DOCSLIB.ORG

PEA-15 Potentiates H-Ras-Mediated Epithelial Cell Transformation Through Phospholipase D

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
PEA-15 Potentiates H-Ras-Mediated Epithelial Cell Transformation Through Phospholipase D Oncogene (2012) 31, 3547–3560 & 2012 Macmillan Publishers Limited All rights reserved 0950-9232/12 www.nature.com/onc ORIGINAL ARTICLE PEA-15 potentiates H-Ras-mediated epithelial cell transformation through phospholipase D FJ Sulzmaier1,2, MKG Valmiki1,2, DA Nelson3, MJ Caliva1,4, D Geerts5, ML Matter4, EP White3 and JW Ramos1,2 1Cancer Biology Program, University of Hawaii Cancer Center, University of Hawaii at Manoa, Honolulu, HI, USA; 2Department of Molecular Biosciences and Bioengineering, University of Hawai’i at Manoa, Honolulu, HI, USA; 3The Cancer Institute of New Jersey, New Brunswick, NJ, USA; 4Department of Cell and Molecular Biology, John A Burns School of Medicine, University of Hawaii at Manoa, Honolulu, HI, USA and 5Department of Pediatric Oncology/Hematology, Sophia Children’s Hospital, Rotterdam, The Netherlands The small GTPase H-Ras is a proto-oncogene that Introduction activates a variety of different pathways including the extracellular-signal-regulated kinase (ERK)/mitogen- The small GTPase Ras is frequently mutated in cancer activated protein kinase pathway. H-Ras is mutated in and contributes to transformation and tumor progres- many human malignancies, and these mutations cause the sion (Dunn et al., 2005; Tan et al., 2005). Physiologi- protein to be constitutively active. Phosphoprotein cally, Ras is involved in a variety of signaling pathways enriched in astrocytes, 15 kDa (PEA-15) blocks ERK- regulating different outcomes (Cordova-Alarcon et al., dependent gene transcription and inhibits proliferation 2005). Guanosine-triphosphate-loaded Ras activates by sequestering ERK in the cytoplasm. We therefore many pathways, including the mitogen-activated protein investigated whether PEA-15 influences H-Ras-mediated kinase/extracellular-signal-regulated kinase (MAPK/ transformation. We found that PEA-15 does not block H- ERK) pathway, the PI3 kinase pathway and the Ras-activated proliferation when H-Ras is constitutively RalGDS pathway, all of which have known roles in active. We show instead that in H-Ras-transformed mouse cancer via diverse effects both on a cellular and kidney epithelial cells, co-expression of PEA-15 resulted physiological level. For example, Ras/MAPK/ERK in enhanced soft agar colony growth and increased tumor signaling modulates transcription, cell cycle progression, growth in vivo. Overexpression of both H-Ras and PEA- proliferation and differentiation, as well as senescence 15 resulted in accelerated G1/S cell cycle transition and and cell death (Ramos, 2008; Castaneda et al., 2010; increased activation of the ERK signaling pathway. PEA- Ferro and Trabalzini, 2010). 15 mediated these effects through activation of its binding Mutations in Ras genes such as K-Ras, N-Ras or partner phospholipase D1 (PLD1). Inhibition of PLD1 or H-Ras can be found in various tumors and are often interference with PEA-15/PLD1 binding blocked PEA- associated with poor prognosis (Cordova-Alarcon et al., 15’s ability to increase ERK activation. Our findings 2005; Zeng et al., 2010). Mutated H-Ras is associated reveal a novel mechanism by which PEA-15 positively with elevated expression of key cell cycle regulatory regulates Ras/ERK signaling and increases the prolifera- proteins, such as cyclin D1 and cdk4, in in vivo models tion of H-Ras-transformed epithelial cells through of oral squamous-cell carcinoma of the head and neck enhanced PLD1 expression and activation. Thus, our region (Sathyan et al., 2007). Cordova-Alarcon et al. work provides a surprising mechanism by which PEA-15 (2005) showed that mutational activation of H-Ras augments H-Ras-driven transformation. These data reveal accelerates G1/S cell cycle transition in a human cervical that PEA-15 not only suppresses ERK signaling and cancer-derived cell line in vitro and has an important tumorigenesis but also alternatively enhances tumorigenesis role in