DOCSLIB.ORG

Supporting Information

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Supporting Information Supporting Information Ortiz et al. 10.1073/pnas.1401952111 SI Materials and Methods monoclonal, 5 μg/mL) was added followed by Tyramide-Alexa Magnetic Resonance Imaging. Brains of injected mice were scanned Fluor 488 (Invitrogen, cat. no. T20922). at 8-, 11-, 16-, and 20-wk postinjection with a 200 MHz Bruker 4.7T Biospec MRI scanner equipped with a 560 mT/m ID 12-cm Immunostaining Image Analysis. Whole slides were scanned with gradient (Bruker Biospin MRI; Resonance Research). For mouse Pannoramic Flash Scanner (3DHistech). Image analysis of tumor brain imaging, brain coronal T2-weighted images using fast spin- areas was performed with Metamorph software (Molecular echo RARE sequence (Rapid Acquisition with Relaxation En- Devices). For analysis of immunohistochemistry images, color hancement) was acquired with TR 1.5 s, TE 50 ms, RARE factor thresholds were set for brown positive staining and for total area + of 8, slice thickness of 0.7 mm, field of view 30 × 20mm, in-plane (brown staining blue nuclei). Percent of brown staining to total resolution of 117 × 125 μM, and 24 averages. Tumor volume was area was calculated. For analysis of dual immunofluorescence calculated by contouring, measuring the tumor areas, and cal- images, a grayscale threshold and standard area was set for green culating the sum of the areas multiplied by the distance between (Iba1), red (p-Stat3), and blue (DAPI). For each sample, the the centers of two adjacent slices. number of DAPI-positive nuclei within each stained area, and the number of DAPI within colocalized areas was calculated. Histology and Immunostaining. Brains from mice were collected and fixed with 10% (vol/vol) neutral buffered formalin (Sigma). Human Tumor Collection, Tissue Lysates, and Immunoblotting. Fresh Tissues were embedded in paraffin and 5-μm sections were used human glioblastoma multiforme (GBM) tissue samples were for analysis. Sections were stained with H&E or used for im- obtained from patients who consented under an Institutional munohistochemical analysis. Review Board-approved protocol. Tumor lysates were lysed in Immunohistochemistry was performed at the Molecular Cytol- CelLytic MT Mammalian Tissue Lysis/Extraction Reagent (Sigma) ogy Core Facility of Memorial Sloan-Kettering Cancer Center supplemented with Complete Mini EDTA-free (Roche) and using Discovery XT processor (Ventana Medical Systems). Tissue PhosSTOP (Roche) protease inhibitor mixes. Protein lysates sections were stained with the following antibodies: p-Stat3 Tyr-705 were run in SDS/PAGE gels and transferred to nitrocellulose (Cell Signaling, cat. no. 9145; rabbit monoclonal, 0.5 μg/mL), Stat3 for immunoblotting. The following antibodies were used: p-STAT3 (Cell Signaling, cat. no. 9132; rabbit polyclonal, 0.16 μg/mL), Iba1 Tyr-705 (Cell Signaling, cat. no. 9145; rabbit monoclonal, 1:2,000) (Wako Chemicals, cat. no. 019–19741; rabbit polyclonal, 0.5 μg/mL), and STAT3 (Cell Signaling, cat. no. 9132; rabbit polyclonal, 1:1,000). Gfap (Dako, cat. no. Z033429-2; rabbit polyclonal, 1 μg/mL), Olig2 Quantification of Western blot by densitometry analysis was per- formed using ImageJ software. (Millipore, cat. no. AB9610; rabbit polyclonal, 2 μg/mL), and Nestin (BD Pharmigen, cat. no. 556309; mouse monoclonal, μ Flow Sorting of RCAS PDGF-GFP Tumors. Tumors were dissected 5 g/mL). For rabbit antibodies, tissue sections were blocked in from mice injected with replication-competent avian sarcoma- 10% (vol/vol) goat serum with 2% (wt/vol) BSA in PBS. For the leukosis virus long terminal repeat with splice acceptor retrovirus mouse antibody, tissues were blocked with Biotinylated Mouse on (RCAS) PDGF-GFP and enzymatically and mechanically dis- Mouse (M.O.M.) anti-Mouse Ig Reagent (Vector Labs, cat. no. sociated into single-cell suspensions by treatment with papain and MKB-2225). Primary antibody incubation was