20-yr-old male pygmy hippopotamus (Hexaprotodon liberiensis), weighing 250 kg, arrived at Zoological Society London Whipsnade Zoo (United Kingdom) from a captive collection in Portugal. A quarantine health check was performed including a comparative intradermal tuberculosis (IDTB) test. Assessment of the comparative IDTB test at 72 hr revealed a strong positive reaction at the bovine site. Serum was tested with a rapid immunochromatographic assay (TB STAT-PAK) and was positive for tuberculosis antibodies. The tuberculosis tests were repeated 6 wk later with the same positive test outcome. In addition, a broncho-alveolar lavage (BAL) was submitted for mycobacterial culture. The positive IDTB test and TB STAT-PAK results were supported by multiantigen print immunoassay (MAPIA). Based on these results, the animal was suspected to be infected with Mycobacterium tuberculosis complex organisms and was euthanized. No gross or histologic signs of tuberculosis were found at postmortem examination. Mycobacterium interjectum was cultured from the BAL but not from necropsy samples. The antigens used in the TB STAT-PAK and MAPIA tests are reportedly specific for the M. tuberculosis complex, and so it is possible this animal presented with a latent case of tuberculosis or had a previous tuberculosis infection that resolved prior to testing. Cross-reactions with nontuberculous mycobacteria have been described with TB STAT-PAK and MAPIA tests. However, Western blotting analysis using serum from this animal did not recognize M. interjectum proteins of equivalent size to the M. tuberculosis-Mycobacterium bovis proteins recognized in the MAPIA. Thus, antigenic cross-reactivity with M. interjectum can be deemed less likely, but other nontuberculous mycobacterial proteins cannot be ruled out. It is therefore possible that false-positive reactions were obtained. These results highlight the difficulty of diagnosing tuberculosis in the absence of pathology and the presence of

  3. USSR Report, Life Sciences Biomedical and Behavioral Sciences

    DTIC Science & Technology

    1984-02-15

    Continuous Measuring of Glucose Concentration in Cerebral Tissue Using Enzyme Microelements (G. S. Kilibayeva, I. T. Demchenko; FIZIOLOGICHESKIY...Yu. G. Pak, et al.; ZHURNAL MIKROBIOLOGII, EPIDEMIOLOGII I IMMUNOBIOLOGII, No 10, Oct 83) 41 Activities of ’Pathogenicity’ Enzymes and Pesticin...AKADEMII NAUK UKRAINSKOY SSR, No 9, Sep 83) 43 g ~ PHARMACOLOGY AND TOXICOLOGY Effects of T-2 Toxin on Organelle-Specific Enzymes in Rat Tissues

  4. USNA: A Dual-Classifier Approach to Contextual Sentiment Analysis

    DTIC Science & Technology

    2013-06-01

    algorithms use emoticons as seman- tic indicators of polarity. For instance, a tweet that contains a sad face likely contains a negative polar- ity (Read...2005; Go et al., 2009; Bifet and Frank, 2010; Pak and Paroubek, 2010; Davidov et al., 2010; Kouloumpis et al., 2011). In a similar vein, hash- tags ...topic (e.g., “US President”), and retrieve tweets that contain the word (O’Connor et al., 2010; Tumasjan et al., 2010; Tan et al., 2011). These sys

  5. Global Phenomena from Local Rules: Peer-to-Peer Networks and Crystal Steps

    DTIC Science & Technology

    2007-01-01

    degree of Doctor of Philosophy 2007 Advisory Committee: Professor James Yorke, Chair/Advisor Professor Brian Hunt, Co-Advisor Professor Dionisios Margetis...Massachusetts, USA, June 2007. [62] Pak-Wing Fok, Rodolfo R. Rosales, and Dionisios Margetis. Unification of step bunching phenomena on vicinal...vicinal substrates. Physical Review Letters, 89(6):1268–1271, February 1998. [73] Dionisios Margetis, Michael J. Aziz, and Howard A. Stone. Continuum

  6. Environmental Fate Studies of HMX

    DTIC Science & Technology

    1983-09-01

    through tetra -nitroso derivatives of HPIX which eventually were- Tctabolized to 1, I-dimetbvilivdrazine. Computer simulations of the Holston River and...cell-I hr- 1 under anaerobic conditions. The metabolites resulting from both aerobic and anaerobic transformation were the mono- through tetra -nitroso...samples. In these analyses, a Water’s radial compression module equipped with a Water’s Radial- Pak C cartridge was used in con- junction with a water

  7. Progress on the Synthesis of Meso-Substituted Metallotetrabenzporphyrins

    DTIC Science & Technology

    1986-11-01

    Tetraphenyltetrabenzporphyrin 5 ZicTta(-ahhl)ttaezopyi Zinc Tetra (-npthayl) tetrabenzporphyrin 6 0 Zinc Tetra (p-N,N-dimethylaminophenyl...tetrabenzporphyrin 7 Zinc Tetra (p-rnthoxyph-enyl) tetrabenzporphyrin 7 ZicTta(-y.ohnl eraezopyi Zinc Tetra (pr-cyanophenyl) tetrabenzporphyrin 8 DISCUSS ION 9...was performed on a Waters 440 Liquid Chromatograph with a Hewlett-Packard HP1040A Spectrophotometric Detector using an 8-ram i.d. silica gel Radial Pak

  8. Neurotoxin Mitigation

    DTIC Science & Technology

    2007-11-01

    biotinamidopentyldecanamide HPLC high performance liquid chromatography ip intraperitoneal LC/MS/MS liquid chromatography coupled to tandem mass...outdated human plasma by ion exchange chromatography at pH 4.0, affinity chromatography on procainamide- Sepharose, and HPLC on a Protein Pak DEAE Anion...porcine trypsin (V5113C sequencing grade modified trypsin in 50 mM acetic acid, Promega) at 37˚C for 5.5 h. HPLC to purify the active site peptide

  9. DNA studies are necessary for accurate patient diagnosis in compound heterozygosity for Hb Adana (HBA2:c.179>A) with deletional or nondeletional α-thalassaemia

    PubMed Central

    Tan, Jin Ai Mary Anne; Kho, Siew Leng; Ngim, Chin Fang; Chua, Kek Heng; Goh, Ai Sim; Yeoh, Seoh Leng; George, Elizabeth

    2016-01-01

    Haemoglobin (Hb) Adana (HBA2:c.179>A) interacts with deletional and nondeletional α-thalassaemia mutations to produce HbH disorders with varying clinical manifestations from asymptomatic to severe anaemia with significant hepatosplenomegaly. Hb Adana carriers are generally asymptomatic and haemoglobin subtyping is unable to detect this highly unstable α-haemoglobin variant. This study identified 13 patients with compound heterozygosity for Hb Adana with either the 3.7 kb gene deletion (-α3.7), Hb Constant Spring (HbCS) (HBA2:c.427T>C) or Hb Paksé (HBA2:429A>T). Multiplex Amplification Refractory Mutation System was used for the detection of five deletional and six nondeletional α-thalassaemia mutations. Duplex-PCR was used to confirm Hb Paksé and HbCS. Results showed 84.6% of the Hb Adana patients were Malays. Using DNA studies, compound heterozygosity for Hb Adana and HbCS (αcodon 59α/αCSα) was confirmed in 11 patients. A novel point in this investigation was that DNA studies confirmed Hb Paksé for the first time in a Malaysian patient (αcodon 59α/αPakséα) after nine years of being misdiagnosis with Hb Adana and HbCS (αcodon 59α/αCSα). Thus, the reliance on haematology studies and Hb subtyping to detect Hb variants is inadequate in countries where thalassaemia is prevalent and caused by a wide spectrum of mutations. PMID:27271331

  10. JPRS Report, Soviet Union, International Affairs.

    DTIC Science & Technology

    1987-11-27

    Revolutionaries" by B. Pak; the conclud- ing part of I. Belyaev’s article "Egypt. The Revolution and the President" "Mahatmah Gandhi as Described by...O. Martyshin. Mahatmah Gandhi in the Evaluations of Soviet Researchers (p 22) Problems and Opinions VI. Li, G. Mirskiy. Socialist Orientation in...Scientific Life V. Rozhkov. A Forum for Mongolian Scholars (p 44) Culture, Literature, Art V. Ozhogin. Lev Tolstoy and Japanese Culture (p 46

  11. Energy to the Edge (E2E) U.S. Army Rapid Equipping Force

    DTIC Science & Technology

    2014-03-21

    efficient systems, warfighter doctrine, training and combat operations. The greatest impact of Operational Energy is that it is not just about saving...Edge Hybrid Energy System Patrol Pak Description: • necessary to keep the appliance loads operating. • Open Architecture allows multiple sources of...power generation • 2kW-h energy storage (batteries) • Field serviceable • Area: 10-200 Sq ft • High- Efficiency power model - Power transfers in