the development of cervical tumors. Oncogenic in the context of active Ras. H-Ras also transforms NIH3T3 fibroblasts, at least in Oncogene (2012) 31, 3547–3560; doi:10.1038/onc.2011.514; part, through upregulation of cyclin D1 due to an published online 21 November 2011 overexpression of Kruppel-like factor 5 (Nandan et al., Keywords: PEA-15; PLD; cell cycle; H-Ras; cell 2004). Interestingly, mutations in H-Ras not only affect transformation gene transcription but also contribute to tumor cell transformation by inducing anti-apoptotic signals. In immortal baby mouse kidney epithelial cells (iBMK), a constitutively activated form of H-Ras inhibited pacli- Correspondence: Dr JW Ramos, Cancer Biology Program, University taxel-induced accumulation of the proapoptotic BH3- of Hawaii Cancer Center, University of Hawaii at Manoa, Biosciences only protein BIM, thereby preventing BIM-dependent Building, Room 241C, 651 Ilalo Street, Honolulu, HI 96813, USA. apoptosis. In this model, H-Ras to ERK signaling E-mail: [email protected] Received 28 March 2011; revised 18 September 2011; accepted 9 October resulted in phosphorylation of BIM, leading to its 2011; published online 21 November 2011 proteasomal degradation (Tan et al., 2005). PEA-15 enhances H-Ras-mediated cell transformation FJ Sulzmaier et al 3548 The small death effector domain-containing protein PLD1 mutant preventing binding to PEA-15 blocked phosphoprotein enriched in astrocytes, 15 kDa (PEA-15) activation of the ERK/MAPK pathway. We have thus is a molecular scaffold that regulates several pathways identified a novel mechanism by which PEA-15 can including the Ras/MAPK/ERK signaling cascade positively regulate Ras/ERK/MAPK signaling and (Ramos, 2008; Revet et al., 2008). It has been shown increase the proliferation of H-Ras-transformed epithe- to bind to ERK and prevent ERK-mediated gene lial cells. The enhancement of Ras signaling was in part transcription (Formstecher et al., 2001; Whitehurst due to increased activity of PLD1 resulting from PEA-15 et al., 2004). This can result in a downregulation of cell expression. This is the first evidence to link PEA-15 proliferation, as shown in murine T cells (Pastorino regulation of Ras signaling to effects on PLD. et al., 2010). Interaction of PEA-15 with specific partners is controlled by its phosphorylation and subcellular localization. These factors therefore deter- Results mine the effects of PEA-15 on cellular signaling processes (Renganathan et al., 2005). Phospholipase PEA-15 enhances H-Ras-mediated epithelial cell D (PLD) binds to PEA-15 and is a widely expressed transformation in vitro in an adhesion-independent manner enzyme that catalyzes the hydrolysis of phosphatidyl- Expression of oncogenic H-Ras leads to a transformed choline to phosphatidic acid (PA) and choline (Wang phenotype in iBMK cells, resulting in adhesion- et al., 1999; Yang et al., 2001). Although the independent colony formation in soft agar assays physiological function of this interaction remains (Degenhardt and White, 2006). PEA-15 negatively regulates elusive, PEA-15 stabilizes PLD expression and may H-Ras to ERK signaling and blocks H-Ras-mediated activate it by a yet undefined mechanism (Zhang et al., proliferation signals by sequestering ERK in the 2000). PLD has been shown to modulate the Ras–ERK cytoplasm (Formstecher et al., 2001; Pastorino et al., pathway and regulate the cell cycle and cell prolifera- 2010). The ability of cells to grow in suspension is a tion (Yang et al., 2001; Donaldson, 2009). PLD activity hallmark of transformation. We therefore analyzed is crucial for Ras activation in NIH3T3 cells, where whether co-expression of PEA-15 would abolish PLD-produced PA is necessary for the recruitment of H-Ras-mediated transformation in iBMK cells and son of sevenless (SOS), a direct activator of Ras, to the inhibit Ras-induced growth in soft agar assays. Surprisingly, membrane (Zhao et al., 2007). In addition, in Rat-1 stable co-expression of PEA-15 in the presence of active fibroblasts PA can associate with Raf-1 and target it to H-Ras enhanced soft agar colony growth. Cells stably