done for 5 h, ovomucoid as previously described (1). Single-cell suspensions + − followed by a 60-min incubation with biotinylated goat anti- were made in PBS with 10% (vol/vol) FBS, and GFP DAPI rabbit IgG (Vector Labs, cat. no. PK6101; 1:200 dilution) or cells were sorted on a MoFlo Cell Sorter (Dako Cytomation). biotinylated anti-mouse IgG (Vector Labs, cat. no. BMK-2202) ’ according to the manufacturer s instructions. Detection was per- Human Genetic Analysis and Microarray Analysis of Mouse Tumor formed with Blocker D, Streptavidin-HRP and DAB kit (Ventana Cells. The Cancer Genome Atlas data used was previously de- ’ Medical Systems) according to the manufacturer s instructions. scribed (2, 3). The GBM oncoprint was generated using the cBio Slides were counterstained with hematoxylin and cover-slipped Portal as previously described (4) + − with Permount (Fisher Scientific). Sorted GFP DAPI tumor cells from mice injected with CD34 staining was performed on the Leica Bond RX (Leica RCAS-PDGF-GFP were stored at −80 °C in TRIzol LS Reagent Biosystems) using the Bond Polymer Refine Detection Kit (Leica (Ambion). Samples were processed by the Memorial Sloan- Biosystems, DS980). Anti-CD34 antibody (Abcam, cat. no. Kettering Cancer Center Genomics Core. Briefly, RNA was ex- ab8158; rat monoclonal, 16 μg/mL) was added for 30 min fol- tracted and quality checked using a bioanalyzer. RNA was analyzed lowed by a 30-min incubation of biotinylated anti-rat (Vector using the Affymetrix MOE 430A 2.0 chip following the manu- Labs, cat. no. BA-4001; 1:100 dilution). TUNEL staining was facturer’s instructions. Differentially expressed genes were deter- performed with the following reaction mixture: 0.1 M sodium mined using ANOVA. Principal component analysis and hierar- cacodylate pH 7, 0.1 mM DTT, 0.05 mg/mL bovine serum al- chical analysis were performed using the Partek Software Suite. bumin, 2 U/μl terminal transferase, 0.2 nm biotin-16-dUTP, and Genes were considered to be differentially expressed if their fold- 2.5 mM cobalt chloride for 1 h at 37°. The reaction was termi- change > 1.8 and P value < 0.05. Enriched pathways were iden- nated with 300 mM sodium chloride and 30 mM sodium citrate tified using Ingenuity Pathway Analysis (Ingenuity Systems, at room temperature for 15 min, incubated in avidin-biotin for Qiagen). Pathways with Benjami–Hochberg multiple testing cor- 30 min, and developed with 3,3′-diaminobenzidine for 3 min. rection P value < 0.05 and Bias-corrected z-score > 2werecon- Dual immunofluorescence of p-Stat3 and Iba1 or Gfap were sidered to be significant. performed using Discovery XT processor (Ventana Medical Systems). Staining with p-Stat3 Tyr-705 (Cell Signaling, cat. no. Side Population Assay. Tumors were dissected from mice injected 9145; rabbit monoclonal, 0.5 μg/mL) followed by Tyramide Alexa with RCAS PDGF-GFP and dissociated into single-cell sus- Fluor 568 (Invitrogen, cat. no T20914) was performed first. Next, pensions, as previously described (1). Single-cell suspensions Iba1 (Wako Chemicals, cat. no. 019–19741; rabbit polyclonal, were made in unsupplemented basal neural stem cell media 0.5 μg/mL) or Gfap (BD Pharmigen, cat. no. 561483; mouse (Lonza) and counted using a hemocytometer. Side population Ortiz et al. www.pnas.org/cgi/content/short/1401952111 1of10 analysis was performed by Hoechst 33342 exclusion as previously at room temperature. Cells were washed two times with HBSS:2% described (5). Cells were incubated with or without verapamil (vol/vol) FBS and suspended in PBS:10% (vol/vol) FBS for analysis (Sigma) and fumitremorgin c (Sigma) ABC inhibitors to set side with a FACSCalibur (BD Biosciences). Analysis was performed population gates (5). Flow cytometry was performed with a MoFlo using FloJo software (Treestar Inc.). Cell Sorter (Dako Cytomation) and analysis was performed using Flojo software (Treestar Inc.). Flow Cytometry for Nestin. Single-cell suspensions of neonatal mouse days 1–3 brains were