  12. Korean Affairs Report No. 321

    DTIC Science & Technology

    2007-11-02

    the incident might be caused by an internal foe as in the case of the assassination of Pak Jung -hi in 1979. Another paper of the country MERDEKA 11...Comrade Henryk Jablonski, president of the Council of State of the Polish People’s Republic; Comrade Gustav Husak, general secretary of the Central...president, and Dominic Mintoff, prime minister, of the Republic of Malta; Carl Gustaf, king of Sweden; Rudolf Kirchschlager, president of the

  13. Behavior of Materials at Cold Regions Temperatures. Part 1. Program Rationale and Test Plan

    DTIC Science & Technology

    1988-07-01

    10 15 i" 0% I 0 0 0 2 3 4 5a 0 1 2 3 4 5 6 7 lf CSTRAIN (IN/INbST AI OW N V, Figure B47. Static stress vs strain curves for resilient expanded ... polystyrene foam (Resilo-Pak) , at temperature extremes (0.5, 0. 75 and 1.3 lb Ifft3 densities) (Titus 1967). so ,eSF 50 -- 8 * TEMPERATURE. K 0 so 100 is

  14. Estrogen and the Dietary Phytoestrogen Tesveratrol as Regulators of the Rho GTPase Rac in Breast Cancer Research

    DTIC Science & Technology

    2008-06-01

    AD_________________ Award Number: W81XWH-07-1-0330 TITLE: Estrogen and the Dietary Phytoestrogen ...COVERED 7 May 2007 – 6 May 2008 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Estrogen and the Dietary Phytoestrogen Tesveratrol as Regulators of the... estrogen (E2), or resveratrol (Res). PAK-PBD-GST beads were used to pull-down active GTP- bound Rac from the cell lysates. Active and total Rac levels

  15. New Developments on the Core Function for Efficient Implementation of the Difficult Residue Number System Operations.

    DTIC Science & Technology

    1985-02-01

    throughput, and hardware complexity. A considerable body of Soviet work exists in the open literature, dating back to the mid 1960 ’s. This includes...Amerbaev, V. Burcev, I. Pak, and D. Yuditskii. Their results have both paralleled and diverged from Western work. In the late 1960 ’s, hardware...processing. A recurrent theme in much of the work from the mid 1960 ’s to the present has been the study of positional characteristics of residue encoded

  16. High yield one-pot production of [(18)F]FCH via a modified TRACERlab FxFN module.

    PubMed

    Huang, Ya-Yao; Tsai, Chia-Ling; Wen, Hsiang-Ping; Tzen, Kai-Yuan; Yen, Ruoh-Fen; Shiue, Chyng-Yann

    2017-10-01

    [(18)F]Fluoromethylcholine ([(18)F]FCH) is a potent tumors imaging agent. In order to fulfill the demand of pre-clinical and clinical studies, we have developed an automated high yield one-pot synthesis of this potent tumors imaging agent. [(18)F]FCH was synthesized using a modified TRACERlab FxFN module. Briefly, dibromomethane (10% in CH3CN) was fluorinated with K[(18)F]/K 2.2.2 in a glassy carbon reaction vessel at 120°C for about 5min to generate [(18)F]fluorobromomethane ([(18)F]FBM). The resulting [(18)F]FBM was then bubbling (He, 700mL/min) through four Sep-Pak® Silica Plus Long cartridges to react with dimethylaminoethanol (10% DMAE in 0.3mL DMSO) which was pre-loaded on Sep-Pak® C18 Plus Short cartridge. The [(18)F]FCH was purified by solid-phase extraction (SPE) using one Sep-Pak® C18 Plus Short and one Sep-Pak® CM Plus Short in series. The quality of [(18)F]FCH synthesized by this method was verified by HPLC and TLC as compared to authentic sample. Using this improved one-pot method, the RCY of [(18)F]FCH was 18.8 ± 2.1% (EOB, n = 27) in a synthesis time of 49 ± 5min from EOB. The radiochemical purity of [(18)F]FCH was greater than 90% and the residual DMAE concentration in the final product was less than 10ppm. This optimized method could fulfill the demand of [(18)F]FCH for both pre-clinical and clinical studies, especially for nearby study sites without a cyclotron. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. In vivo yeast cell morphogenesis is regulated by a p21-activated kinase in the human pathogen Penicillium marneffei.

    PubMed

    Boyce, Kylie J; Schreider, Lena; Andrianopoulos, Alex

    2009-11-01

    Pathogens have developed diverse strategies to infect their hosts and evade the host defense systems. Many pathogens reside within host phagocytic cells, thus evading much of the host immune system. For dimorphic fungal pathogens which grow in a multicellular hyphal form, a central attribute which facilitates growth inside host cells without rapid killing is the capacity to switch from the hyphal growth form to a unicellular yeast form. Blocking this transition abolishes or severely reduces pathogenicity. Host body temperature (37 degrees C) is the most common inducer of the hyphal to yeast transition in vitro for many dimorphic fungi, and it is often assumed that this is the inducer in vivo. This work describes the identification and analysis of a new pathway involved in sensing the environment inside a host cell by a dimorphic fungal pathogen, Penicillium marneffei. The pakB gene, encoding a p21-activated kinase, defines this pathway and operates independently of known effectors in P. marneffei. Expression of pakB is upregulated in P. marneffei yeast cells isolated from macrophages but absent from in vitro cultured yeast cells produced at 37 degrees C. Deletion of pakB leads to a failure to produce yeast cells inside macrophages but no effect in vitro at 37 degrees C. Loss of pakB also leads to the inappropriate production of yeast cells at 25 degrees C in vitro, and the mechanism underlying this requires the activity of the central regulator of asexual development. The data shows that this new pathway is central to eliciting the appropriate morphogenetic response by the pathogen to the host environment independently of the common temperature signal, thus clearly separating the temperature- and intracellular-dependent signaling systems.

  18. Dynamics of Flagellum- and Pilus-Mediated Association of Pseudomonas aeruginosa with Contact Lens Surfaces▿

    PubMed Central

    Tran, Victoria B.; Fleiszig, Suzanne M. J.; Evans, David J.; Radke, Clayton J.

    2011-01-01

    Flagella and pili are appendages that modulate attachment of Pseudomonas aeruginosa to solid surfaces. However, previous studies have mostly reported absolute attachment. Neither the dynamic roles of these appendages in surface association nor those of attachment phenotypes have been quantified. We used video microscopy to address this issue. Unworn, sterile, soft contact lenses were placed in a laminar-flow optical chamber. Initial lens association kinetics for P. aeruginosa strain PAK were assessed in addition to lens-surface association phenotypes. Comparisons were made to strains with mutations in flagellin (fliC) or pilin (pilA) or those in flagellum (motAB) or pilus (pilU) function. PAK and its mutants associated with the contact lens surface at a constant rate according to first-order kinetics. Nonswimming mutants associated ∼30 to 40 times slower than the wild type. PAK and its pilA mutant associated at similar rates, but each ∼4 times faster than the pilU mutant. Lens attachment by wild-type PAK induced multiple phenotypes (static, lateral, and rotational surface movement), each showing only minor detachment. Flagellin (fliC) and flagellar-motility (motAB) mutants did not exhibit surface rotation. Conversely, strains with mutations in pilin (pilA) and pilus retraction (pilU) lacked lateral-surface movement but displayed enhanced surface rotation. Slower surface association of swimming-incapable P. aeruginosa mutants was ascribed to lower convective-diffusion-arrival rates, not to an inability to adhere. Flagellum function (swimming) enhanced lens association, attachment, and rotation; hyperpiliation hindered lens association. P. aeruginosa bound through three different adhesion sites: flagellum, pili, and body. Reduction of bacterial attachment to contact lenses thus requires blockage of multiple adhesion phenotypes. PMID:21498762

