DOCSLIB.ORG

For Antigen-Specific Immune Responses Dispensable Intracellular Igg and Igm but Is Fc Receptor-Like a Associates With

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
For Antigen-Specific Immune Responses Dispensable Intracellular Igg and Igm but Is Fc Receptor-Like a Associates With The Journal of Immunology Fc Receptor-Like A Associates with Intracellular IgG and IgM but Is Dispensable for Antigen-Specific Immune Responses Timothy J. Wilson, Susan Gilfillan, and Marco Colonna FcR-like (FcRL) proteins comprise a family of lymphocyte receptors with homology to FcgRI. Among these receptors, FcRLA is uniquely interesting due to its intracellular localization, unusual structural features, and high expression within human germinal center and marginal zone B cells. Our analysis of human cell lines has confirmed that this receptor is not secreted but is maintained as an intracellular protein in B cells where it interacts with Igs, consistent with a possible role in Ab assembly. By generating FcRLA-specific antisera as well as knockout mice, we were able to unequivocally demonstrate that FcRLA protein is expressed exclusively in all mouse B cells. We also found that FcRLA is not required for the generation of Ag-specific humoral immune responses to T-dependent or T-independent Ags. However, given its highly conserved structure and universal expression within B cells, it is probable that FcRLA functions similarly in humans and mice. Cumulatively, our data suggest that FcRLA plays a role in Ig assembly that can be compensated for by other proteins. The Journal of Immunology, 2010, 185: 2960–2967. cRs bind Igs with a range of specificities and affinities FcRLA and FcRLB, are highly conserved and share unique fea- and can either activate or inhibit cellular responses. They tures; 1) they have no transmembrane domain and hence are F assist essential effector functions of Abs by mediating Ab- expressed as intracellular receptors; 2) both have a C terminus dependent cell-mediated cytotoxicity, cellular activation, recogni- containing a proline-rich stalk region followed by a leucine-rich tion and capture of opsonized pathogens, uptake of Ag for presen- coiled-coil motif. Their differences lie in their N-terminal domain tation to T cells, and regulation of B cell responses (1). Additionally, structures and expression patterns. Whereas FcRLB has Ig-like some FcRs mediate specialized functions for Ig transport and recy- domains with homology to all three domains of FcgRI, FcRLA cling, including the polymeric Ig receptor and the neonatal FcR (2). contains Ig-like domains homologous to only the second and third FcR-like (FcRL) proteins were identified as a family of cell sur- domains of FcgR1, preceded by an N-terminal domain rich in face receptors with homology to FcgRI in one or more domains acidic residues as well as potentially unpaired cysteines. In addi- and are expressed differentially on B cells at various stages of tion, FcRLB seems to be expressed in only a tiny fraction of non- differentiation (3–7). More recently, an additional member of dividing B cells, whereas FcRLA is expressed at low levels in the family (FcRL6) was found primarily on the surfaces of NK mantle zone B cells and at higher levels within germinal centers cells and cytotoxic T cells (8, 9). FcRL1–6 are type I transmem- and splenic marginal zone B cells in humans (13, 22–27). brane proteins of the Ig superfamily, and all have one or more Currently, the ligands for the ectodomains of the human FcRL canonical ITAMs or ITIMs in their cytoplasmic tails. The ability proteins are unknown, although anecdotal reports in the literature to recruit tyrosine kinases or protein tyrosine phosphatases has have implicated Ig binding by FcRL4 and FcRL5 (5, 28). In this been reported for all of these receptors (8–15), with bona fide article, we demonstrate the clear association of FcRLA with in- activating potential found for FcRL1 (12) and potent inhibitory tracellular Ig, illustrate the expression of FcRLA within