made using papain digestion as done K i67 and GFP Flow Cytometry. Tumors were dissected from mice previously. Staining was performed as described previously using injected with RCAS PDGF-GFP and dissociated into single-cell Nestin antibody (BD Pharmigen, mouse monoclonal, cat. no. suspensions as previously described (1). Single-cell suspensions μ were made in PBS and counted using a hemocytometer. The 556309; 1.7 g/mL) and goat anti-mouse Alexa 568 (Life Tech- μ following protocol was performed to minimize quenching of native nologies, cat. no. A21124; 33 g/mL) (1). Flow cytometry was GFP in the tumor cells. One million cells were fixed with 1% performed on the FACSCalibur (BD Biosciences), and analysis paraformaldehyde in a buffer made of HBSS and 2% (vol/vol) was performed using FloJo software (Treestar Inc.). fetal bovine calf serum overnight at 4 °C. Cells were permeabilized using 0.5% TritonX-100 in HBSS and 2% (vol/vol) FBS for 10 min Statistical Analysis. Unless noted, Student t test was performed for at room temperature. Ki67 antibody conjugated to Alexa 647 (BD all statistical analysis. Log-rank statistical analysis was performed Pharmigen, cat no. 561126; mouse monoclonal, 1/60 dilution) for Kaplan–Meier curves and Fisher’s exact test was performed was made in HBSS:2% (vol/vol) FBS and applied to cells for 1 h for the tumor grade analysis. 1. Ciznadija D, Liu Y, Pyonteck SM, Holland EC, Koff A (2011) Cyclin D1 and cdk4 mediate 4. Cerami E, et al. (2012) The cBio cancer genomics portal: an open platform for exploring development of neurologically destructive oligodendroglioma. Cancer Res 71(19): multidimensional cancer genomics data. Cancer Discov 2(5):401–404. 6174–6183. 5. Bleau AM, et al. (2009) PTEN/PI3K/Akt pathway regulates the side population phenotype 2. Cancer Genome Atlas Research Network (2008) Comprehensive genomic characteriza- and ABCG2 activity in glioma tumor stem-like cells. Cell Stem Cell 4(3):226–235. tion defines human glioblastoma genes and core pathways. Nature 455(7216):1061–1068. 3. Verhaak RG, et al.; Cancer Genome Atlas Research Network (2010) Integrated genomic analysis identifies clinically relevant subtypes of glioblastoma characterized by abnormalities in PDGFRA, IDH1, EGFR, and NF1.
Load more

Recommended publications
  • Multigene Panels in Prostate Cancer Risk Assessment
    Evidence Report/Technology Assessment Number 209 Multigene Panels in Prostate Cancer Risk Assessment Evidence-Based Practice Evidence Report/Technology Assessment Number 209 Multigene Panels in Prostate Cancer Risk Assessment Prepared for: Agency for Healthcare Research and Quality U.S. Department of Health and Human Services 540 Gaither Road Rockville, MD 20850 www.ahrq.gov Contract No. 290-2007-10060-1 Prepared by: McMaster University Evidence-based Practice Center Hamilton, ON, Canada Investigators: Julian Little, Ph.D. Brenda Wilson, M.B.Ch.B., M.Sc., M.R.C.P. (UK), FFPH Ron Carter, Ph.D. Kate Walker, M.Sc.PT. Pasqualina Santaguida, Ph.D. Eva Tomiak, M.D. Joseph Beyene, Ph.D. Parminder Raina, Ph.D. AHRQ Publication No. 12-E020-EF July 2012 This report is based on research conducted by the McMaster University Evidence-based Practice Center (EPC) under contract to the Agency for Healthcare Research and Quality (AHRQ), Rockville, MD (Contract No. 290-2007-10060-1). The findings and conclusions in this document are those of the authors, who are responsible for its contents; the findings and conclusions do not necessarily represent the views of AHRQ. Therefore, no statement in this report should be construed as an official position of AHRQ or of the U.S. Department of Health and Human Services. The information in this report is intended to help health care decisionmakers—patients and clinicians, health system leaders, and policymakers, among others—make well-informed decisions and thereby improve the quality of health care services. This report is not intended to be a substitute for the application of clinical judgment.