  19. Altered expression of CDC42 signaling pathway components in cortical layer 3 pyramidal cells in schizophrenia.

    PubMed

    Datta, Dibyadeep; Arion, Dominique; Corradi, John P; Lewis, David A

    2015-12-01

    Cognitive dysfunction in schizophrenia is associated with a lower density of dendritic spines on deep layer 3 pyramidal cells in the dorsolateral prefrontal cortex (DLPFC). These alterations appear to reflect dysregulation of the actin cytoskeleton required for spine formation and maintenance. Consistent with this idea, altered expression of genes in the cell division cycle 42 (CDC42)-CDC42 effector protein (CDC42EP) signaling pathway, a key organizer of the actin cytoskeleton, was previously reported in DLPFC gray matter from subjects with schizophrenia. We examined the integrity of the CDC42-p21-activated serine/threonine protein kinases (PAK)-LIM domain-containing serine/threonine protein kinases (LIMK) signaling pathway in schizophrenia in a layer-specific and cell type-specific fashion in DLPFC deep layer 3. Using laser microdissection, samples of DLPFC deep layer 3 were collected from 56 matched pairs of subjects with schizophrenia and comparison subjects, and levels of CDC42-PAK-LIMK pathway messenger RNAs were measured by quantitative polymerase chain reaction. These same transcripts also were quantified by microarray in samples of individually microdissected deep layer 3 pyramidal cells from a subset of the same subjects and from monkeys exposed to antipsychotics. Relative to comparison subjects, CDC42EP4, LIMK1, LIMK2, ARHGDIA, and PAK3 messenger RNA levels were significantly upregulated in subjects with schizophrenia in laminar and cellular samples. In contrast, CDC42 and PAK1 messenger RNA levels were significantly downregulated specifically in deep layer 3 pyramidal cells. These differences were not attributable to psychotropic medications or other comorbid factors. Findings from the present and prior studies converge on synergistic alterations in CDC42 signaling pathway that could destabilize actin dynamics and produce spine deficits preferentially in deep layer 3 pyramidal cells in schizophrenia. Copyright © 2015 Society of Biological Psychiatry

  20. Delayed fluorescence spectra of intact leaves photoexcited by sunlight measured with a multichannel Fourier-transform chemiluminescence spectrometer

    NASA Astrophysics Data System (ADS)

    Akita, Saeka; Yano, Ayako; Ishii, Hiroshi; Satoh, Chikahiro; Akai, Nobuyuki; Nakata, Munetaka

    2013-06-01

    Delayed fluorescence spectra of intact leaves of Green pak choi (Brassica rapa var. chinensis) were measured with a multichannel Fourier-transform chemiluminescence spectrometer, which we developed recently. The intact samples, photoexcited by sunlight without artificial light sources, showed delayed fluorescence around 740 nm with a lifetime of ˜6 s. The observed spectra were deconvoluted into two Gaussian bands: the delayed fluorescence from photosystem II and photosystem I complexes. Their relative intensities depended on the chlorophyll concentration, but their wavelengths were unchanged.

  1. Dissociation of Crk-associated substrate from the vimentin network is regulated by p21-activated kinase upon acetylcholine activation of airway smooth muscle

    PubMed Central

    Wang, Ruping; Li, Qing-Fen; Anfinogenova, Yana; Tang, Dale D.

    2006-01-01

    The intermediate filament protein vimentin has been shown to be required for smooth muscle contraction. The adapter protein p130 Crk-associated substrate (CAS) participates in the signaling processes that regulate force development in smooth muscle. However, the interaction of vimentin filaments with CAS has not been well elucidated. In the present study, stimulation of tracheal smooth muscle strips with acetylcholine (ACh) resulted in the increase in ratios of soluble vimentin to insoluble vimentin (an index of vimentin disassembly) in association with force development. Activation with ACh also induced vimentin phosphorylation at Ser-56 as assessed by immunoblot analysis. More importantly, CAS was found in the cytoskeletal vimentin fraction, and the amount of CAS in cytoskeletal vimentin was reduced in smooth muscle strips upon contractile stimulation. CAS redistributed from the myoplasm to the periphery during ACh activation of smooth muscle cells. The decrease in distribution of CAS in cytoskeletal vimentin elicited by ACh was attenuated by the downregulation of p21-activated kinase (PAK) 1 with antisense oligodeoxynucleotides. Vimentin phosphorylation at this residue, the ratio of soluble vimentin to insoluble vimentin, and active force in smooth muscle strips induced by ACh were also reduced in PAK-depleted tissues. These results suggest that PAK may regulate CAS release from the vimentin intermediate filaments by mediating vimentin phosphorylation at Ser-56 and the transition of cytoskeletal vimentin to soluble vimentin. The PAK-mediated the dissociation of CAS from the vimentin network may participate in the cellular processes that affect active force development during acetylcholine activation of tracheal smooth muscle tissues. PMID:16997882

  2. Building a Data-Driven Vital Sign Indicator for an Economically Optimized Component Replacement Policy

    DTIC Science & Technology

    2014-10-02

    Jardine & Tsang, 2013). The optimization of replacement decision policy based on component failure predictions has been critical in the area of...PROGNOSTICS AND HEALTH MANAGEMENT SOCIETY 2014 2 (Banjevic, Jardine , Makis, & Ennis, 2001)( Jardine , Banjevic, Montgomery, & Pak, 2008). In practice, the...of the American Statistical Association, 101(473). Banjevic, D., Jardine , A., Makis, V., & Ennis, M. (2001). A control-limit policy and software for

  3. Conference Proceedings of NASA/DoD Controls-Structures Interaction Technology Held in San Diego, California on 29 January-2 February 1989,

    DTIC Science & Technology

    1989-08-01

    Suite H 2015 Neil Avenue San Diego, CA 92024 Columbus. OH1 43210 Young H. Pak Richard S. Pappa Bus. Phor e: 714-896-4682 Bus. Phone: 804-864-4321...of the FMI was inspired by the work of Mike Shao of the Harvard Smithsonian Astrophysical Observatory. Dr. Shao currently has in operation, on Mount...1995 to 2015 (Astronomy and Astrophysics), National Academy Press, Washington, DC, 1988. 2. NASA Office of Space Science and Applications 1988

  4. Pakistan: Frontline State Again?

    DTIC Science & Technology

    1995-12-01

    leader Mohammed Ali Jinnah embarked on a new strategy against Indian Congress domination stating that Islam was in danger and the congress was...A. OUTLINE OF THE STUDY ............................. 4 II. A COLD W AR W ITHIN ...................................... 9 A. LEGACY OF PARTITION ...in the region, one absolute of Indo-Pak relations remains after the end of the cold war. Since the partition of India in 1947, Pakistan has been

  5. Tagging b jets associated with heavy neutral MSSM Higgs bosons

    NASA Astrophysics Data System (ADS)

    Heikkinen, A.; Lehti, S.

    2006-04-01

    Since a neural network (NN) approach has been shown to be applicable to the problem of Higgs boson detection at LHC [I. Iashvili, A. Kharchilava, CMS TN-1996/100; M. Mjahed, Nucl. Phys. B 140 (2005) 799], we study the use of NNs in the problem of tagging b jets in pp →bb¯HSUSY, HSUSY→ττ in the Compact Muons Solenoid experiment [F. Hakl, et al., Nucl. Instr. and Meth. A 502 (2003) 489; S. Lehti, CMS NOTE-2001/019; G. Segneri, F. Palla, CMS NOTE-2002/046]. B tagging is an important tool for separating the Higgs events with associated b jets from the Drell-Yan background Z,γ*→ττ, for which the associated jets are mostly light quark and gluon jets. We teach multi-layer perceptrons (MLPs) available in the object oriented implementation of data analysis framework ROOT [ROOT—An Object Oriented Data Analysis Framework, in: Proceedings of the AIHENP'96 Workshop, Lausanne, September 1996, Nucl. Instr. and Meth. A 389 (1997) 81]. The following learning methods are evaluated: steepest descent algorithm, (BFGS) Broyden-Fletcher-Goldfarb-Shanno algorithm, and variants of conjugate gradients. The ROOT code generation feature of standalone C++ classifiers is utilized. We compare the b tagging performance of MLPs with another ROOT based feed forward NN tool NeuNet [J.P. Ernenwein, NeuNet software for ROOT], which uses a common back-propagation learning method. In addition, we demonstrate the use of the self-organizing map program package (SOM_PAK) and the learning vector quantization program package (LVQ_PAK) [T. Kohonen, et al., SOM_PAK: the self-organizing map program package, Technical Report A31; T. Kohonen, et al., LVQ_PAK: the learning vector quantization program package, Technical Report A30, Laboratory of Computer and Information Science, Helsinki University of Technology, FIN-02150 Espoo, Finland, 1996] in the b tagging problem.

  6. The Soviet Union in Afghanistan: Benefits and Costs.

    DTIC Science & Technology

    1980-06-12

    COLLEGE Carlisle Barracks, Pennsylvania THE SOVIET NION IN4FGHANISTAN: S BENFITS AND COSTS, by SS-hirin/Fahir-Kheli ,/;/ 12 June480./ -/ . DISTRIBUTION...1973. The Soviet Union committed itself to 20 major projects in agriculture, irrigation, electric power, oil and gas exploration , mineral and metal...regime spread, Pak-Afghan relations deteriorated. Pakistan counted 56 violations of its air and ground space (penetratioins of up to three miles above

  7. Altered expression of CDC42 signaling pathway components in cortical layer 3 pyramidal cells in schizophrenia

    PubMed Central

    Datta, Dibyadeep; Arion, Dominique; Corradi, John P.; Lewis, David A.