the membrane. This Ras-independent recruitment is overexpressing both H-Ras and PEA-15 formed important for the subsequent activation of Raf-1 by significantly (Po0.0003) more colonies in soft agar Ras in these cells and is required for further down- compared with cells expressing H-Ras alone (Figure 1a). stream signaling (Rizzo et al., 2000). Elevated PLD Neither the control cell lines nor the cell lines stably activity is found in several tumors, including breast, expressing PEA-15 alone were able to form soft agar gastric and renal cancers (Uchida et al., 1997, 1999; colonies. Co-expression of PEA-15 in cells expressing Noh et al., 2000; Zhao et al.,2000).PLDhasalsobeen the constitutively active form of H-Ras not only shown to have transforming properties in fibroblasts augmented the number of soft agar colonies formed, with aberrant tyrosine kinase activity and may con- but also increased the diameter of the colonies (Figures tribute to tumor progression in this context (Lu et al., 1b and c). We verified the overexpression of PEA-15 2000; Joseph et al.,2001).Furthermore,inv-Raf and H-Ras after stable transfection by immunoblot transformed NIH3T3 cells, increased PLD activity was (Figure 1d). able to overcome Raf-induced cell cycle arrest (Frankel et al.,1999;Josephet al., 2002). PEA-15 promotes G1- to S-phase transition and does not In this study, we analyzed the effects of PEA-15 alter anoikis expression on H-Ras-transformed mouse kidney epithe- The Ras/MAPK/ERK signaling cascade has been lial cells (iBMK). Surprisingly, we found that PEA-15 shown to be involved in the regulation of cell cycle does not block H-Ras proliferation signals as previously progression both in physiological and pathological described, but in this cellular context enhanced H-Ras- conditions (Cordova-Alarcon et al., 2005; Sathyan driven transformation independent of apoptosis. Co- et al., 2007; Viparelli et al., 2008). We therefore analyzed expression of H-Ras and PEA-15 resulted in enhanced whether the increased colony formation in soft agar was tumor formation in vivo and increased colony formation due to influences of PEA-15 expression on cell cycle in soft agar assays in vitro.
Load more

Recommended publications
  • ATP-Induced Focal Adhesion Kinase Activity Is Negatively Modulated by Phospholipase D2 in PC12 Cells
    EXPERIMENTAL and MOLECULAR MEDICINE, Vol. 33, No. 3, 150-155, September 2001 ATP-induced focal adhesion kinase activity is negatively modulated by phospholipase D2 in PC12 cells Yoe-Sik Bae1 and Sung Ho Ryu1,2 Introduction 1 Division of Molecular and Life Sciences, Pohang University of Purinergic receptors have been reported to play impor- Science and Technology, Pohang 790-784, Korea tant roles on the regulation of neuronal cell functions 2 Corresponding author: Tel, +82-54-279-2292; (Communi et al., 2000; Di Iorio et al., 1998). ATP, a Fax, +82-54-279-2199; E-mail, [email protected] ligand for the receptors modulate various cellular re- sponses such as mitogenic and morphogenic activity in Accepted 18 September 2001 PC12 rat pheochromocytoma cells (Neary et al., 1996; Soltoff et al., 1998; Schindelholz et al., 2000). Stimu- Abbreviations: Fak, focal adhesion kinase; PLD, phospholipase D; lation of cells with ATP induces tyrosine phosphorylation PA, phosphatidic acid; PC, phosphatidylcholine; DAG, diacylglyc- of several cytoskeletal proteins and focal adhesion erol; PBt, phosphatidylbutanol; PKC, protein kinase C; PAP, phos- molecules such as focal adhesion kinase (Fak), proline- phatidic acid phosphohydrolase rich tyrosine kinase (Pyk2), and paxillin (Soltoff et al., 1998; Schindelholz et al., 2000). Since these cytosk- eleton-associated proteins have been regarded as important factors for the regulation of neuronal cell Abstract functions, the study on the regulatory mechanism for the proteins remains an important issue. Extracellular ATP has been known to modulate vari- Phospholipase D (PLD) catalyzes the hydrolysis of ous cellular responses including mitogenesis, secre- phosphatidylcholine (PC) into phosphatidic acid (PA) tion and morphogenic activity in neuronal cells.