    [Show full text]
  • Supplemental Figure 1. Vimentin
    Double mutant specific genes Transcript gene_assignment Gene Symbol RefSeq FDR Fold- FDR Fold- FDR Fold- ID (single vs. Change (double Change (double Change wt) (single vs. wt) (double vs. single) (double vs. wt) vs. wt) vs. single) 10485013 BC085239 // 1110051M20Rik // RIKEN cDNA 1110051M20 gene // 2 E1 // 228356 /// NM 1110051M20Ri BC085239 0.164013 -1.38517 0.0345128 -2.24228 0.154535 -1.61877 k 10358717 NM_197990 // 1700025G04Rik // RIKEN cDNA 1700025G04 gene // 1 G2 // 69399 /// BC 1700025G04Rik NM_197990 0.142593 -1.37878 0.0212926 -3.13385 0.093068 -2.27291 10358713 NM_197990 // 1700025G04Rik // RIKEN cDNA 1700025G04 gene // 1 G2 // 69399 1700025G04Rik NM_197990 0.0655213 -1.71563 0.0222468 -2.32498 0.166843 -1.35517 10481312 NM_027283 // 1700026L06Rik // RIKEN cDNA 1700026L06 gene // 2 A3 // 69987 /// EN 1700026L06Rik NM_027283 0.0503754 -1.46385 0.0140999 -2.19537 0.0825609 -1.49972 10351465 BC150846 // 1700084C01Rik // RIKEN cDNA 1700084C01 gene // 1 H3 // 78465 /// NM_ 1700084C01Rik BC150846 0.107391 -1.5916 0.0385418 -2.05801 0.295457 -1.29305 10569654 AK007416 // 1810010D01Rik // RIKEN cDNA 1810010D01 gene // 7 F5 // 381935 /// XR 1810010D01Rik AK007416 0.145576 1.69432 0.0476957 2.51662 0.288571 1.48533 10508883 NM_001083916 // 1810019J16Rik // RIKEN cDNA 1810019J16 gene // 4 D2.3 // 69073 / 1810019J16Rik NM_001083916 0.0533206 1.57139 0.0145433 2.56417 0.0836674 1.63179 10585282 ENSMUST00000050829 // 2010007H06Rik // RIKEN cDNA 2010007H06 gene // --- // 6984 2010007H06Rik ENSMUST00000050829 0.129914 -1.71998 0.0434862 -2.51672
    [Show full text]
  • Molecular Profile of Tumor-Specific CD8+ T Cell Hypofunction in a Transplantable Murine Cancer Model
    Downloaded from http://www.jimmunol.org/ by guest on September 25, 2021 T + is online at: average * The Journal of Immunology , 34 of which you can access for free at: 2016; 197:1477-1488; Prepublished online 1 July from submission to initial decision 4 weeks from acceptance to publication 2016; doi: 10.4049/jimmunol.1600589 http://www.jimmunol.org/content/197/4/1477 Molecular Profile of Tumor-Specific CD8 Cell Hypofunction in a Transplantable Murine Cancer Model Katherine A. Waugh, Sonia M. Leach, Brandon L. Moore, Tullia C. Bruno, Jonathan D. Buhrman and Jill E. Slansky J Immunol cites 95 articles Submit online. Every submission reviewed by practicing scientists ? is published twice each month by Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts http://jimmunol.org/subscription Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html http://www.jimmunol.org/content/suppl/2016/07/01/jimmunol.160058 9.DCSupplemental This article http://www.jimmunol.org/content/197/4/1477.full#ref-list-1 Information about subscribing to The JI No Triage! Fast Publication! Rapid Reviews! 30 days* Why • • • Material References Permissions Email Alerts Subscription Supplementary The Journal of Immunology The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2016 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. This information is current as of September 25, 2021. The Journal of Immunology Molecular Profile of Tumor-Specific CD8+ T Cell Hypofunction in a Transplantable Murine Cancer Model Katherine A.