mouse potential found for FcRL4 and FcRL5 (10, 11). This potential to B cells, and examine the consequence of FcRLA deficiency on regulate B cell responses appears to be conserved in mouse FcR humoral immune responses in vivo. homolog 3/FcRL5 (16). The evolutionary divergence of most FcRL family members from Materials and Methods humans to rodents has been extensive, with significant structural 2 2 Generation of Fcrla / mice changes or gene deletions affecting all of the cell surface FcRL 2/2 proteins (9, 17–21). However, two members of the FcRL family, Fcrla mice were generated in E14.1 (129P2/OlaHsd) embryonic stem cells by targeted disruption and replacement of the third and fourth exons of the Fcrla gene with an MC1-neor expression cassette. Chimeras Department of Pathology and Immunology, Washington University School of Med- resulting from the injection of targeted embryonic stem cells into icine, St. Louis, MO 63110 C57BL/6 blastocysts were bred to C57BL/6 transgenic mice expressing Received for publication April 29, 2010. Accepted for publication June 24, 2010. Cre recombinase under the control of the CMV promoter to excise the MC1-neor cassette. Heterozygous Fcrla2/+ mice were intercrossed, and the This work was supported in part by the Center for HIV/AIDS Vaccine Immunology. resulting homozygous Fcrla+/+ and Fcrla2/2 progeny were used for analysis T.J.W. was sponsored by a predoctoral training grant in Tumor Immunology from the Cancer Research Institute. of humoral immune responses in vivo. All of the animal procedures in this study have been reviewed and approved by the Washington University Address correspondence and reprint requests to Prof. Marco Colonna, Department of Animal Care and Use Committee. Pathology and Immunology, Washington University School of Medicine, 660 South Euclid Avenue, Campus Box 8118, St. Louis, MO 63110. E-mail address: mcolonna@ Abs used in ELISA, flow cytometry, and immunohistochemistry pathology.wustl.edu Abbreviations used in this paper: FcRL, FcR-like; IP, immunoprecipitation; KLH, Rabbit antisera specific for human and mouse FcRLA were raised by Pacific keyhole limpet hemocyanin; NP, nitrophenyl; SRBC, sheep RBC. Immunology Company (San Diego, CA). Peptides SGHQKPGTTKATAE for human FcRLA and SVYLKPGTTKVADK for mouse FcRLA were Copyright Ó 2010 by The American Association of Immunologists, Inc. 0022-1767/10/$16.00 conjugated to keyhole limpet hemocyanin (KLH) and immunized according www.jimmunol.org/cgi/doi/10.4049/jimmunol.1001428 The Journal of Immunology 2961 to established protocols. Rabbit antisera (numbers 839 and 840 for human prior to immunoprecipitations (IPs). For IP from CESS cells, lysates were and 1391 and 1392 for mouse) were stored in 50% glycerol at 280˚C for incubated with protein G-, protein A-, protein L-, or streptavidin-coated aga- preservation and 220˚C for working stocks. Purified mAbs specific for hu- rose beads (Thermo Scientific Pierce, Rockford, IL). As a positive control for man FcRLA have been described previously (24). For immunohistological FcRLA IP, monoclonal anti-FcRLA (24) on protein G beads was used. For IP assessment of lymphoid architecture and flow cytometric analysis of IgM from Daudi and BJAB cell lines, streptavidin-coupled agarose beads of lymphocyte development and FcRLA expression, the following Abs were were precoated with biotinylated F(ab9)2 anti-human IgM, anti-k, or anti-L used. Anti-mouse CD3ε-FITC, CD4-PE, CD8-biotin, IgD-FITC, IgM-PE, (BD Pharmingen). Western blotting for FcRLA was performed using rabbit CD23-PE, CD21-FITC, B220-biotin, CD11b-FITC, CD5-FITC, and strepta- anti-FcRLA antiserum 840 and goat anti-rabbit IgG-HRP. vidin-allophycocyanin were obtained from BD Pharmingen (San Diego, CA). Biotinylated peanut agglutinin was obtained from Vector Laboratories Measurement of Ag-specific Ab responses (Burlingame, CA) and alkaline phosphatase-conjugated anti-rat IgG was obtained from Southern Biotechnology Associates (Birmingham, AL). For determination of the effect of FcRLA