    [Show full text]
  • PARSANA-DISSERTATION-2020.Pdf
    DECIPHERING TRANSCRIPTIONAL PATTERNS OF GENE REGULATION: A COMPUTATIONAL APPROACH by Princy Parsana A dissertation submitted to The Johns Hopkins University in conformity with the requirements for the degree of Doctor of Philosophy Baltimore, Maryland July, 2020 © 2020 Princy Parsana All rights reserved Abstract With rapid advancements in sequencing technology, we now have the ability to sequence the entire human genome, and to quantify expression of tens of thousands of genes from hundreds of individuals. This provides an extraordinary opportunity to learn phenotype relevant genomic patterns that can improve our understanding of molecular and cellular processes underlying a trait. The high dimensional nature of genomic data presents a range of computational and statistical challenges. This dissertation presents a compilation of projects that were driven by the motivation to efficiently capture gene regulatory patterns in the human transcriptome, while addressing statistical and computational challenges that accompany this data. We attempt to address two major difficulties in this domain: a) artifacts and noise in transcriptomic data, andb) limited statistical power. First, we present our work on investigating the effect of artifactual variation in gene expression data and its impact on trans-eQTL discovery. Here we performed an in-depth analysis of diverse pre-recorded covariates and latent confounders to understand their contribution to heterogeneity in gene expression measurements. Next, we discovered 673 trans-eQTLs across 16 human tissues using v6 data from the Genotype Tissue Expression (GTEx) project. Finally, we characterized two trait-associated trans-eQTLs; one in Skeletal Muscle and another in Thyroid. Second, we present a principal component based residualization method to correct gene expression measurements prior to reconstruction of co-expression networks.
    [Show full text]
  • PLCG1) Mutations in Sézary Syndrome
    This electronic thesis or dissertation has been downloaded from the King’s Research Portal at https://kclpure.kcl.ac.uk/portal/ Functional interrogations of Phospholipase C Gamma 1 (PLCG1) mutations in Sézary Syndrome Patel, Varsha Maheshkumar Awarding institution: King's College London The copyright of this thesis rests with the author and no quotation from it or information derived from it may be published without proper acknowledgement. END USER LICENCE AGREEMENT Unless another licence is stated on the immediately following page this work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International licence. https://creativecommons.org/licenses/by-nc-nd/4.0/ You are free to copy, distribute and transmit the work Under the following conditions: Attribution: You must attribute the work in the manner specified by the author (but not in any way that suggests that they endorse you or your use of the work). Non Commercial: You may not use this work for commercial purposes. No Derivative Works - You may not alter, transform, or build upon this work. Any of these conditions can be waived if you receive permission from the author. Your fair dealings and other rights are in no way affected by the above. Take down policy If you believe that this document breaches copyright please contact [email protected] providing details, and we will remove access to the work immediately and investigate your claim. Download date: 11. Oct. 2021 Functional interrogations of Phospholipase C Gamma 1 (PLCG1) mutations in Sézary Syndrome Varsha Maheshkumar Patel Skin Tumour Unit, St John’s Institute of Dermatology, School of Basic and Medical Biosciences, King’s College London.