    [Show full text]
  • Supplementary Table S1. Upregulated Genes Differentially
    Supplementary Table S1. Upregulated genes differentially expressed in athletes (p < 0.05 and 1.3-fold change) Gene Symbol p Value Fold Change 221051_s_at NMRK2 0.01 2.38 236518_at CCDC183 0.00 2.05 218804_at ANO1 0.00 2.05 234675_x_at 0.01 2.02 207076_s_at ASS1 0.00 1.85 209135_at ASPH 0.02 1.81 228434_at BTNL9 0.03 1.81 229985_at BTNL9 0.01 1.79 215795_at MYH7B 0.01 1.78 217979_at TSPAN13 0.01 1.77 230992_at BTNL9 0.01 1.75 226884_at LRRN1 0.03 1.74 220039_s_at CDKAL1 0.01 1.73 236520_at 0.02 1.72 219895_at TMEM255A 0.04 1.72 201030_x_at LDHB 0.00 1.69 233824_at 0.00 1.69 232257_s_at 0.05 1.67 236359_at SCN4B 0.04 1.64 242868_at 0.00 1.63 1557286_at 0.01 1.63 202780_at OXCT1 0.01 1.63 1556542_a_at 0.04 1.63 209992_at PFKFB2 0.04 1.63 205247_at NOTCH4 0.01 1.62 1554182_at TRIM73///TRIM74 0.00 1.61 232892_at MIR1-1HG 0.02 1.61 204726_at CDH13 0.01 1.6 1561167_at 0.01 1.6 1565821_at 0.01 1.6 210169_at SEC14L5 0.01 1.6 236963_at 0.02 1.6 1552880_at SEC16B 0.02 1.6 235228_at CCDC85A 0.02 1.6 1568623_a_at SLC35E4 0.00 1.59 204844_at ENPEP 0.00 1.59 1552256_a_at SCARB1 0.02 1.59 1557283_a_at ZNF519 0.02 1.59 1557293_at LINC00969 0.03 1.59 231644_at 0.01 1.58 228115_at GAREM1 0.01 1.58 223687_s_at LY6K 0.02 1.58 231779_at IRAK2 0.03 1.58 243332_at LOC105379610 0.04 1.58 232118_at 0.01 1.57 203423_at RBP1 0.02 1.57 AMY1A///AMY1B///AMY1C///AMY2A///AMY2B// 208498_s_at 0.03 1.57 /AMYP1 237154_at LOC101930114 0.00 1.56 1559691_at 0.01 1.56 243481_at RHOJ 0.03 1.56 238834_at MYLK3 0.01 1.55 213438_at NFASC 0.02 1.55 242290_at TACC1 0.04 1.55 ANKRD20A1///ANKRD20A12P///ANKRD20A2///
    [Show full text]
  • RNA Epigenetics: Fine-Tuning Chromatin Plasticity and Transcriptional Regulation, and the Implications in Human Diseases
    G C A T T A C G G C A T genes Review RNA Epigenetics: Fine-Tuning Chromatin Plasticity and Transcriptional Regulation, and the Implications in Human Diseases Amber Willbanks, Shaun Wood and Jason X. Cheng * Department of Pathology, Hematopathology Section, University of Chicago, Chicago, IL 60637, USA; [email protected] (A.W.); [email protected] (S.W.) * Correspondence: [email protected] Abstract: Chromatin structure plays an essential role in eukaryotic gene expression and cell identity. Traditionally, DNA and histone modifications have been the focus of chromatin regulation; however, recent molecular and imaging studies have revealed an intimate connection between RNA epigenetics and chromatin structure. Accumulating evidence suggests that RNA serves as the interplay between chromatin and the transcription and splicing machineries within the cell. Additionally, epigenetic modifications of nascent RNAs fine-tune these interactions to regulate gene expression at the co- and post-transcriptional levels in normal cell development and human diseases. This review will provide an overview of recent advances in the emerging field of RNA epigenetics, specifically the role of RNA modifications and RNA modifying proteins in chromatin remodeling, transcription activation and RNA processing, as well as translational implications in human diseases. Keywords: 5’ cap (5’ cap); 7-methylguanosine (m7G); R-loops; N6-methyladenosine (m6A); RNA editing; A-to-I; C-to-U; 2’-O-methylation (Nm); 5-methylcytosine (m5C); NOL1/NOP2/sun domain Citation: Willbanks, A.; Wood, S.; (NSUN); MYC Cheng, J.X. RNA Epigenetics: Fine-Tuning Chromatin Plasticity and Transcriptional Regulation, and the Implications in Human Diseases. Genes 2021, 12, 627.