deficiency on the development of Abs used for ELISAs to quantify total mouse serum Ig or Ag-specific Ig humoral immune responses, mice were immunized by i.p. injection of Ag at were as follows. Purified anti-IgM, anti-IgG2b, anti-IgG3, anti-IgA, and the doses and time points indicated in the Results section. For consistency anti-IgE were obtained from BD Pharmingen along with biotinylated and to reduce injection errors, all of the Ags were administered in a total Abs for detection of IgG3 and IgA. Purified anti-IgG1 and anti-IgG2a were volume of 200 ml in sterile PBS. For assays using sheep RBCs (SRBCs) as obtained from Southern Biotechnology Associates along with HRP- an Ag, SRBCs were freshly prepared from whole sheep blood (Colorado conjugated Abs specific for mouse Ig-k, Ig-l, IgM, IgG1, IgG2a, IgG2c, Serum Company, Denver, CO). Measurement of total serum Igs was per- and IgE as well as biotinylated anti-IgG2b and streptavidin-HRP. Purified formed by sandwich ELISA. Measurement of Ag-specific Ig was per- anti-IgG2c was obtained from Bethyl Laboratories (Montgomery, TX). formed using direct ELISA by coating wells with 100 mlofa5 mg/ml Ag solution. For measurement of anti-SRBC Abs, plates were Immunohistochemistry and FcRLA secretion ELISA coated to confluency with glutaraldehyde-fixed SRBCs. Endogenous per- oxidase was quenched with 0.3% hydrogen peroxide prior to incubation The expression pattern of mouse FcRLA within spleen was determined by 2/2 with diluted mouse sera. The substrate used for all of the ELISAs was 1 immunohistochemistry. Wild-type C57BL/6 and Fcrla mice were im- mg/ml ortho-phenylenediamine dihydrochloride (Sigma-Aldrich). Report- munized with 50 mg nitrophenyl (NP)-KLH (Biosearch Technologies, ing of ELISA data occurs in one of three ways: mean absorbance at 490 Novato, CA), and organs were harvested 10 d later and frozen in OCT nm; by titers defined as the mean absorbance plus or minus three standard compound. Six-micrometer cryosections were stained with the indicated deviations of the last three (negative) wells in the titration; and relative Abs in PBS/0.1% Tween 20/5% goat serum after a 10-min pretreatment units derived from direct comparison of the experimental serum with a hy- with 0.3% hydrogen peroxide to quench endogenous peroxidase activity.
Load more

Recommended publications
  • Fcrl5 and T-Bet Define Influenza-Specific Memory B Cells That Predict Long-Lived Antibody 2 Responses 3 4 Anoma Nellore1, Christopher D
    bioRxiv preprint doi: https://doi.org/10.1101/643973; this version posted May 20, 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. 1 Fcrl5 and T-bet define influenza-specific memory B cells that predict long-lived antibody 2 responses 3 4 Anoma Nellore1, Christopher D. Scharer2, Rodney G. King3, Christopher M. Tipton4, Esther 5 Zumaquero3, Christopher Fucile3,5, Betty Mousseau3, John E. Bradley6, Kevin Macon3, Tian Mi2, 6 Paul A. Goepfert1,3, John F. Kearney,3 Jeremy M. Boss,2 Troy D. Randall6, Ignacio Sanz4, 7 Alexander Rosenberg3,5, Frances E. Lund3 8 9 10 11 1Dept. of Medicine, Division of Infectious Disease, 3Dept. of Microbiology, 5Informatics Institute, 12 6Dept of Medicine, Division of Clinical Immunology and Rheumatology, at The University of 13 Alabama at Birmingham, Birmingham, AL 35294 USA 14 15 2Dept. of Microbiology and Immunology and 4Department of Medicine, Division of 16 Rheumatology, at Emory University, Atlanta, GA 30322, USA 17 18 19 20 21 22 23 24 25 26 27 28 Lead Contact and to whom correspondence should be addressed: [email protected] 29 30 Mailing address: Frances E. Lund, PhD 31 Charles H. McCauley Professor and Chair 32 Dept of Microbiology 33 University of Alabama at Birmingham (UAB) 34 Dept of Microbiology 35 University of Alabama at Birmingham (UAB) 36 276 BBRB Box 11 37 1720 2nd Avenue South 38 Birmingham AL 35294-2170 39 40 SHORT RUNNING TITLE: Effector memory B cell development after influenza vaccination 41 bioRxiv preprint doi: https://doi.org/10.1101/643973; this version posted May 20, 2019.