    [Show full text]
  • (4,5) Bisphosphate-Phospholipase C Resynthesis Cycle: Pitps Bridge the ER-PM GAP
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by UCL Discovery Topological organisation of the phosphatidylinositol (4,5) bisphosphate-phospholipase C resynthesis cycle: PITPs bridge the ER-PM GAP Shamshad Cockcroft and Padinjat Raghu* Dept. of Neuroscience, Physiology and Pharmacology, Division of Biosciences, University College London, London WC1E 6JJ, UK; *National Centre for Biological Sciences, TIFR-GKVK Campus, Bellary Road, Bangalore 560065, India Address correspondence to: Shamshad Cockcroft, University College London UK; Phone: 0044-20-7679-6259; Email: [email protected] Abstract Phospholipase C (PLC) is a receptor-regulated enzyme that hydrolyses phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) at the plasma membrane (PM) triggering three biochemical consequences, the generation of soluble inositol 1,4,5-trisphosphate (IP3), membrane– associated diacylglycerol (DG) and the consumption of plasma membrane PI(4,5)P2. Each of these three signals triggers multiple molecular processes impacting key cellular properties. The activation of PLC also triggers a sequence of biochemical reactions, collectively referred to as the PI(4,5)P2 cycle that culminates in the resynthesis of this lipid. The biochemical intermediates of this cycle and the enzymes that mediate these reactions are topologically distributed across two membrane compartments, the PM and the endoplasmic reticulum (ER). At the plasma membrane, the DG formed during PLC activation is rapidly converted to phosphatidic acid (PA) that needs to be transported to the ER where the machinery for its conversion into PI is localised. Conversely, PI from the ER needs to be rapidly transferred to the plasma membrane where it can be phosphorylated by lipid kinases to regenerate PI(4,5)P2.
    [Show full text]
  • Identification of Lithium-Regulated Genes in Cultured Lymphoblasts of Lithium Responsive Subjects with Bipolar Disorder
    Neuropsychopharmacology (2004) 29, 799–804 & 2004 Nature Publishing Group All rights reserved 0893-133X/04 $25.00 www.neuropsychopharmacology.org Identification of Lithium-Regulated Genes in Cultured Lymphoblasts of Lithium Responsive Subjects with Bipolar Disorder 1 1 1 2 3 4 Xiujun Sun , L Trevor Young , Jun-Feng Wang , Paul Grof , Gustavo Turecki , Guy A Rouleau ,5 and Martin Alda* 1Department of Psychiatry, University of Toronto, Toronto, Canada; 2Department of Psychiatry, University of Ottawa, Ottawa, Canada; 3 4 Department of Psychiatry, McGill University, Montreal, Canada; Center for Research in Neuroscience, McGill University, Montreal, Canada; 5 Department of Psychiatry, Dalhousie University, Halifax, Canada Lithium, a common drug for the treatment of bipolar disorder (BD), requires chronic administration to prevent recurrences of the illness. The necessity for long-term treatment suggests that changes in genes expression are involved in the mechanism of its action. We studied effects of lithium on gene expression in lymphoblasts from BD patients, all excellent responders to lithium prophylaxis. Gene expression was analyzed using cDNA arrays that included a total of 2400 cDNAs. We found that chronic lithium treatment at a therapeutically relevant concentration decreased the expression of seven genes in lymphoblasts from lithium responders. Five of these candidate lithium- regulated genes, including alpha1B-adrenoceptor (a1B-AR), acetylcholine receptor protein alpha chain precursor (ACHR), cAMP- 0 0 dependent 3 ,5 -cyclic phosphodiesterase 4D (PDE4D), substance-P receptor (SPR), and ras-related protein RAB7, were verified by Northern blotting analysis in lithium responders. None of these genes were regulated by lithium in healthy control subjects. When we compared the expression of these five genes between bipolar subjects and healthy control subjects at baseline, prior to lithium administration, we found that a1B-AR gene expression was higher in bipolar subjects than in healthy control subjects.