    [Show full text]
  • A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
    Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated.
    [Show full text]
  • Exosome-Mediated MIR211 Modulates Tumor Microenvironment Via the DUSP6-ERK5 Axis and Contributes to BRAFV600E Inhibitor Resistan
    bioRxiv preprint doi: https://doi.org/10.1101/548818; this version posted February 13, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Exosome-mediated MIR211 modulates tumor microenvironment via the DUSP6-ERK5 axis and contributes to BRAFV600E inhibitor resistance in melanoma. Bongyong Lee1,3, Anupama Sahoo3, Junko Sawada1,3, John Marchica1,3, Sanjay Sahoo3, Fabiana I. A. L. Layng4, Darren Finlay4, Joseph Mazar5, Piyush Joshi1, Masanobu Komatsu1,3, Kristiina Vuori4, Garth Powis4, Petrus R. de Jong4, Animesh Ray6,7, and Ranjan J. Perera 1,2,3* 1 Department of Cancer and Blood Disorders Institute, Johns Hopkins All Children’s Hospital, St. Petersburg, FL 33701 USA 2 Department of Oncology, Sydney Kimmel Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287 USA 3 Sanford Burnham Prebys Medical Discovery Institute, Orlando, FL 32827 USA 4 Sanford Burnham Prebys Medical Discovery Institute, NCI-Designated Cancer Center, La Jolla, CA 92037, USA 5 Nemours Children Hospital, Orlando, FL 32827 USA 6 Keck Graduate Institute, Claremont CA 91711 USA, 7 Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125 USA. *Correspondence: Ranjan Perera Tel: 1-727-767-3491 Email: [email protected] 1 bioRxiv preprint doi: https://doi.org/10.1101/548818; this version posted February 13, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. ABSTRACT The microRNA MIR211 is an important regulator of melanoma tumor cell behavior.
    [Show full text]
  • 9. Atypical Dusps: 19 Phosphatases in Search of a Role
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Digital.CSIC Transworld Research Network 37/661 (2), Fort P.O. Trivandrum-695 023 Kerala, India Emerging Signaling Pathways in Tumor Biology, 2010: 185-208 ISBN: 978-81-7895-477-6 Editor: Pedro A. Lazo 9. Atypical DUSPs: 19 phosphatases in search of a role Yolanda Bayón and Andrés Alonso Instituto de Biología y Genética Molecular, CSIC-Universidad de Valladolid c/ Sanz y Forés s/n, 47003 Valladolid, Spain Abstract. Atypical Dual Specificity Phosphatases (A-DUSPs) are a group of 19 phosphatases poorly characterized. They are included among the Class I Cys-based PTPs and contain the active site motif HCXXGXXR conserved in the Class I PTPs. These enzymes present a phosphatase domain similar to MKPs, but lack any substrate targeting domain similar to the CH2 present in this group. Although most of these phosphatases have no more than 250 amino acids, their size ranges from the 150 residues of the smallest A-DUSP, VHZ/DUSP23, to the 1158 residues of the putative PTP DUSP27. The substrates of this family include MAPK, but, in general terms, it does not look that MAPK are the general substrates for the whole group. In fact, other substrates have been described for some of these phosphatases, like the 5’CAP structure of mRNA, glycogen, or STATs and still the substrates of many A-DUSPs have not been identified. In addition to the PTP domain, most of these enzymes present no additional recognizable domains in their sequence, with the exception of CBM-20 in laforin, GTase in HCE1 and a Zn binding domain in DUSP12.