    2015-01-01

    Background Cognitive dysfunction in schizophrenia is associated with a lower density of dendritic spines on deep layer 3 pyramidal cells in the dorsolateral prefrontal cortex (DLPFC). These alterations appear to reflect dysregulation of the actin cytoskeleton required for spine formation and maintenance. Consistent with this idea, altered expression of genes in the CDC42 (cell division cycle 42)-CDC42 effector protein signaling pathway, a key organizer of the actin cytoskeleton, was previously reported in DLPFC gray matter from subjects with schizophrenia. Here, we examined the integrity in schizophrenia of the CDC42-PAK-LIMK signaling pathway in a layer- and cell type-specific fashion in DLPFC deep layer 3. Methods Using laser microdissection, we collected samples of DLPFC deep layer 3 from 56 matched pairs of schizophrenia and comparison subjects and measured levels of CDC42-PAK-LIMK pathway mRNAs by qPCR. These same transcripts were also quantified by microarray in samples of individually microdissected deep layer 3 pyramidal cells from a subset of the same subjects and from antipsychotic-exposed monkeys. Results Relative to comparison subjects, CDC42EP4, LIMK1, LIMK2, ARHGDIA and PAK3 mRNA levels were significantly up-regulated in schizophrenia subjects in both laminar and cellular samples. In contrast, CDC42 and PAK1 mRNA levels were significantly down-regulated specifically in deep layer 3 pyramidal cells. These differences were not attributable to psychotropic medications or other co-morbid factors. Conclusions Findings from the present and prior studies converge on synergistic alterations in CDC42 signaling pathway that could destabilize actin dynamics and produce spine deficits preferentially in deep layer 3 pyramidal cells in schizophrenia. PMID:25981171

  8. On Target: Organizing and Executing the Strategic Air Campaign Against Iraq

    DTIC Science & Technology

    2002-01-01

    and the Arab powers in the Persian Gulf as dramatic as that of Egyptian President Anwar Sadat over the Israelis in October 1973.2 This action followed...between intelligence, operations, and planners. 23. Intvw, Col. Crigger, Mar 5, 1992. Wild Weasels operating from Shaikh Isa in Bahrain, also close to...Mohammed Sabah al-Salam: 27, 278 Sadat, Anwar : 1 Saddam International Airport (IAP): 201 Safwan airfield: 290 Salah Al Din SAM plant: 269 Salman Pak AM

  9. Criminal Acts Against Civil Aviation

    DTIC Science & Technology

    1991-01-01

    Services (ACS), a private company, was located on the bottom floor. Seven persons, including six who were in the ACS office, were killed in the...explosion. Reportedly, the device accidentally exploded in the hands of i’ Pak iinda, gU-irilla. whose intended target was the British Consulate, located ...Airlines was located . The explosion caused damage and one injury. The Greek terrorist group, People’s Struggle (LEA), claimed responsibility for the

  10. Performance of converted pressure cookers and two conventional jars for anaerobic bacterial culture.

    PubMed

    Gargan, R A; Phillips, I

    1978-05-01

    The simple conversion of commercial pressure cookers into inexpensive anaerobic jars is described. These containers were shown to be as good as the small conventional BBL polycarbonate GasPak and large vented 150 gas-replacement jars when assessed by means of three biological indicators: Pseudomonas aeruginosa, Bacteroides melaninogenicus, and Bacteroides fragilis. Ps. aeruginosa seeded on Simmond's citrate agar was shown to be the most sensitive indicator of the three for traces of oxygen.

  11. Simultaneous extraction of. beta. -endorphin and leu- and met-enkephalins from human and rat plasma

    SciTech Connect

    Bhathena, S.J.; Smith, P.M.; Kennedy, B.W. ); Voyles, N.R.; Recant, L. )

    1989-01-01

    A simple, rapid and reliable procedure is described to simultaneously concentrated and purify {beta}-endorphin, leu-and met-enkephalins from small volumes of human and rat plasma before radioimmunoassay is performed. It uses C{sub 18} Sep-Pak reverse phase cartridges. The effectiveness of different protease inhibitors in preventing degradation of opiates by plasma and different solvent systems for eluting opiates is also evaluated.

  12. Complete Genome Sequence of a Velogenic Newcastle Disease Virus Strain Isolated from a Clinically Healthy Exotic Parakeet (Melopsittacus undulatus) in Pakistan

    PubMed Central

    Wajid, Abdul; Basharat, Asma; Khan, Taseer Ahmed; Wasim, Muhammad

    2017-01-01

    ABSTRACT The complete genome sequence of a virulent Newcastle disease virus (vNDV) strain isolated from an exotic parakeet (Melopsittacus undulatus) is described here. The virulent strain parakeet/Pak/R-Pindi/SFR-16/2016 was isolated from a bird reared as a pet in the province of Punjab in the northern region of Pakistan in 2016. Phylogenetic analysis classified the isolate as a member of NDV class II, subgenotype VIIi, in genotype VII. PMID:28183762

  13. JPRS Report, Science & Technology, Europe & Latin America

    DTIC Science & Technology

    1987-07-23

    We developed the protocol according to the LSV 2 prescriptions of Siemens. Data transmission speed can be 1200 -9600 baud. In the course of spatial...for two reactor types-- VVER -440 and VVER -IO00. (In the case of the Paks nuclear power plant the first four reactor blocks belong to the former type...reactor and of its behavior under breakdown or accident conditions (thermohydraulics), In the case of the VVER -440 this separation has not yet led to

  14. 78 FR 17992 - Additional Designation of Three North Korean Individuals Pursuant to Executive Order 13382

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-25

    ... of State. ACTION: Designation of Pak To-Chun, Chu Kyu-Chang, and O Kuk-Ryol Pursuant to E.O. 13382...-Chang, and O Kuk-Ryol have engaged, or attempted to engage, in activities or transactions that have..., Chagang Province, DPRK CHU KYU-CHANG A.K.A.: Chu Kyu-Ch'ang A.K.A.: Ju Kyu-Chang D.O.B.: November 25, 1928...

  15. Amino acid sequence of atrial natriuretic peptides in human coronary sinus plasma.

    PubMed

    Yandle, T; Crozier, I; Nicholls, G; Espiner, E; Carne, A; Brennan, S

    1987-07-31

    Two atrial natriuretic peptides were purified from pooled human coronary sinus plasma by Sep-Pak extraction, immunoaffinity chromatography and reverse phase HPLC. The amino acid sequences of the two peptides were homologous with 99-126 human atrial natriuretic peptide (hANP) and 106-126 hANP, the latter being most probably linked to 99-105 ANP by the disulphide bond. The molar ratio of the peptides in plasma, as assessed by radioimmunoassay was 10:3.

  16. Saugus River and Tributaries Flood Damage Reduction Study: Lynn, Malden, Revere and Saugus, Massachusetts. Volume 7. Appendix J. Feasibility Study and EIS/EIR Comments and Responses. Section C. Final Report Review,

    DTIC Science & Technology

    1990-04-01

    a:least the year. NO PORN . REL. [TAR). Block 3. Tys of Reort agg Oates Covred. 0 e .053.4 itbur State whethier resort is Interim, final. etc If 00-Se1enn...also MWd sex OP, and provide the real estate and relocation requirements estimated at $9,200,000 for Stony Broo* Reser’abons Al, MebtoPaks MetroParkways

  17. Testing the Role of p21-Activated Kinases in Schwannoma Formation Using a Novel Genetically Engineered Murine Model that Closely Phenocopies Human NF2 Disease

    DTIC Science & Technology

    2015-06-01

    2 (NF2) is an autosomal dominant genetic disease characterized by benign schwannomas that grow on the cranial and spinal nerves . While technically...DRG and spinal nerves compared to Postn-Cre+;Nf2flox/flox mice. Diffuse spinal nerve hyperplasia and pseudo onion bulb formation of proliferating...schwann cells observed in the nerve of a Postn-Cre+;Nf2flox/flox;Pak1-/- mouse; however, frequency is lower. Original magnification 40x.   5

  18. Radial Artery Pulse Change During Cold Pressor Test

    DTIC Science & Technology

    2007-11-02

    KRF-2000-015-PP0177) REFERENCES [1] C.H. Chen, C.T. Ting, A. Nussbacher, E. Nevo , D. A. Kass, P. Pak, S. P. Wang, M. S Chang, C.P. Frank, “Validation...E. Nevo , B. Fetics, H. Peter, F. C.P. Yin, W. L. Maughan,s D. A. Kass “Estimation of Central Aortic Pressure Waveform by Mathematical Transformation

  19. Targeting and activation of Rac1 are mediated by the exchange factor β-Pix

    PubMed Central

    ten Klooster, Jean Paul; Jaffer, Zahara M.; Chernoff, Jonathan; Hordijk, Peter L.