    [Show full text]
  • The Genomic Organization and Expression Pattern of the Low-Affinity Fc Gamma Receptors (Fcγr) in the Göttingen Minipig
    Immunogenetics (2019) 71:123–136 https://doi.org/10.1007/s00251-018-01099-1 ORIGINAL ARTICLE The genomic organization and expression pattern of the low-affinity Fc gamma receptors (FcγR) in the Göttingen minipig Jerome Egli1 & Roland Schmucki1 & Benjamin Loos1 & Stephan Reichl1 & Nils Grabole1 & Andreas Roller1 & Martin Ebeling1 & Alex Odermatt2 & Antonio Iglesias1 Received: 9 August 2018 /Accepted: 24 November 2018 /Published online: 18 December 2018 # The Author(s) 2018 Abstract Safety and efficacy of therapeutic antibodies are often dependent on their interaction with Fc receptors for IgG (FcγRs). The Göttingen minipig represents a valuable species for biomedical research but its use in preclinical studies with therapeutic antibodies is hampered by the lack of knowledge about the porcine FcγRs. Genome analysis and sequencing now enabled the localization of the previously described FcγRIIIa in the orthologous location to human FCGR3A. In addition, we identified nearby the gene coding for the hitherto undescribed putative porcine FcγRIIa. The 1′241 bp long FCGR2A cDNA translates to a 274aa transmembrane protein containing an extracellular region with high similarity to human and cattle FcγRIIa. Like in cattle, the intracellular part does not contain an immunoreceptor tyrosine-based activation motif (ITAM) as in human FcγRIIa. Flow cytometry of the whole blood and single-cell RNA sequencing of peripheral blood mononuclear cells (PBMCs) of Göttingen minipigs revealed the expression profile of all porcine FcγRs which is compared to human and mouse. The new FcγRIIa is mainly expressed on platelets making the minipig a good model to study IgG-mediated platelet activation and aggregation. In contrast to humans, minipig blood monocytes were found to express inhibitory FcγRIIb that could lead to the underestimation of FcγR-mediated effects of monocytes observed in minipig studies with therapeutic antibodies.
    [Show full text]
  • Genetic and Genomic Analysis of Hyperlipidemia, Obesity and Diabetes Using (C57BL/6J × TALLYHO/Jngj) F2 Mice
    University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Nutrition Publications and Other Works Nutrition 12-19-2010 Genetic and genomic analysis of hyperlipidemia, obesity and diabetes using (C57BL/6J × TALLYHO/JngJ) F2 mice Taryn P. Stewart Marshall University Hyoung Y. Kim University of Tennessee - Knoxville, [email protected] Arnold M. Saxton University of Tennessee - Knoxville, [email protected] Jung H. Kim Marshall University Follow this and additional works at: https://trace.tennessee.edu/utk_nutrpubs Part of the Animal Sciences Commons, and the Nutrition Commons Recommended Citation BMC Genomics 2010, 11:713 doi:10.1186/1471-2164-11-713 This Article is brought to you for free and open access by the Nutrition at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Nutrition Publications and Other Works by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. Stewart et al. BMC Genomics 2010, 11:713 http://www.biomedcentral.com/1471-2164/11/713 RESEARCH ARTICLE Open Access Genetic and genomic analysis of hyperlipidemia, obesity and diabetes using (C57BL/6J × TALLYHO/JngJ) F2 mice Taryn P Stewart1, Hyoung Yon Kim2, Arnold M Saxton3, Jung Han Kim1* Abstract Background: Type 2 diabetes (T2D) is the most common form of diabetes in humans and is closely associated with dyslipidemia and obesity that magnifies the mortality and morbidity related to T2D. The genetic contribution to human T2D and related metabolic disorders is evident, and mostly follows polygenic inheritance. The TALLYHO/ JngJ (TH) mice are a polygenic model for T2D characterized by obesity, hyperinsulinemia, impaired glucose uptake and tolerance, hyperlipidemia, and hyperglycemia.