    [Show full text]
  • The Protein Phosphatase PP2A Plays Multiple Roles in Plant Development by Regulation of Vesicle Traffic—Facts and Questions
    International Journal of Molecular Sciences Review The Protein Phosphatase PP2A Plays Multiple Roles in Plant Development by Regulation of Vesicle Traffic—Facts and Questions Csaba Máthé *, Márta M-Hamvas, Csongor Freytag and Tamás Garda Department of Botany, Faculty of Science and Technology, University of Debrecen, H-4032 Debrecen, Hungary; [email protected] (M.M.-H.); [email protected] (C.F.); [email protected] (T.G.) * Correspondence: [email protected] Abstract: The protein phosphatase PP2A is essential for the control of integrated eukaryotic cell functioning. Several cellular and developmental events, e.g., plant growth regulator (PGR) mediated signaling pathways are regulated by reversible phosphorylation of vesicle traffic proteins. Reviewing present knowledge on the relevant role of PP2A is timely. We discuss three aspects: (1) PP2A regulates microtubule-mediated vesicle delivery during cell plate assembly. PP2A dephosphorylates members of the microtubule associated protein family MAP65, promoting their binding to microtubules. Regulation of phosphatase activity leads to changes in microtubule organization, which affects vesicle traffic towards cell plate and vesicle fusion to build the new cell wall between dividing cells. (2) PP2A-mediated inhibition of target of rapamycin complex (TORC) dependent signaling pathways contributes to autophagy and this has possible connections to the brassinosteroid signaling pathway. (3) Transcytosis of vesicles transporting PIN auxin efflux carriers. PP2A regulates vesicle localization and recycling of PINs related to GNOM (a GTP–GDP exchange factor) mediated pathways. The proper intracellular traffic of PINs is essential for auxin distribution in the plant body, thus in whole Citation: Máthé, C.; M-Hamvas, M.; plant development.
    [Show full text]
  • 1. Introduction
    1 1. Introduction 1.1 Phospholipids in cellular signalling Signalling through lipid metabolites has gained a lot of attention since the finding of MABEL R. and LOWELL E. HOKIN (1953) that [32P]-orthophosphate is incorporated into phosphatidylinositol upon acetylcholine stimulation. This reaction step is now recognised as part of the phosphoinositide cascade. Its activation results in the release of D-meso-inositol-1,4,5-trisphosphate and sn-1,2-diacylglycerol (DAG) from phosphatidyl- inositol-4,5-bisphosphate (PIP2) (BERRIDGE and IRVINE, 1984). These second messengers are products of a phospholipase C (PLC), other second messengers are generated by phospholi- pase A2 (PLA2) and phospholipase D (PLD) (Fig. 1). O O O- P R3 R1 O O O phospholipase D R2 O phospholipase C O phospholipase A2 Figure 1. Mechanism of action of phospholipases involved in signalling on phospholipids. R1 is mostly a saturated, R2 an unsaturated carbon chain and R3 is a polar head group like choline, ethanolamine or inositol. Activation of the phosphoinositide cascade occurs through many cell surface receptors and has been characterised in great detail. Its second messenger D-meso-inositol-1,4,5-tris- phosphate releases Ca2+ from intracellular stores whereas sn-1,2-diacylglycerol activates protein kinase C (PKC). At present, three PIP2-specific phospholipase C families are characte- rised, referred to as phospholipase Cb, phospholipase Cg and phospholipase Cd (REBECCHI and PENTYALA, 2000). Each of these families comprises several members and is used in different signal transduction pathways. Receptors coupled to heterotrimeric G-proteins activate phospholipase Cb through the latter while activation of phospholipase Cg is mediated via tyrosine kinase receptors.
    [Show full text]
  • Aneuploidy: Using Genetic Instability to Preserve a Haploid Genome?
    Health Science Campus FINAL APPROVAL OF DISSERTATION Doctor of Philosophy in Biomedical Science (Cancer Biology) Aneuploidy: Using genetic instability to preserve a haploid genome? Submitted by: Ramona Ramdath In partial fulfillment of the requirements for the degree of Doctor of Philosophy in Biomedical Science Examination Committee Signature/Date Major Advisor: David Allison, M.D., Ph.D. Academic James Trempe, Ph.D. Advisory Committee: David Giovanucci, Ph.D. Randall Ruch, Ph.D. Ronald Mellgren, Ph.D. Senior Associate Dean College of Graduate Studies Michael S. Bisesi, Ph.D. Date of Defense: April 10, 2009 Aneuploidy: Using genetic instability to preserve a haploid genome? Ramona Ramdath University of Toledo, Health Science Campus 2009 Dedication I dedicate this dissertation to my grandfather who died of lung cancer two years ago, but who always instilled in us the value and importance of education. And to my mom and sister, both of whom have been pillars of support and stimulating conversations. To my sister, Rehanna, especially- I hope this inspires you to achieve all that you want to in life, academically and otherwise. ii Acknowledgements As we go through these academic journeys, there are so many along the way that make an impact not only on our work, but on our lives as well, and I would like to say a heartfelt thank you to all of those people: My Committee members- Dr. James Trempe, Dr. David Giovanucchi, Dr. Ronald Mellgren and Dr. Randall Ruch for their guidance, suggestions, support and confidence in me. My major advisor- Dr. David Allison, for his constructive criticism and positive reinforcement.