    2006-01-01

    Rho guanosine triphosphatases (GTPases) are critical regulators of cytoskeletal dynamics and control complex functions such as cell adhesion, spreading, migration, and cell division. It is generally accepted that localized GTPase activation is required for the proper initiation of downstream signaling events, although the molecular mechanisms that control targeting of Rho GTPases are unknown. In this study, we show that the Rho GTPase Rac1, via a proline stretch in its COOH terminus, binds directly to the SH3 domain of the Cdc42/Rac activator β-Pix (p21-activated kinase [Pak]–interacting exchange factor). The interaction with β-Pix is nucleotide independent and is necessary and sufficient for Rac1 recruitment to membrane ruffles and to focal adhesions. In addition, the Rac1–β-Pix interaction is required for Rac1 activation by β-Pix as well as for Rac1-mediated spreading. Finally, using cells deficient for the β-Pix–binding kinase Pak1, we show that Pak1 regulates the Rac1–β-Pix interaction and controls cell spreading and adhesion-induced Rac1 activation. These data provide a model for the intracellular targeting and localized activation of Rac1 through its exchange factor β-Pix. PMID:16492808

  20. Novel regulatory mechanisms for the Dbl family guanine nucleotide exchange factor Cool-2/alpha-Pix.

    PubMed

    Feng, Qiyu; Baird, Daniel; Cerione, Richard A

    2004-09-01

    The Cool-2 (cloned-out of library-2) protein (identical to alpha-Pix for Pak-interactive exchange factor) has been implicated in various biological responses including chemoattractant signaling and in certain forms of mental retardation. We show that when Cool-2 exists as a dimer, it functions as a Rac-specific guanine nucleotide exchange factor (GEF). Dimerization of Cool-2 enables its Dbl (diffuse B-cell lymphoma) and pleckstrin homology domains to work together (in trans) to bind specifically to Rac-GDP. Dissociation of dimeric Cool-2 into its monomeric form allows it to act as a GEF for Cdc42 as well as for Rac. The binding of either PAK (p21-activated kinase) or Cbl (Casitas B-lymphoma) to the SH3 domain of monomeric Cool-2 is necessary for the functional interactions between GDP-bound Cdc42 or Rac and the Cool-2 monomer. The betagamma subunit complex of large GTP-binding proteins, by interacting with PAK, stimulates the dissociation of the Cool-2 dimer and activates its GEF activity for Cdc42. Overall, these findings highlight novel mechanisms by which extracellular signals can direct the specific activation of Rac versus Cdc42 by Cool-2/alpha-Pix.

  1. Targeting and activation of Rac1 are mediated by the exchange factor beta-Pix.

    PubMed

    ten Klooster, Jean Paul; Jaffer, Zahara M; Chernoff, Jonathan; Hordijk, Peter L

    2006-02-27

    Rho guanosine triphosphatases (GTPases) are critical regulators of cytoskeletal dynamics and control complex functions such as cell adhesion, spreading, migration, and cell division. It is generally accepted that localized GTPase activation is required for the proper initiation of downstream signaling events, although the molecular mechanisms that control targeting of Rho GTPases are unknown. In this study, we show that the Rho GTPase Rac1, via a proline stretch in its COOH terminus, binds directly to the SH3 domain of the Cdc42/Rac activator beta-Pix (p21-activated kinase [Pak]-interacting exchange factor). The interaction with beta-Pix is nucleotide independent and is necessary and sufficient for Rac1 recruitment to membrane ruffles and to focal adhesions. In addition, the Rac1-beta-Pix interaction is required for Rac1 activation by beta-Pix as well as for Rac1-mediated spreading. Finally, using cells deficient for the beta-Pix-binding kinase Pak1, we show that Pak1 regulates the Rac1-beta-Pix interaction and controls cell spreading and adhesion-induced Rac1 activation. These data provide a model for the intracellular targeting and localized activation of Rac1 through its exchange factor beta-Pix.

  2. DEVELOPMENT OF A HYDRONIC ROOFTOP UNIT -- HYPAK

    SciTech Connect

    Eric Lee; Dick Bourne; Mark Berman

    2004-03-25

    The majority of US commercial floor space is cooled by rooftop HVAC units (RTU's). RTU popularity derives chiefly from their low initial cost and relative ease of service access without disturbing building occupants. Unfortunately, current RTU's are inherently inefficient due to a combination of characteristics that unnecessarily increase cooling loads and energy use. Existing RTU's in the U.S. consume an estimated 2.4 quads annually. Inefficient RTU's create an estimated 3.5% of U.S. CO{sub 2} emissions, thus contributing significantly to global warming. Also, RTU's often fail to maintain adequate ventilation air and air filtration. This project was developed to evaluate the feasibility of a radically new ''HyPak'' RTU design that significantly and cost-effectively increases RTU performance and delivered air quality. The objective of the HyPak Project was to design, develop and test a hydronic RTU that provides a quantum improvement over conventional RTU performance. Our proposal targeted 60% and 50% reduction in electrical energy use by the HyPak RTU for dry and humid climates, respectively, when compared with a conventional unit.

  3. Determination of tetracyclines in bovine and porcine muscle by high-performance liquid chromatography using solid-phase extraction.

    PubMed

    Walsh, J R; Walker, L V; Webber, J J

    1992-04-10

    A method is presented for the determination of the three tetracyclines oxytetracycline, tetracycline and chlortetracycline in muscle, spiked at 100 ng/g, using high-performance liquid chromatography (HPLC). The concentration and extraction steps are carried out using Waters Environmental Sep-Pak cartridges. The principal steps involve homogenizing the sample in EDTA-McIlvaine buffer followed by centrifugation and precipitation of the supernatant using trichloroacetic acid. After further filtration and concentration on a Sep-Pak cartridge, the sample is eluted and analysed by HPLC with UV detection and confirmation by diode-array. The column used is a Nova-Pak C18 (4 microns) cartridge (10 cm x 8 mm I.D.). A phosphate-citrate-acetonitrile buffer, utilizing ion suppression, is the mobile phase. The analytes are detectable at levels down to 10 ng/g. The analyte identity can be confirmed at 20 ng/g by the use of diode-array detection and spectral library comparison.

  4. The assembly of a GTPase–kinase signalling complex by a bacterial catalytic scaffold

    PubMed Central

    Selyunin, Andrey S.; Sutton, Sarah E.; Weigele, Bethany A.; Reddick, L. Evan; Orchard, Robert C.; Bresson, Stefan M.; Tomchick, Diana R.; Alto, Neal M.

    2011-01-01

    The fidelity and specificity of information flow within a cell is controlled by scaffolding proteins that assemble and link enzymes into signalling circuits1,2. These circuits can be inhibited by bacterial effector proteins that post-translationally modify individual pathway components3–6. However, there is emerging evidence that pathogens directly organize higher-order signalling networks through enzyme scaffolding7,8, and the identity of the effectors and their mechanisms of action are poorly understood. Here we identify the enterohaemorrhagic Escherichia coli O157:H7 type III effector EspG as a regulator of endomembrane trafficking using a functional screen, and report ADP-ribosylation factor (ARF) GTPases and p21-activated kinases (PAKs) as its relevant host substrates. The 2.5 Å crystal structure of EspG in complex with ARF6 shows how EspG blocks GTPase-activating-protein-assisted GTP hydrolysis, revealing a potent mechanism of GTPase signalling inhibition at organelle membranes. In addition, the 2.8 Å crystal structure of EspG in complex with the autoinhibitory Iα3-helix of PAK2 defines a previously unknown catalytic site in EspG and provides an allosteric mechanism of kinase activation by a bacterial effector. Unexpectedly, ARF and PAKs are organized on adjacent surfaces of EspG, indicating its role as a ‘catalytic scaffold’ that effectively reprograms cellular events through the functional assembly of GTPase-kinase signalling complex. PMID:21170023

  5. Nutritional performance and activity of some digestive enzymes of the cotton bollworm, Helicoverpa armigera, in response to seven tested bean cultivars.