    [Show full text]
  • Supplementary Table 1: Adhesion Genes Data Set
    Supplementary Table 1: Adhesion genes data set PROBE Entrez Gene ID Celera Gene ID Gene_Symbol Gene_Name 160832 1 hCG201364.3 A1BG alpha-1-B glycoprotein 223658 1 hCG201364.3 A1BG alpha-1-B glycoprotein 212988 102 hCG40040.3 ADAM10 ADAM metallopeptidase domain 10 133411 4185 hCG28232.2 ADAM11 ADAM metallopeptidase domain 11 110695 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 195222 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 165344 8751 hCG20021.3 ADAM15 ADAM metallopeptidase domain 15 (metargidin) 189065 6868 null ADAM17 ADAM metallopeptidase domain 17 (tumor necrosis factor, alpha, converting enzyme) 108119 8728 hCG15398.4 ADAM19 ADAM metallopeptidase domain 19 (meltrin beta) 117763 8748 hCG20675.3 ADAM20 ADAM metallopeptidase domain 20 126448 8747 hCG1785634.2 ADAM21 ADAM metallopeptidase domain 21 208981 8747 hCG1785634.2|hCG2042897 ADAM21 ADAM metallopeptidase domain 21 180903 53616 hCG17212.4 ADAM22 ADAM metallopeptidase domain 22 177272 8745 hCG1811623.1 ADAM23 ADAM metallopeptidase domain 23 102384 10863 hCG1818505.1 ADAM28 ADAM metallopeptidase domain 28 119968 11086 hCG1786734.2 ADAM29 ADAM metallopeptidase domain 29 205542 11085 hCG1997196.1 ADAM30 ADAM metallopeptidase domain 30 148417 80332 hCG39255.4 ADAM33 ADAM metallopeptidase domain 33 140492 8756 hCG1789002.2 ADAM7 ADAM metallopeptidase domain 7 122603 101 hCG1816947.1 ADAM8 ADAM metallopeptidase domain 8 183965 8754 hCG1996391 ADAM9 ADAM metallopeptidase domain 9 (meltrin gamma) 129974 27299 hCG15447.3 ADAMDEC1 ADAM-like,
    [Show full text]
  • Cellular and Molecular Signatures in the Disease Tissue of Early
    Cellular and Molecular Signatures in the Disease Tissue of Early Rheumatoid Arthritis Stratify Clinical Response to csDMARD-Therapy and Predict Radiographic Progression Frances Humby1,* Myles Lewis1,* Nandhini Ramamoorthi2, Jason Hackney3, Michael Barnes1, Michele Bombardieri1, Francesca Setiadi2, Stephen Kelly1, Fabiola Bene1, Maria di Cicco1, Sudeh Riahi1, Vidalba Rocher-Ros1, Nora Ng1, Ilias Lazorou1, Rebecca E. Hands1, Desiree van der Heijde4, Robert Landewé5, Annette van der Helm-van Mil4, Alberto Cauli6, Iain B. McInnes7, Christopher D. Buckley8, Ernest Choy9, Peter Taylor10, Michael J. Townsend2 & Costantino Pitzalis1 1Centre for Experimental Medicine and Rheumatology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, Charterhouse Square, London EC1M 6BQ, UK. Departments of 2Biomarker Discovery OMNI, 3Bioinformatics and Computational Biology, Genentech Research and Early Development, South San Francisco, California 94080 USA 4Department of Rheumatology, Leiden University Medical Center, The Netherlands 5Department of Clinical Immunology & Rheumatology, Amsterdam Rheumatology & Immunology Center, Amsterdam, The Netherlands 6Rheumatology Unit, Department of Medical Sciences, Policlinico of the University of Cagliari, Cagliari, Italy 7Institute of Infection, Immunity and Inflammation, University of Glasgow, Glasgow G12 8TA, UK 8Rheumatology Research Group, Institute of Inflammation and Ageing (IIA), University of Birmingham, Birmingham B15 2WB, UK 9Institute of
    [Show full text]
  • Characteristics of B Cell-Associated Gene Expression in Patients With
    MOLECULAR MEDICINE REPORTS 13: 4113-4121, 2016 Characteristics of B cell-associated gene expression in patients with coronary artery disease WENWEN YAN*, HAOMING SONG*, JINFA JIANG, WENJUN XU, ZHU GONG, QIANGLIN DUAN, CHUANGRONG LI, YUAN XIE and LEMIN WANG Department of Internal Medicine, Division of Cardiology, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China Received May 19, 2015; Accepted February 12, 2016 DOI: 10.3892/mmr.2016.5029 Abstract. The current study aimed to identify differentially with the two other groups. Additionally the gene expression expressed B cell-associated genes in peripheral blood mono- levels of B cell regulatory genes were measured. In patients nuclear cells and observe the changes in B cell activation at with AMI, CR1, LILRB2, LILRB3 and VAV1 mRNA expres- different stages of coronary artery disease. Groups of patients sion levels were statistically increased, whereas, CS1 and IL4I1 with acute myocardial infarction (AMI) and stable angina (SA), mRNAs were significantly reduced compared with the SA and as well as healthy volunteers, were recruited into the study control groups. There was no statistically significant difference (n=20 per group). Whole human genome microarray analysis in B cell-associated gene expression levels between patients was performed to examine the expression of B cell-associated with SA and the control group. The present study identified the genes among these three groups. The mRNA expression levels downregulation of genes associated with BCRs, B2 cells and of 60 genes associated with B cell activity and regulation were B cell regulators in patients with AMI, indicating a weakened measured using reverse transcription-quantitative polymerase T cell-B cell interaction and reduced B2 cell activation during chain reaction.
