title: Redemption of autoantibodies on anergic B cells by variable-region glycosylation and mutation away from self-reactivity.
abstract: The best-understood mechanisms for achieving antibody self/non-self discrimination discard self-reactive antibodies before they can be tested for binding microbial antigens, potentially creating holes in the repertoire. Here we provide evidence for a complementary mechanism: retaining autoantibodies in the repertoire displayed as low levels of IgM and high IgD on anergic B cells, masking a varying proportion of autoantibody-binding sites with carbohydrates, and removing their self-reactivity by somatic hypermutation and selection in germinal centers (GCs). Analysis of human antibody sequences by deep sequencing of isotype-switched memory B cells or in IgG antibodies elicited against allogeneic RhD+ erythrocytes, vaccinia virus, rotavirus, or tetanus toxoid provides evidence for reactivation of anergic IgM(low) IgD+ IGHV4-34+ B cells and removal of cold agglutinin self-reactivity by hypermutation, often accompanied by mutations that inactivated an N-linked glycosylation sequon in complementarity-determining region 2 (CDR2). In a Hy10 antibody transgenic model where anergic B cells respond to a biophysically defined lysozyme epitope displayed on both foreign and self-antigens, cell transfers revealed that anergic IgM(low) IgD+ B cells form twice as many GC progeny as naïve IgM(hi) IgD+ counterparts. Their GC progeny were rapidly selected for CDR2 mutations that blocked 72% of antigen-binding sites with N-linked glycan, decreased affinity 100-fold, and then cleared the binding sites of blocking glycan. These results provide evidence for a mechanism to acquire self/non-self discrimination by somatic mutation away from self-reactivity, and reveal how varying the efficiency of N-glycosylation provides a mechanism to modulate antibody avidity.
authors.affiliation: Department of Immunology, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia;
authors.forename: Zahra
authors.initials: Z
authors.lastname: Sabouri
authors.affiliation: Immunology Division, Garvan Institute of Medical Research and.
authors.forename: Peter
authors.initials: P
authors.lastname: Schofield
authors.affiliation: Department of Immunology, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia;
authors.forename: Keisuke
authors.initials: K
authors.lastname: Horikawa
authors.affiliation: Department of Immunology, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia;
authors.forename: Emily
authors.initials: E
authors.lastname: Spierings
authors.affiliation: Institute of Cancer and Genetics, School of Medicine, Cardiff University, Cardiff CF14 4XN, United Kingdom; and.
authors.forename: David
authors.initials: D
authors.lastname: Kipling
authors.affiliation: Department of Immunology, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia;
authors.forename: Katrina L
authors.initials: KL
authors.lastname: Randall
authors.affiliation: Immunology Division, Garvan Institute of Medical Research and.
authors.forename: David
authors.initials: D
authors.lastname: Langley
authors.affiliation: Immunology Division, Garvan Institute of Medical Research and.
authors.forename: Brendan
authors.initials: B
authors.lastname: Roome
authors.affiliation: Immunology Division, Garvan Institute of Medical Research and.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Vazquez-Lombardi
authors.affiliation: Immunology Division, Garvan Institute of Medical Research and.
authors.forename: Romain
authors.identifier: http://orcid.org/0000-0003-4210-9613
authors.initials: R
authors.lastname: Rouet
authors.affiliation: Immunology Division, Garvan Institute of Medical Research and.
authors.forename: Jana
authors.initials: J
authors.lastname: Hermes
authors.affiliation: Immunology Division, Garvan Institute of Medical Research andSt Vincent's Clinical School, University of New South Wales, Darlinghurst, NSW 2010, Australia;
authors.forename: Tyani D
authors.initials: TD
authors.lastname: Chan
authors.affiliation: Immunology Division, Garvan Institute of Medical Research andSt Vincent's Clinical School, University of New South Wales, Darlinghurst, NSW 2010, Australia;
authors.forename: Robert
authors.initials: R
authors.lastname: Brink
authors.affiliation: Department of Immunobiology, King's College London School of Medicine, London SE1 9RT, United Kingdom.
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Dunn-Walters
authors.affiliation: Immunology Division, Garvan Institute of Medical Research andSt Vincent's Clinical School, University of New South Wales, Darlinghurst, NSW 2010, Australia; d.christ@garvan.org.au chris.goodnow@anu.edu.au.
authors.forename: Daniel
authors.initials: D
authors.lastname: Christ
authors.affiliation: Department of Immunology, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia; d.christ@garvan.org.au chris.goodnow@anu.edu.au.
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Goodnow
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D001323:Autoantibodies / Q000235:genetics / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D017635:Clonal Anergy / Q000276:immunology*; D005260:Female; D018858:Germinal Center / Q000276:immunology*; D006031:Glycosylation; D006801:Humans; D007072:Immunoglobulin D / Q000235:genetics / Q000276:immunology; D007075:Immunoglobulin M / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008875:Middle Aged; D027041:Somatic Hypermutation, Immunoglobulin / Q000235:genetics / Q000276:immunology*
chemical_list: D001323:Autoantibodies; D007072:Immunoglobulin D; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region
keywords: affinity maturation; autoimmunity; clonal selection; self-tolerance