    PubMed

    Namin, Foroogh Rahimi; Naseri, Bahram; Razmjou, Jabraeil

    2014-01-01

    Nutritional performance and activity of some digestive enzymes (protease and α-amylase) of Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) in response to feeding on bean (Phaseolus vulgaris L. (Fabales: Fabaceae)) cultivars (Shokufa, Akhtar, Sayyad, Naz, Pak, Daneshkadeh, and Talash) were evaluated under laboratory conditions (25 ± 1°C, 65 ± 5% RH, and a 16:8 L:D photoperiod). The highest and lowest respective values of approximate digestibility were observed when fourth, fifth, and sixth larval instar H. armigera were fed red kidney bean Akhtar and white kidney bean Daneshkadeh. The efficiency of conversion of ingested and digested food was highest when H. armigera was fed red kidney beans Akhtar and Naz and lowest when they were fed white kidney bean Pak. The highest protease activity of fifth instars was observed when they were fed red kidney bean Naz, and the highest amylase activity of fifth instars was observed when they were fed red kidney bean Sayyad. Sixth instar larvae that fed on red kidney bean Sayyad showed the highest protease activity. Larvae reared on common bean Talash and white kidney bean Pak showed the highest amylase activity. Among bean cultivars tested, red kidney bean Sayyad was the most unsuitable host for feeding H. armigera.

  6. Microtubule-dependent transport of vimentin filament precursors is regulated by actin and by the concerted action of Rho- and p21-activated kinases.

    PubMed

    Robert, Amélie; Herrmann, Harald; Davidson, Michael W; Gelfand, Vladimir I

    2014-07-01

    Intermediate filaments (IFs) form a dense and dynamic network that is functionally associated with microtubules and actin filaments. We used the GFP-tagged vimentin mutant Y117L to study vimentin-cytoskeletal interactions and transport of vimentin filament precursors. This mutant preserves vimentin interaction with other components of the cytoskeleton, but its assembly is blocked at the unit-length filament (ULF) stage. ULFs are easy to track, and they allow a reliable and quantifiable analysis of movement. Our results show that in cultured human vimentin-negative SW13 cells, 2% of vimentin-ULFs move along microtubules bidirectionally, while the majority are stationary and tightly associated with actin filaments. Rapid motor-dependent transport of ULFs along microtubules is enhanced ≥ 5-fold by depolymerization of actin cytoskeleton with latrunculin B. The microtubule-dependent transport of vimentin ULFs is further regulated by Rho-kinase (ROCK) and p21-activated kinase (PAK): ROCK inhibits ULF transport, while PAK stimulates it. Both kinases act on microtubule transport independently of their effects on actin cytoskeleton. Our study demonstrates the importance of the actin cytoskeleton to restrict IF transport and reveals a new role for PAK and ROCK in the regulation of IF precursor transport.-Robert, A., Herrmann, H., Davidson, M. W., and Gelfand, V. I. Microtubule-dependent transport of vimentin filament precursors is regulated by actin and by the concerted action of Rho- and p21-activated kinases.

  7. Performance of rapid tests and algorithms for HIV screening in Abidjan, Ivory Coast.

    PubMed

    Loukou, Y G; Cabran, M A; Yessé, Zinzendorf Nanga; Adouko, B M O; Lathro, S J; Agbessi-Kouassi, K B T

    2014-01-01

    Seven rapid diagnosis tests (RDTs) of HIV were evaluated by a panel group who collected serum samples from patients in Abidjan (HIV-1 = 203, HIV-2 = 25, HIV-dual = 25, HIV = 305). Kit performances were recorded after the reference techniques (enzyme-linked immunosorbent assay). The following RDTs showed a sensitivity of 100% and a specificity higher than 99%: Determine, Oraquick, SD Bioline, BCP, and Stat-Pak. These kits were used to establish infection screening strategies. The combination with 2 or 3 of these tests in series or parallel algorithms showed that series combinations with 2 tests (Oraquick and Bioline) and 3 tests (Determine, BCP, and Stat-Pak) gave the best performances (sensitivity, specificity, positive predictive value, and negative predictive value of 100%). However, the combination with 2 tests appeared to be more onerous than the combination with 3 tests. The combination with Determine, BCP, and Stat-Pak tests serving as a tiebreaker could be an alternative to the HIV/AIDS serological screening in Abidjan.

  8. Filamin, a synaptic organizer in Drosophila, determines glutamate receptor composition and membrane growth

    PubMed Central

    Lee, GaYoung; Schwarz, Thomas L

    2016-01-01

    Filamin is a scaffolding protein that functions in many cells as an actin-crosslinker. FLN90, an isoform of the Drosophila ortholog Filamin/cheerio that lacks the actin-binding domain, is here shown to govern the growth of postsynaptic membrane folds and the composition of glutamate receptor clusters at the larval neuromuscular junction. Genetic and biochemical analyses revealed that FLN90 is present surrounding synaptic boutons. FLN90 is required in the muscle for localization of the kinase dPak and, downstream of dPak, for localization of the GTPase Ral and the exocyst complex to this region. Consequently, Filamin is needed for growth of the subsynaptic reticulum. In addition, in the absence of filamin, type-A glutamate receptor subunits are lacking at the postsynapse, while type-B subunits cluster correctly. Receptor composition is dependent on dPak, but independent of the Ral pathway. Thus two major aspects of synapse formation, morphological plasticity and subtype-specific receptor clustering, require postsynaptic Filamin. DOI: http://dx.doi.org/10.7554/eLife.19991.001 PMID:27914199

  9. Technical failure of the pancreas after SPK transplant: are these patients good candidates for later pancreas retransplant?

    PubMed

    Wang, Shen-Nien; Sturdevant, Mark; Kandaswamy, Raja; Gruessner, Rainer G W; Sutherland, David E R; Humar, Abhinav

    2008-01-01

    Technical failure of the pancreas graft after a simultaneous pancreas-kidney (SPK) transplant is not uncommon, affecting roughly 10% of SPK recipients. These patients often recover with good kidney function, but have persistent issues related to their diabetes. The aim of this study was to determine if these patients were good candidates for a later pancreas retransplant. Outcomes were compared between 21 PASPK (pancreas after SPK) recipients and 361 recipients of a primary pancreas after kidney (PAK) transplant. Except for kidney graft source, there was no significant difference in the demographic characteristics between these two groups. In general, early surgical complications were more common in PASPK than PAK recipients (47.6% vs. 35.5%, p = 0.15), although the difference was not statistically significant. The incidence of acute rejection was no different between these two groups (28% vs. 33%, p = NS). At three yr post-transplant, patient and pancreas graft survival rates were also no different between the two groups (p = NS). The most common cause for graft loss in both groups was acute or chronic rejection. In conclusion, pancreas retransplant is a viable option for SPK recipients experiencing early technical failure of the pancreas graft. These recipients are not at higher immunologic risk vs. primary PAK recipients.

  10. Comparison of particulate verification techniques study

    NASA Astrophysics Data System (ADS)

    Rivera, Rachel

    2006-08-01

    The efficacy of five particulate verification techniques on four types of materials was studied. Statistical Analysis Software/JMP 6.0 was used to create a statistically valid design of experiments. In doing so, 35 witness coupons consisting of the four types of materials being studied, were intentionally contaminated with particulate fallout. Image Analysis was used to characterize the extent of particulate fallout on the coupons and was used to establish a baseline, or basis of comparison, against the five techniques that were studied. The five particulate verification techniques were the Tapelift, the Particulate Solvent Rinse, the GelPak lift, an in-line vacuum filtration probe, and the Infinity Focusing Microscope (IFM). The four types of materials consisted of magnesium flouride (MgF II) coated mirrors, composite coated silver aluminum (CCAg), Z93 and NS43G coated aluminum, and silicon (si) wafers. The vacuum probe was determined to be most effective for Z93, the tapelift or vacuum probe for MgF2, and the GelPak Lift for CCAg and si substrates. A margin of error for each technique, based on experimental data from two experiments, for si wafer substrates, yielded the following: Tapelift - 67%, Solvent Rinse - 58%, GelPak- 26%, Vacuum Probe - 93%, IFM-to be determined.

  11. Structural constraints on human immunodeficiency virus type 1 Nef function

    SciTech Connect

    Raney, Alexa; Shaw, Alice Y.; Foster, John L.; Garcia, J. Victor

    2007-11-10

    HIV-1 Nef is a multifunctional protein that exerts its activities through interactions with multiple cellular partners. Nef uses different domains and mechanisms to exert its functions including cell surface down-modulation of CD4 and MHC-I receptors and activation of the serine/threonine kinase PAK-2. We inserted tags at the C-terminus and proximal to the N-terminus of Nef and the effects on Nef's structure/function relationships were examined. We discovered significant defects in MHC-I down-modulation with the insertion of HA/FLAG tags at either region. We also found impaired PAK-2 activation with a C-terminal fusion with GFP. Interestingly, Nef-GFP and Nef-GH{sub 7} induced MHC-I down-modulation, suggesting that the negative charge of the HA/FLAG tag could contribute to the observed defect. Together, these observations highlight elements of Nef's functional complexity and demonstrate previously unsuspected structural requirements for PAK-2 activation and MHC-1 down-modulation in Nef's flexible N- and C-terminal regions.