    [Show full text]
  • Substantial Conformational Change Mediated by Charge-Triad Residues of the Death Effector Domain in Protein-Protein Interactions
    Substantial Conformational Change Mediated by Charge-Triad Residues of the Death Effector Domain in Protein-Protein Interactions Edward C. Twomey1,2,3, Dana F. Cordasco1,2, Stephen D. Kozuch2, Yufeng Wei1* 1 Institute of NeuroImmune Pharmacology, Seton Hall University, South Orange, New Jersey, United States of America, 2 Department of Chemistry and Biochemistry, Seton Hall University, South Orange, New Jersey, United States of America, 3 Integrated Program in Cellular, Molecular, and Biomedical Studies, Columbia University, New York, New York, United States of America Abstract Protein conformational changes are commonly associated with the formation of protein complexes. The non-catalytic death effector domains (DEDs) mediate protein-protein interactions in a variety of cellular processes, including apoptosis, proliferation and migration, and glucose metabolism. Here, using NMR residual dipolar coupling (RDC) data, we report a conformational change in the DED of the phosphoprotein enriched in astrocytes, 15 kDa (PEA-15) protein in the complex with a mitogen-activated protein (MAP) kinase, extracellular regulated kinase 2 (ERK2), which is essential in regulating ERK2 cellular distribution and function in cell proliferation and migration. The most significant conformational change in PEA-15 happens at helices a2, a3, and a4, which also possess the highest flexibility among the six-helix bundle of the DED. This crucial conformational change is modulated by the D/E-RxDL charge-triad motif, one of the prominent structural features of DEDs, together with a number of other electrostatic and hydrogen bonding interactions on the protein surface. Charge- triad motif promotes the optimal orientation of key residues and expands the binding interface to accommodate protein- protein interactions.
    [Show full text]
  • The Role of Phospholipase D (Pld) and Grb2 in Chemotaxis
    THE ROLE OF PHOSPHOLIPASE D (PLD) AND GRB2 IN CHEMOTAXIS A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science By KATIE J. KNAPEK B.S., Indiana University of Pennyslvania, 2006 2008 Wright State University WRIGHT STATE UNIVERSITY SCHOOL OF GRADUATE STUDIES December 19, 2008 I HEREBY RECOMMEND THAT THE THESIS PREPARED UNDER MY SUPERVISON BY Katie J. Knapek ENTITLED The Role of Phospholipase D (PLD) and Grb2 in Chemotaxis BE ACCEPTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF Master of Science. _____________________________ Julian Gomez-Cambronero, Ph. D. Thesis Director _____________________________ Barbara Hull, Ph. D. Program Director Committee on Final Examination _______________________ Julian Gomez-Cambronero, Ph. D. _______________________ Nancy Bigley, Ph. D. _______________________ Mill Miller, Ph. D. ________________________ Joseph F. Thomas Jr., Ph. D. Dean, School of Graduate Studies ABSTRACT Knapek, Katie J. M.S., Department of Biological Sciences, Microbiology and Immunology Program, Wright State University, 2008. The Role of Phospholipase D (PLD) and Grb2 in Chemotaxis. Phospholipase D (PLD) is an enzyme that hydrolyzes phosphatidylcholine yielding choline and phosphatidic acid. PLD is activated by mitogens (lead to cell division) and motogens (leading to cell migration). PLD is known to contribute to cellular proliferation and deregulated expression of PLD has been implicated in several human cancers. PLD has been found to play a role in leukocyte chemotaxis and adhesion as studied through the formation of chemokine gradients. We have established a model of cell migration comprising three cell lines: macrophages RAW 264.7 and LR-5 (for innate defense), and fibroblast COS-7 cells (for wound healing).