    [Show full text]
  • Mouse Fcrla Conditional Knockout Project (CRISPR/Cas9)
    https://www.alphaknockout.com Mouse Fcrla Conditional Knockout Project (CRISPR/Cas9) Objective: To create a Fcrla conditional knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering. Strategy summary: The Fcrla gene (NCBI Reference Sequence: NM_001160215 ; Ensembl: ENSMUSG00000038421 ) is located on Mouse chromosome 1. 5 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 5 (Transcript: ENSMUST00000046322). Exon 2~4 will be selected as conditional knockout region (cKO region). Deletion of this region should result in the loss of function of the Mouse Fcrla gene. To engineer the targeting vector, homologous arms and cKO region will be generated by PCR using BAC clone RP23-100H5 as template. Cas9, gRNA and targeting vector will be co-injected into fertilized eggs for cKO Mouse production. The pups will be genotyped by PCR followed by sequencing analysis. Note: Mice homozygous for a targeted allele exhibit largely normal T-dependent and T-independent antibody responses with an increase in IgG1 after secondary challenge with sheep red blood cells. Exon 2~4 is not frameshift exon, and covers 69.69% of the coding region. The size of intron 1 for 5'-loxP site insertion: 5054 bp, and the size of intron 4 for 3'-loxP site insertion: 2439 bp. The size of effective cKO region: ~2074 bp. The cKO region does not have any other known gene. Page 1 of 8 https://www.alphaknockout.com Overview of the Targeting Strategy Wildtype allele 5' gRNA region gRNA region 3' 1 2 3 4 5 Targeting vector Targeted allele Constitutive KO allele (After Cre recombination) Legends Exon of mouse Fcrla Homology arm cKO region loxP site Page 2 of 8 https://www.alphaknockout.com Overview of the Dot Plot Window size: 10 bp Forward Reverse Complement Sequence 12 Note: The sequence of homologous arms and cKO region is aligned with itself to determine if there are tandem repeats.
    [Show full text]
  • Type of the Paper (Article
    Supplementary figures and tables E g r 1 F g f2 F g f7 1 0 * 5 1 0 * * e e e * g g g * n n n * a a a 8 4 * 8 h h h * c c c d d d * l l l o o o * f f f * n n n o o o 6 3 6 i i i s s s s s s e e e r r r p p p x x x e e e 4 2 4 e e e n n n e e e g g g e e e v v v i i i t t t 2 1 2 a a a l l l e e e R R R 0 0 0 c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d J a k 2 N o tc h 2 H if1 * 3 4 6 * * * e e e g g g n n n a a * * a * h h * h c c c 3 * d d * d l l l * o o o f f 2 f 4 n n n o o o i i i s s s s s s e e e r r 2 r p p p x x x e e e e e e n n n e e 1 e 2 g g g e e 1 e v v v i i i t t t a a a l l l e e e R R R 0 0 0 c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d Z e b 2 C d h 1 S n a i1 * * 7 1 .5 4 * * e e e g g g 6 n n n * a a a * h h h c c c 3 * d d d l l l 5 o o o f f f 1 .0 * n n n * o o o i i i 4 * s s s s s s e e e r r r 2 p p p x x x 3 e e e e e e n n n e e e 0 .5 g g g 2 e e e 1 v v v i i i t t t a a a * l l l e e e 1 * R R R 0 0 .0 0 c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d M m p 9 L o x V im 2 0 0 2 0 8 * * * e e e * g g g 1 5 0 * n n n * a a a * h h h * c c c 1 5 * 6 d d d l l l 1 0 0 o o o f f f n n n o o o i i i 5 0 s s s s s s * e e e r r r 1 0 4 3 0 p p p * x x x e e e * e e e n n n e e e 2 0 g g g e e e 5 2 v v v i i i t t t a a a l l l 1 0 e e e R R R 0 0 0 c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d Supplementary Figure 1.