  12. Pulsed Light-Emitting Diodes for a Higher Phytochemical Level in Microgreens.

    PubMed

    Vaštakaitė, Viktorija; Viršilė, Akvilė; Brazaitytė, Aušra; Samuolienė, Giedrė; Jankauskienė, Julė; Novičkovas, Algirdas; Duchovskis, Pavelas

    2017-08-09

    A novel research of pulsed light-emitting diode (LED) lighting versus continuous lighting was conducted by analyzing phytochemical levels in microgreens. Red pak choi (Brassica rapa var. chinensis), mustard (Brassica juncea L.), and tatsoi (Brassica rapa var. rosularis) were grown indoors under HPS lamps supplemented with monochromatic (455, 470, 505, 590, and 627 nm) LEDs [total photosynthetic photon flux density (PPFD) of 200 ± 10 μmol m(-2) s(-1), for 16 h day(-1)]. For pulsed light treatments, the frequencies at 2, 32, 256, and 1024 Hz with a duty cycle of 50% monochromatic LEDs were applied. The results were compared to those under the continuous light (0 Hz) condition in terms of total phenolic content, anthocyanins, and antiradical activity (DPPH). The summarized data suggested that pulsed light affected accumulation of secondary metabolites both positive and negative in microgreens. The significant differences in the response of phytochemicals between pulsed light at several frequencies and continuous light were determined. The most positive effects of 2, 256, and 1024 Hz for total phenolic compounds in mustard under all wavelength LEDs were achieved. The LED frequencies at 2 and 32 Hz were the most suitable for accumulation of anthocyanins in red pak choi and tatsoi. The highest antiradical activity under the treatments of 32, 256, and 1024 Hz in mustard and under the 2 Hz frequency in red pak choi and tatsoi was determined.

  13. Different Rho GTPase–dependent signaling pathways initiate sequential steps in the consolidation of long-term potentiation

    PubMed Central

    Chen, Lulu Y.; Sharma, Anupam; Liu, Jihua; Babayan, Alex H.; Gall, Christine M.; Lynch, Gary

    2009-01-01

    The releasable factor adenosine blocks the formation of long-term potentiation (LTP). These experiments used this observation to uncover the synaptic processes that stabilize the potentiation effect. Brief adenosine infusion blocked stimulation-induced actin polymerization within dendritic spines along with LTP itself in control rat hippocampal slices but not in those pretreated with the actin filament stabilizer jasplakinolide. Adenosine also blocked activity-driven phosphorylation of synaptic cofilin but not of synaptic p21-activated kinase (PAK). A search for the upstream origins of these effects showed that adenosine suppressed RhoA activity but only modestly affected Rac and Cdc42. A RhoA kinase (ROCK) inhibitor reproduced adenosine's effects on cofilin phosphorylation, spine actin polymerization, and LTP, whereas a Rac inhibitor did not. However, inhibitors of Rac or PAK did prolong LTP's vulnerability to reversal by latrunculin, a toxin which blocks actin filament assembly. Thus, LTP induction initiates two synaptic signaling cascades: one (RhoA-ROCK-cofilin) leads to actin polymerization, whereas the other (Rac-PAK) stabilizes the newly formed filaments. PMID:19596849

  14. Fission products from the damaged Fukushima reactor observed in Hungary.

    PubMed

    Bihari, Árpád; Dezső, Zoltán; Bujtás, Tibor; Manga, László; Lencsés, András; Dombóvári, Péter; Csige, István; Ranga, Tibor; Mogyorósi, Magdolna; Veres, Mihály

    2014-01-01

    Fission products, especially (131)I, (134)Cs and (137)Cs, from the damaged Fukushima Dai-ichi nuclear power plant (NPP) were detected in many places worldwide shortly after the accident caused by natural disaster. To observe the spatial and temporal variation of these isotopes in Hungary, aerosol samples were collected at five locations from late March to early May 2011: Institute of Nuclear Research, Hungarian Academy of Sciences (ATOMKI, Debrecen, East Hungary), Paks NPP (Paks, South-Central Hungary) as well as at the vicinity of Aggtelek (Northeast Hungary), Tapolca (West Hungary) and Bátaapáti (Southwest Hungary) settlements. In addition to the aerosol samples, dry/wet fallout samples were collected at ATOMKI, and airborne elemental iodine and organic iodide samples were collected at Paks NPP. The peak in the activity concentration of airborne (131)I was observed around 30 March (1-3 mBq m(-3) both in aerosol samples and gaseous iodine traps) with a slow decline afterwards. Aerosol samples of several hundred cubic metres of air showed (134)Cs and (137)Cs in detectable amounts along with (131)I. The decay-corrected inventory of (131)I fallout at ATOMKI was 2.1±0.1 Bq m(-2) at maximum in the observation period. Dose-rate contribution calculations show that the radiological impact of this event at Hungarian locations was of no considerable concern.

  15. MEK kinases are regulated by EGF and selectively interact with Rac/Cdc42.

    PubMed Central

    Fanger, G R; Johnson, N L; Johnson, G L

    1997-01-01

    MEK kinases (MEKKs) 1, 2, 3 and 4 are members of sequential kinase pathways that regulate MAP kinases including c-Jun NH2-terminal kinases (JNKs) and extracellular regulated kinases (ERKs). Confocal immunofluorescence microscopy of COS cells demonstrated differential MEKK subcellular localization: MEKK1 was nuclear and in post-Golgi vesicular-like structures; MEKK2 and 4 were localized to distinct Golgi-associated vesicles that were dispersed by brefeldin A. MEKK1 and 2 were activated by EGF, and kinase-inactive mutants of each MEKK partially inhibited EGF-stimulated JNK activity. Kinase-inactive MEKK1, but not MEKK2, 3 or 4, strongly inhibited EGF-stimulated ERK activity. In contrast to MEKK2 and 3, MEKK1 and 4 specifically associated with Rac and Cdc42 and kinase-inactive mutants blocked Rac/Cdc42 stimulation of JNK activity. Inhibitory mutants of MEKK1-4 did not affect p21-activated kinase (PAK) activation of JNK, indicating that the PAK-regulated JNK pathway is independent of MEKKs. Thus, in different cellular locations, specific MEKKs are required for the regulation of MAPK family members, and MEKK1 and 4 are involved in the regulation of JNK activation by Rac/Cdc42 independent of PAK. Differential MEKK subcellular distribution and interaction with small GTP-binding proteins provides a mechanism to regulate MAP kinase responses in localized regions of the cell and to different upstream stimuli. PMID:9305638

  16. Inhibition of Pseudomonas aeruginosa biofilm formation by 2,2’-bipyridyl, lipoic, kojic and picolinic acids

    PubMed Central

    Çevik, Kübra; Ulusoy, Seyhan

    2015-01-01

    Objective(s): The inhibitory effects of iron chelators, and FeCl3 chelation on biofilm formation and swarming motility were investigated against an opportunistic human pathogen Pseudomonas aeruginosa. Materials and Methods: The inhibitory activity of 2,2’-bipyridyl, lipoic acid, kojic acid and picolinic acid on biofilm formation of P. aeruginosa strain PAO1 and three clinical isolates (P. aeruginosa PAK01, P. aeruginosa PAK02 and P. aeruginosa PAK03) were investigated, based on crystal violet assay, and swarming motility test. Results: The kojic, lipoic and picolinic acid inhibited biofilm formation by 5-33% in all tested P. aeruginosa isolates. When chelated iron was added, biofilm inhibition rates were determined to be 39-57%. Among the tested chelators against P. aeruginosa, lipoic acid (84%) and kojic acid (68%) presented the highest inhibition of swarming motility. This is the first study to report the inhibitory effect of lipoic acid on biofilm formation and swarming motility of P. aeruginosa. Conclusion: It is considered that lipoic and picolinic acids can serve as alternatives for the treatment of the P. aeruginosa infections by inhibiting biofilm formation. PMID:26557964

  17. Glycemic Control in Simultaneous Islet-Kidney Versus Pancreas-Kidney Transplantation in Type 1 Diabetes: A Prospective 13-Year Follow-up.

