title: Uncommon structural motifs dominate the antigen binding site in human autoantibodies reactive with basement membrane collagen.
abstract: Autoantibodies mediate organ destruction in multiple autoimmune diseases, yet their origins in patients remain poorly understood. To probe the genetic origins and structure of disease-associated autoantibodies, we engrafted immunodeficient mice with human CD34+ hematopoietic stem cells and immunized with the non-collagenous-1 (NC1) domain of the alpha3 chain of type IV collagen. This antigen is expressed in lungs and kidneys and is targeted by autoantibodies in anti-glomerular basement membrane (GBM) nephritis and Goodpasture syndrome (GPS), prototypic human organ-specific autoimmune diseases. Using Epstein Barr virus transformation and cell fusion, six human anti-alpha3(IV)NC1 collagen monoclonal autoantibodies (mAb) were recovered, including subsets reactive with human kidney and with epitopes recognized by patients' IgG. Sequence analysis reveals a long to exceptionally long heavy chain complementarity determining region3 (HCDR3), the major site of antigen binding, in all six mAb. Mean HCDR3 length is 25.5 amino acids (range 20-36), generated from inherently long DH and JH genes and extended regions of non-templated N-nucleotides. Long HCDR3 are suited to forming noncontiguous antigen contacts and to binding recessed, immunologically silent epitopes hidden from conventional antibodies, as seen with self-antigen crossreactive broadly neutralizing anti-HIV Ig (bnAb). The anti-alpha3(IV)NC1 collagen mAb also show preferential use of unmutated variable region genes that are enriched among human chronic lymphocytic leukemia antibodies that share features with natural polyreactive Ig. Our findings suggest unexpected relationships between pathogenic anti-collagen Ig, bnAb, and autoreactive Ig associated with malignancy, all of which arise from B cells expressing unconventional structural elements that may require transient escape from tolerance for successful expansion.
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, N.C., USA; Durham VA Medical Center, Durham, N.C., USA; Duke Cancer Institute, Duke University Medical Center, Durham, N.C., USA. Electronic address: mary.h.foster@dm.duke.edu.
authors.forename: Mary H
authors.initials: MH
authors.lastname: Foster
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, N.C., USA. Electronic address: elizabeth.buckley@duke.edu.
authors.forename: Elizabeth S
authors.initials: ES
authors.lastname: Buckley
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, N.C., USA; Duke Cancer Institute, Duke University Medical Center, Durham, N.C., USA. Electronic address: benny.chen@dm.duke.edu.
authors.forename: Benny J
authors.initials: BJ
authors.lastname: Chen
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, N.C., USA; Duke Human Vaccine Institute, Duke University Medical Center, Durham, N.C., USA. Electronic address: kwan-ki.hwang@dm.duke.edu.
authors.forename: Kwan-Ki
authors.initials: KK
authors.lastname: Hwang
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, N.C., USA; Durham VA Medical Center, Durham, N.C., USA. Electronic address: amy.clark@duke.edu.
authors.forename: Amy G
authors.initials: AG
authors.lastname: Clark
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology; D001323:Autoantibodies / Q000235:genetics / Q000276:immunology*; D001324:Autoantigens / Q000276:immunology; D001485:Basement Membrane / Q000276:immunology*; D024141:Collagen Type IV / Q000276:immunology*; D022801:Complementarity Determining Regions / Q000235:genetics / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice
chemical_list: D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D001324:Autoantigens; D024141:Collagen Type IV; D022801:Complementarity Determining Regions
keywords: Anti-collagen autoimmunity; HCDR3 (heavy chain complementarity determining region 3); Human monoclonal autoantibody