    [Show full text]
  • Dual-Specificity Phosphatase 3 Deletion Protects Female, but Not
    Published August 28, 2017, doi:10.4049/jimmunol.1602092 The Journal of Immunology Dual-Specificity Phosphatase 3 Deletion Protects Female, but Not Male, Mice from Endotoxemia-Induced and Polymicrobial-Induced Septic Shock Maud M. Vandereyken,*,1 Pratibha Singh,*,1 Caroline P. Wathieu,* Sophie Jacques,* Tinatin Zurashvilli,* Lien Dejager,†,‡ Mathieu Amand,* Lucia Musumeci,* Maneesh Singh,* Michel P. Moutschen,* Claude R. F. Libert,†,‡ and Souad Rahmouni* Dual-specificity phosphatase 3 (DUSP3) is a small phosphatase with poorly known physiological functions and for which only a few substrates are known. Using knockout mice, we recently reported that DUSP3 deficiency confers resistance to endotoxin- and polymicrobial-induced septic shock. We showed that this protection was macrophage dependent. In this study, we further investigated the role of DUSP3 in sepsis tolerance and showed that the resistance is sex dependent. Using adoptive-transfer experiments and ovariectomized mice, we highlighted the role of female sex hormones in the phenotype. Indeed, in ovariec- tomized females and in male mice, the dominance of M2-like macrophages observed in DUSP32/2 female mice was reduced, suggesting a role for this cell subset in sepsis tolerance. At the molecular level, DUSP3 deletion was associated with estrogen- dependent decreased phosphorylation of ERK1/2 and Akt in peritoneal macrophages stimulated ex vivo by LPS. Our results demonstrate that estrogens may modulate M2-like responses during endotoxemia in a DUSP3-dependent manner. The Journal of Immunology, 2017, 199: 000–000. epsis and septic shock are complex clinical syndromes that ally, death (4). Sepsis occurrence and outcome depend on arise when the local body response to pathogens becomes pathogen characteristics, as well as on risk factors, such as age S systemic and injures its own tissues and organs (1).
    [Show full text]
  • CDH12 Cadherin 12, Type 2 N-Cadherin 2 RPL5 Ribosomal
    5 6 6 5 . 4 2 1 1 1 2 4 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 2 2 A A A A A A A A A A A A A A A A A A A A C C C C C C C C C C C C C C C C C C C C R R R R R R R R R R R R R R R R R R R R B , B B B B B B B B B B B B B B B B B B B , 9 , , , , 4 , , 3 0 , , , , , , , , 6 2 , , 5 , 0 8 6 4 , 7 5 7 0 2 8 9 1 3 3 3 1 1 7 5 0 4 1 4 0 7 1 0 2 0 6 7 8 0 2 5 7 8 0 3 8 5 4 9 0 1 0 8 8 3 5 6 7 4 7 9 5 2 1 1 8 2 2 1 7 9 6 2 1 7 1 1 0 4 5 3 5 8 9 1 0 0 4 2 5 0 8 1 4 1 6 9 0 0 6 3 6 9 1 0 9 0 3 8 1 3 5 6 3 6 0 4 2 6 1 0 1 2 1 9 9 7 9 5 7 1 5 8 9 8 8 2 1 9 9 1 1 1 9 6 9 8 9 7 8 4 5 8 8 6 4 8 1 1 2 8 6 2 7 9 8 3 5 4 3 2 1 7 9 5 3 1 3 2 1 2 9 5 1 1 1 1 1 1 5 9 5 3 2 6 3 4 1 3 1 1 4 1 4 1 7 1 3 4 3 2 7 6 4 2 7 2 1 2 1 5 1 6 3 5 6 1 3 6 4 7 1 6 5 1 1 4 1 6 1 7 6 4 7 e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l l p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m m
    [Show full text]
  • DUSP10/MKP5 Antibody A