    PubMed

    Lehmann, Roger; Graziano, Jessica; Brockmann, Jens; Pfammatter, Thomas; Kron, Philipp; de Rougemont, Olivier; Mueller, Thomas; Zuellig, Richard A; Spinas, Giatgen A; Gerber, Philipp A

    2015-05-01

    In patients with type 1 diabetes and end-stage renal disease, combined transplantation of a kidney together with a pancreas or isolated pancreatic islets are options to improve glycemic control. The aim of this study was to compare their long-term outcome with regard to metabolic control and surgical complication rate, as well as function of the transplanted kidney. We conducted a prospective cohort study in consecutive patients receiving either a pancreas or islet transplant simultaneously with or after kidney transplantation (simultaneous pancreas-kidney [SPK]/pancreas-after-kidney [PAK] or simultaneous islet-kidney [SIK]/islet-after-kidney [IAK] transplantation). Ninety-four patients who had undergone SPK/PAK transplantation were compared with 38 patients who had undergone SIK/IAK transplantation over a period of up to 13 years. HbA1c levels declined from 7.8 ± 1.3% (62 ± 14 mmol/mol) to 5.9 ± 1.1% (41 ± 12 mmol/mol), and from 8.0 ± 1.3% (64 ± 14 mmol/mol) to 6.5 ± 1.1% (48 ± 12 mmol/mol), respectively, in the SPK/PAK and SIK/IAK groups (P < 0.001 for both) and remained stable during follow-up, despite a reduction in the rate of severe hypoglycemia by >90%. The 5-year insulin independence rate was higher in the SPK/PAK group (73.6 vs. 9.3% in the SIK/IAK group), as was the rate of relaparotomy after transplantation (41.5 vs. 10.5% in the SIK/IAK group). There was no difference in the rate of kidney function decline. During a long-term follow-up, SPK/PAK transplantation as well as SIK/IAK transplantation resulted in a sustained improvement of glycemic control with a slightly higher glycated hemoglobin level in the SIK/IAK group. While insulin independence is more common in whole-organ pancreas recipients, islet transplantation can be conducted with a much lower surgical complication rate and no difference in kidney function decline. © 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is

  18. Influence of thermal maturity on the hydrogen isotope content of extractable hydrocarbons

    NASA Astrophysics Data System (ADS)

    Radke, J.; Bechtel, A.; Püttmann, W.; Gleixner, G.

    2003-04-01

    Based on hydrogen isotope analysis of hydrocarbons from recent sediments it is suggested that compound specific hydrogen isotope ratios are a new proxy to reconstruct the palaeoclimate (Sauer et al., 2001). However, it remains unclear if transformation of carbon bound hydrogen with environmental water during maturation or thermal methanogenesis might influence the observed values. Short-term experiments excluded exchange reactions of deuterium from alkanes (Schimmelmann et al., 1999), however, thermally stressed kerogens are enriched in deuterium (Schoell, 1984). Therefore, we investigated the influence of maturity on the deltaD-values of alkanes and acyclic isoprenoids. In the Kupferschiefer horizon from the Polish Zechstein Basin thermal maturity of organic matter is correlated to burial depth yielding a natural long-term exchange experiment. The deltaD-values of extracted hydrocarbons linearly correlated with thermal maturity. These results enable the correction of deltaD values from biomarkers with known maturity and therefore expanding palaeoclimatic reconstructions using deltaD values to the geological past. References: SAUER, P.E., EGLINTON, T.I., HAYES, J.M., SCHIMMELMANN, A. &SESSIONS, A.L. (2001) Compound specific D/H ratios of lipid biomarkers from sediments as a proxy for environmental and climatic conditions. Geochimica et Cosmochimica Acta, 65(2), 213-222. SCHIMMELMANN, A., LEWAN, M.D. &WINTSCH, R.P. (1999) D/H isotope ratios of kerogen, bitumen, oil, and water in hydrous pyrolysis of source rocks containing kerogen types I, II, IIS, and III. Geochimica et Cosmochimica Acta, 63(22), 3751-3766. SCHOELL, M. (1984) Wasserstoff- und Kohlenstoffisotope in organischen Substanzen, Erdölen und Erdgasen. Schweitzerbart'sche Verlagsbuchhandlung, Stuttgart. Reihe D (67), 161pp.

  19. Alternative anaerobic enrichments to the bacteriological analytical manual culture method for isolation of Shigella sonnei from selected types of fresh produce.

    PubMed

    Jacobson, Andrew P; Thunberg, Richard L; Johnson, Mildred L; Hammack, Thomas S; Andrews, Wallace H

    2004-01-01

    Alternative methods of reducing oxygen during anaerobic enrichment in the Bacteriological Analytical Manual (BAM) Shigella culture method were evaluated and compared to the current and less practical GasPak method. The alternative anaerobic methods included the use of reducing agents in Shigella broth and reducing culture container headspace volume to minimize atmospheric effects on oxygen concentration in Shigella broth during enrichment. The reducing agents evaluated were sodium thioglycollate, L-cystine, L-cysteine, titanium(III) citrate, and dithiothreitol, each at concentrations of 0.1, 0.05, and 0.01%. The use of Oxyrase for Broth with the enrichment medium (Shigella broth) was evaluated at concentrations of 10, 20 and 30 microL/mL. Recoveries of chill- and freeze-stressed S. sonnei strains 357 and 20143 were determined with each anaerobic method, including the GasPak method, using inoculation levels ranging from 10(0)to 10(3) cells. For each anaerobic method, strain, inoculation level, and stress type, 5 replicate enrichments were evaluated by streaking to MacConkey agar for isolation. The numbers of cultures with each method from which S. sonnei was isolated were used to compare the alternative anaerobic methods to the GasPak method. The alternative anaerobic method with which chill- and freeze-stressed S. sonnei strains 357 and 20143 were isolated most consistently was the use of Oxyrase for Broth in Shigella broth at a concentration of 20 microL/mL. This method was compared to the GasPak anaerobic method in evaluations on the recovery of S. sonnei strains 357 and 20143 from artificially contaminated test portions of parsley, cilantro, green onions, strawberries, carrots, and celery. A third anaerobic method included the use of 0.5 cm mineral oil overlay on cultures containing Oxyrase for Broth at concentrations of 20 microL/mL. Recovery rates of strain 357 were significantly greater (p < 0.05) with the GasPak method than with Oxyrase for Broth, with and

  20. Applications of GPS-tracked personal and fixed-location PM(2.5) continuous exposure monitoring.

    PubMed

    Sloan, Chantel D; Philipp, Tyler J; Bradshaw, Rebecca K; Chronister, Sara; Barber, W Bradford; Johnston, James D

    2016-01-01

    Continued development of personal air pollution monitors is rapidly improving government and research capabilities for data collection. In this study, we tested the feasibility of using GPS-enabled personal exposure monitors to collect personal exposure readings and short-term daily PM2.5 measures at 15 fixed locations throughout a community. The goals were to determine the accuracy of fixed-location monitoring for approximating individual exposures compared to a centralized outdoor air pollution monitor, and to test the utility of two different personal monitors, the RTI MicroPEM V3.2 and TSI SidePak AM510. For personal samples, 24-hr mean PM2.5 concentrations were 6.93 μg/m³ (stderr = 0.15) and 8.47 μg/m³ (stderr = 0.10) for the MicroPEM and SidePak, respectively. Based on time-activity patterns from participant journals, exposures were highest while participants were outdoors (MicroPEM = 7.61 µg/m³, stderr = 1.08, SidePak = 11.85 µg/m³, stderr = 0.83) or in restaurants (MicroPEM = 7.48 µg/m³, stderr = 0.39, SidePak = 24.93 µg/m³, stderr = 0.82), and lowest when participants were exercising indoors (MicroPEM = 4.78 µg/m³, stderr = 0.23, SidePak = 5.63 µg/m³, stderr = 0.08). Mean PM(2.5) at the 15 fixed locations, as measured by the SidePak, ranged from 4.71 µg/m³ (stderr = 0.23) to 12.38 µg/m³ (stderr = 0.45). By comparison, mean 24-h PM(2.5) measured at the centralized outdoor monitor ranged from 2.7 to 6.7 µg/m³ during the study period. The range of average PM(2.5) exposure levels estimated for each participant using the interpolated fixed-location data was 2.83 to 19.26 µg/m³ (mean = 8.3, stderr = 1.4). These estimated levels were compared with average exposure from personal samples. The fixed-location monitoring strategy was useful in identifying high air pollution microclimates throughout the county. For 7 of 10 subjects, the fixed-location monitoring strategy more closely approximated individuals' 24-hr breathing zone exposures than