    [Show full text]
  • Monoclonal Antibody-Based Therapy As a New Treatment Strategy in Multiple Myeloma
    Leukemia (2012) 26, 199–213 & 2012 Macmillan Publishers Limited All rights reserved 0887-6924/12 www.nature.com/leu REVIEW Monoclonal antibody-based therapy as a new treatment strategy in multiple myeloma NWCJ van de Donk1, S Kamps1, T Mutis2 and HM Lokhorst1 1Department of Hematology, University Medical Center Utrecht, Utrecht, The Netherlands and 2Department of Clinical Chemistry and Hematology, University Medical Center Utrecht, Utrecht, The Netherlands The introduction of autologous stem cell transplantation the myeloma patients achieved a partial response (PR) or stable combined with the introduction of immunomodulatory drugs disease following rituximab therapy. All these patients expressed (IMiDs) and proteasome inhibitors has significantly improved CD20 on their myeloma cells.2 However, as only B15–20% of survival of multiple myeloma patients. However, ultimately the majority of patients will develop refractory disease, indicating all myeloma patients express CD20 on their bone marrow the need for new treatment modalities. In preclinical and clinical plasma cells, new targets for immunotherapy need to be studies, promising results have been obtained with several identified. The search for other targets has led to the develop- monoclonal antibodies (mAbs) targeting the myeloma tumor ment of mAbs targeting growth factor receptors or adhesion cell or the bone marrow microenvironment. The mechanisms molecules on myeloma cells. Other newly developed mAbs underlying the therapeutic efficacy of these mAbs include are directed against cellular or non-cellular components of the direct induction of tumor cell apoptosis via inhibition or activation of target molecules, complement-dependent cyto- bone marrow microenvironment, resulting in the neutraliza- toxicity and antibody-dependent cell-mediated cytotoxicity tion of growth factors, inhibition of angiogenesis, modulation (ADCC).
    [Show full text]
  • Supplementary Table 1
    Supplementary Table 1. 492 genes are unique to 0 h post-heat timepoint. The name, p-value, fold change, location and family of each gene are indicated. Genes were filtered for an absolute value log2 ration 1.5 and a significance value of p ≤ 0.05. Symbol p-value Log Gene Name Location Family Ratio ABCA13 1.87E-02 3.292 ATP-binding cassette, sub-family unknown transporter A (ABC1), member 13 ABCB1 1.93E-02 −1.819 ATP-binding cassette, sub-family Plasma transporter B (MDR/TAP), member 1 Membrane ABCC3 2.83E-02 2.016 ATP-binding cassette, sub-family Plasma transporter C (CFTR/MRP), member 3 Membrane ABHD6 7.79E-03 −2.717 abhydrolase domain containing 6 Cytoplasm enzyme ACAT1 4.10E-02 3.009 acetyl-CoA acetyltransferase 1 Cytoplasm enzyme ACBD4 2.66E-03 1.722 acyl-CoA binding domain unknown other containing 4 ACSL5 1.86E-02 −2.876 acyl-CoA synthetase long-chain Cytoplasm enzyme family member 5 ADAM23 3.33E-02 −3.008 ADAM metallopeptidase domain Plasma peptidase 23 Membrane ADAM29 5.58E-03 3.463 ADAM metallopeptidase domain Plasma peptidase 29 Membrane ADAMTS17 2.67E-04 3.051 ADAM metallopeptidase with Extracellular other thrombospondin type 1 motif, 17 Space ADCYAP1R1 1.20E-02 1.848 adenylate cyclase activating Plasma G-protein polypeptide 1 (pituitary) receptor Membrane coupled type I receptor ADH6 (includes 4.02E-02 −1.845 alcohol dehydrogenase 6 (class Cytoplasm enzyme EG:130) V) AHSA2 1.54E-04 −1.6 AHA1, activator of heat shock unknown other 90kDa protein ATPase homolog 2 (yeast) AK5 3.32E-02 1.658 adenylate kinase 5 Cytoplasm kinase AK7
    [Show full text]
  • Mouse and Human Fcr Effector Functions