    Revision 1 C 0 2 - t DUSP10/MKP5 Antibody a e r o t S Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) 3 8 Web: [email protected] 4 www.cellsignal.com 3 # 3 Trask Lane Danvers Massachusetts 01923 USA For Research Use Only. Not For Use In Diagnostic Procedures. Applications: Reactivity: Sensitivity: MW (kDa): Source: UniProt ID: Entrez-Gene Id: WB H M R Endogenous 54 Rabbit Q9Y6W6 11221 Product Usage Information 3. Salojin, K. and Oravecz, T. (2007) J Leukoc Biol 81, 860-9. 4. Tanoue, T. et al. (2002) J Biol Chem 277, 22942-9. Application Dilution 5. Dickinson, R.J. and Keyse, S.M. (2006) J Cell Sci 119, 4607-15. 6. Wu, G.S. (2007) Cancer Metastasis Rev 26, 579-85. Western Blotting 1:1000 7. Teng, C.H. et al. (2007) J Biol Chem 282, 28395-407. 8. Zhang, Y. et al. (2004) Nature 430, 793-7. Storage Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody. Specificity / Sensitivity DUSP10/MKP5 Antibody detects endogenous levels of total DUSP10 protein. Species Reactivity: Human, Mouse, Rat Source / Purification Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to human DUSP10. Antibodies are purified by protein A and peptide affinity chromatography. Background MAP kinases are inactivated by dual-specificity protein phosphatases (DUSPs) that differ in their substrate specificity, tissue distribution, inducibility by extracellular stimuli, and cellular localization. DUSPs, also known as MAPK phosphatases (MKP), specifically dephosphorylate both threonine and tyrosine residues in MAPK P-loops and have been shown to play important roles in regulating the function of the MAPK family (1,2).
    [Show full text]
  • The Regulatory Roles of Phosphatases in Cancer
    Oncogene (2014) 33, 939–953 & 2014 Macmillan Publishers Limited All rights reserved 0950-9232/14 www.nature.com/onc REVIEW The regulatory roles of phosphatases in cancer J Stebbing1, LC Lit1, H Zhang, RS Darrington, O Melaiu, B Rudraraju and G Giamas The relevance of potentially reversible post-translational modifications required for controlling cellular processes in cancer is one of the most thriving arenas of cellular and molecular biology. Any alteration in the balanced equilibrium between kinases and phosphatases may result in development and progression of various diseases, including different types of cancer, though phosphatases are relatively under-studied. Loss of phosphatases such as PTEN (phosphatase and tensin homologue deleted on chromosome 10), a known tumour suppressor, across tumour types lends credence to the development of phosphatidylinositol 3--kinase inhibitors alongside the use of phosphatase expression as a biomarker, though phase 3 trial data are lacking. In this review, we give an updated report on phosphatase dysregulation linked to organ-specific malignancies. Oncogene (2014) 33, 939–953; doi:10.1038/onc.2013.80; published online 18 March 2013 Keywords: cancer; phosphatases; solid tumours GASTROINTESTINAL MALIGNANCIES abs in sera were significantly associated with poor survival in Oesophageal cancer advanced ESCC, suggesting that they may have a clinical utility in Loss of PTEN (phosphatase and tensin homologue deleted on ESCC screening and diagnosis.5 chromosome 10) expression in oesophageal cancer is frequent, Cao et al.6 investigated the role of protein tyrosine phosphatase, among other gene alterations characterizing this disease. Zhou non-receptor type 12 (PTPN12) in ESCC and showed that PTPN12 et al.1 found that overexpression of PTEN suppresses growth and protein expression is higher in normal para-cancerous tissues than induces apoptosis in oesophageal cancer cell lines, through in 20 ESCC tissues.
    [Show full text]