    [Show full text]
  • Author's Personal Copy
    Author's personal copy Provided for non-commercial research and educational use only. Not for reproduction, distribution or commercial use. This article was originally published in the book Encyclopedia of Immunobiology, published by Elsevier, and the attached copy is provided by Elsevier for the author's benefit and for the benefit of the author's institution, for non-commercial research and educational use including without limitation use in instruction at your institution, sending it to specific colleagues who you know, and providing a copy to your institution's administrator. All other uses, reproduction and distribution, including without limitation commercial reprints, selling or licensing copies or access, or posting on open internet sites, your personal or institution’s website or repository, are prohibited. For exceptions, permission may be sought for such use through Elsevier’s permissions site at: http://www.elsevier.com/locate/permissionusematerial From Myers, D.R., Roose, J.P., 2016. Kinase and Phosphatase Effector Pathways in T Cells. In: Ratcliffe, M.J.H. (Editor in Chief), Encyclopedia of Immunobiology, Vol. 3, pp. 25–37. Oxford: Academic Press. Copyright © 2016 Elsevier Ltd. unless otherwise stated. All rights reserved. Academic Press Author's personal copy Kinase and Phosphatase Effector Pathways in T Cells Darienne R Myers and Jeroen P Roose, University of California, San Francisco (UCSF), San Francisco, CA, USA Ó 2016 Elsevier Ltd. All rights reserved. Abstract Multiple interconnected effector pathways mediate the activation of T cells following recognition of cognate antigen. These kinase and phosphatase pathways link proximal T cell receptor (TCR) signaling to changes in gene expression and cell physiology.
    [Show full text]
  • Enhancement in Phospholipase D Activity As a New Proposed Molecular Mechanism of Haloperidol-Induced Neurotoxicity
    International Journal of Molecular Sciences Communication Enhancement in Phospholipase D Activity as a New Proposed Molecular Mechanism of Haloperidol-Induced Neurotoxicity Marek Krzystanek 1,*, Ewa Krzystanek 2 , Katarzyna Skałacka 3 and Artur Pałasz 4 1 Department and Clinic of Psychiatric Rehabilitation, Department of Psychiatry and Psychotherapy, Faculty of Medical Sciences, Medical School of Silesia in Katowice, Ziołowa 45/47, 40-635 Katowice, Poland 2 Department of Neurology, Faculty of Medical Sciences, Medical School of Silesia in Katowice, Medyków 14, 40-772 Katowice, Poland; [email protected] 3 Institute of Psychology, University of Opole, Kopernika 11A Street, 45-040 Opole, Poland; [email protected] 4 Department of Histology, Faculty of Medical Sciences, Medical School of Silesia in Katowice, Medyków 18, 40-752 Katowice, Poland; [email protected] * Correspondence: [email protected] or [email protected] Received: 19 November 2020; Accepted: 1 December 2020; Published: 4 December 2020 Abstract: Membrane phospholipase D (PLD) is associated with numerous neuronal functions, such as axonal growth, synaptogenesis, formation of secretory vesicles, neurodegeneration, and apoptosis. PLD acts mainly on phosphatidylcholine, from which phosphatidic acid (PA) and choline are formed. In turn, PA is a key element of the PLD-dependent secondary messenger system. Changes in PLD activity are associated with the mechanism of action of olanzapine, an atypical antipsychotic. The aim of the present study was to assess the effect of short-term administration of the first-generation antipsychotic drugs haloperidol, chlorpromazine, and fluphenazine on membrane PLD activity in the rat brain. Animals were sacrificed for a time equal to the half-life of the antipsychotic drug in the brain, then the membranes in which PLD activity was determined were isolated from the tissue.
    [Show full text]