    Pierre Bruhns Mouse and human FcR effector € Friederike Jonsson functions Authors’ addresses Summary: Mouse and human FcRs have been a major focus of Pierre Bruhns1,2, Friederike J€onsson1,2 attention not only of the scientific community, through the cloning 1Unite des Anticorps en Therapie et Pathologie, and characterization of novel receptors, and of the medical commu- Departement d’Immunologie, Institut Pasteur, Paris, nity, through the identification of polymorphisms and linkage to France. disease but also of the pharmaceutical community, through the iden- 2INSERM, U760, Paris, France. tification of FcRs as targets for therapy or engineering of Fc domains for the generation of enhanced therapeutic antibodies. The Correspondence to: availability of knockout mouse lines for every single mouse FcR, of Pierre Bruhns multiple or cell-specific—‘a la carte’—FcR knockouts and the Unite des Anticorps en Therapie et Pathologie increasing generation of hFcR transgenics enable powerful in vivo Departement d’Immunologie approaches for the study of mouse and human FcR biology. Institut Pasteur This review will present the landscape of the current FcR family, 25 rue du Docteur Roux their effector functions and the in vivo models at hand to study 75015 Paris, France them. These in vivo models were recently instrumental in re-defining Tel.: +33145688629 the properties and effector functions of FcRs that had been over- e-mail: [email protected] looked or discarded from previous analyses. A particular focus will be made on the (mis)concepts on the role of high-affinity Acknowledgements IgG receptors in vivo and on results from antibody engineering We thank our colleagues for advice: Ulrich Blank & Renato to enhance or abrogate antibody effector functions mediated by Monteiro (FacultedeMedecine Site X.
    [Show full text]
  • Genomic Organization of Mouse Fcy Receptor Genes
    Proc. Nail. Acad. Sci. USA Vol. 87, pp. 2856-2860, April 1990 Immunology Genomic organization of mouse Fcy receptor genes (Fc receptors/exon-ntron junctions/protein domains/immunoglobulin gene superfamily) ANTHONY KULCZYCKI, JR.*t, JENNIFER WEBBER*, HAUNANI A. SOARES*, MICHAEL D. ONKEN*, JAMES A. THOMPSON*, DAVID D. CHAPLIN*t, DENNIS Y. LOH*t, AND JEFFREY P. TILLINGHAST* *Department of Medicine and tHoward Hughes Medical Institute, Washington University School of Medicine, Saint Louis, MO 63110 Communicated by David M. Kipnis, January 16, 1990 ABSTRACT We have isolated and characterized the gene responsible for intrathymic deletion of a large fraction of coding for the mouse Fc receptor that is termed Fc,,RHa. The potentially self-reactive T-cell clones (11-13). gene contains five exons and spans approximately 9 kilobases. An understanding of the relationship of the various Fc Unlike most members of the immunoglobulin gene superfam- receptor genes to each other, the factors influencing their ily, this gene utilizes multiple exons to encode its leader peptide. evolution, and molecular mechanisms regulating their The first exon encodes the hydrophobic region of the signal expression requires a detailed analysis of the structures of sequence; the second exon, which contains only 21 base pairs, their genes. Here we report the characterization of two encodes a segment of the signal peptidase recognition site; and members of this gene family. the beginning of the third exon encodes the predicted site of peptidase cleavage. The third and fourth exons each code for immunoglobulin-like extracellular domains. The fifth exon MATERIALS AND METHODS encodes the hydrophobic transmembrane domain and the Screening of Mouse cDNA and Genomic Libraries.
    [Show full text]