RESUMEN
OBJECTIVE: Autoantibodies directed against tyrosine phosphatase IA-2 antibody (IA-2 Ab) are diagnostic for autoimmune type 1 diabetes. Conventional assays target the intracellular domain of IA-2. Among patients with ketosis-prone diabetes (KPD), characterized by presentation with diabetic ketoacidosis (DKA), >60% of adults lack three classic islet autoantibodies-IA-2, GAD65, and ZnT8 Abs-associated with type 1 diabetes. We aimed to determine whether apparently autoantibody-negative ("A-") KPD patients possess occult IA-2 Ab directed against full-length IA-2 (IA-2FL) or its extracellular domain (IA-2EC). RESEARCH DESIGN AND METHODS: We developed an assay that targets IA-2FL and IA-2EC and used it to analyze 288 subjects with A- KPD. RESULTS: Ten A- KPD patients were positive for IA-2EC Ab (3.5%), and three were also positive for IA-2FL Ab (1.0%), similar to frequencies in type 1 and type 2 diabetes. CONCLUSIONS: Measurement of IA-2FL Ab and IA-2EC Ab improves the accuracy of the Aß classification of KPD patients.
Asunto(s)
Autoanticuerpos/sangre , Cetoacidosis Diabética/clasificación , Cetoacidosis Diabética/diagnóstico , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/inmunología , Adolescente , Adulto , Biomarcadores/sangre , Niño , Estudios de Cohortes , Diabetes Mellitus Tipo 1/sangre , Diabetes Mellitus Tipo 1/complicaciones , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/complicaciones , Cetoacidosis Diabética/sangre , Diagnóstico Diferencial , Femenino , Humanos , Inmunoensayo/métodos , Masculino , Persona de Mediana Edad , Dominios Proteicos/inmunología , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Adulto JovenRESUMEN
Small oligomers formed early along human islet amyloid polypeptide (hIAPP) aggregation is responsible for the cell death in Type II diabetes. The epigallocatechin gallate (EGCG), a green tea extract, was found to inhibit hIAPP fibrillation. However, the inhibition mechanism and the conformational distribution of the smallest hIAPP oligomer - dimer are mostly unknown. Herein, we performed extensive replica exchange molecular dynamic simulations on hIAPP dimer with and without EGCG molecules. Extended hIAPP dimer conformations, with a collision cross section value similar to that observed by ion mobility-mass spectrometry, were observed in our simulations. Notably, these dimers adopt a three-stranded antiparallel ß-sheet and contain the previously reported ß-hairpin amyloidogenic precursor. We find that EGCG binding strongly blocks both the inter-peptide hydrophobic and aromatic-stacking interactions responsible for inter-peptide ß-sheet formation and intra-peptide interaction crucial for ß-hairpin formation, thus abolishes the three-stranded ß-sheet structures and leads to the formation of coil-rich conformations. Hydrophobic, aromatic-stacking, cation-π and hydrogen-bonding interactions jointly contribute to the EGCG-induced conformational shift. This study provides, on atomic level, the conformational ensemble of hIAPP dimer and the molecular mechanism by which EGCG inhibits hIAPP aggregation.
Asunto(s)
Catequina/análogos & derivados , Simulación de Dinámica Molecular , Extractos Vegetales/química , Multimerización de Proteína , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Té/química , Catequina/química , Humanos , Espectrometría de Masas , Estructura Cuaternaria de Proteína , Estructura Secundaria de ProteínaRESUMEN
BACKGROUND: ICA512 (or IA-2/PTPRN) is a transmembrane protein-tyrosine phosphatase located in secretory granules of neuroendocrine cells. Previous studies implied its involvement in generation, cargo storage, traffic, exocytosis and recycling of insulin secretory granules, as well as in ß-cell proliferation. While several ICA512 domains have been characterized, the function and structure of a large portion of its N-terminal extracellular (or lumenal) region are unknown. Here, we report a biophysical, biochemical, and functional characterization of ICA512-RESP18HD, a domain comprising residues 35 to 131 and homologous to regulated endocrine-specific protein 18 (RESP18). METHODS: Pure recombinant ICA512-RESP18HD was characterized by CD and fluorescence. Its binding to insulin and proinsulin was characterized by ELISA, surface plasmon resonance, and fluorescence anisotropy. Thiol reactivity was measured kinetically. Targeting of ΔRESP18HD ICA512-GFP to the membrane of insulinoma cells was monitored by immunofluorescence. RESULTS: ICA512-RESP18HD possesses a strong tendency to aggregate and polymerize via intermolecular disulfide formation, particularly at pH>4.5. Its cysteine residues are highly susceptible to oxidation forming an intramolecular disulfide between cysteine 53 and 62 and intermolecular disulfides via cysteine 40 and cysteine 47. The regulated sorting of ICA512 to secretory granules in INS-1 cells was impaired by deletion of RESP18HD. ICA512-RESP18HD binds with high-affinity to insulin and proinsulin. CONCLUSIONS: RESP18HD is required for efficient sorting of ICA512 to secretory granules. GENERAL SIGNIFICANCE: RESP18HD is a key determinant for ICA512 granule targeting.
Asunto(s)
Insulina/metabolismo , Proteínas del Tejido Nervioso/química , Estructura Terciaria de Proteína/genética , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Secuencia de Aminoácidos/genética , Biofisica , Proliferación Celular/genética , Humanos , Insulina/química , Islotes Pancreáticos/química , Islotes Pancreáticos/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Células Neuroendocrinas/química , Células Neuroendocrinas/metabolismo , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/genética , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/metabolismo , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/metabolismo , Vesículas Secretoras/química , Vesículas Secretoras/metabolismoRESUMEN
AIMS/HYPOTHESIS: Insulinoma-associated protein 2 (IA-2) is a major target of autoimmunity in type 1 diabetes. When first detected, IA-2-autoantibodies commonly bind epitopes in the juxtamembrane (JM) domain of IA-2 and antibody responses subsequently spread to the tyrosine phosphatase domain. Definition of structures of epitopes in the JM domain, and genetic requirements for autoimmunity to these epitopes, is important for our understanding of initiation and progression of autoimmunity. The aims of this study were to investigate the contribution of individual amino acids in the IA-2 JM domain to antibody binding to these epitopes and the role of HLA genotypes in determining epitope specificity. METHODS: Regions of the JM domain recognised by autoantibodies were identified by peptide competition and inhibitory effects of alanine substitutions of residues within the JM region. Antibody binding was determined by radioligand binding assays using sera from patients genotyped for HLA-DRB1 and -DQB1 alleles. RESULTS: Patients were categorised into two distinct groups of JM antibody reactivity according to peptide inhibition. Inhibition by substitutions of individual amino acids within the JM domain differed between patients, indicating heterogeneity in epitope recognition. Cluster analysis defined six groups of residues having similar inhibitory effects on antibody binding, with three clusters showing differences in patients affected or unaffected by peptide. One cluster demonstrated significant differences in antibody binding between HLA-DRB1*04 and HLA-DRB1*07 patients and within DRB1*04 individuals; antibody recognition of a second cluster depended on expression of HLA-DQB1*0302. CONCLUSIONS/INTERPRETATION: The results identify amino acids contributing to distinct epitopes on IA-2, with both HLA-DR and HLA-DQ alleles influencing epitope specificity.
Asunto(s)
Autoanticuerpos/inmunología , Diabetes Mellitus Tipo 1/inmunología , Epítopos/inmunología , Cadenas beta de HLA-DQ/genética , Cadenas HLA-DRB1/genética , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/inmunología , Adolescente , Adulto , Alelos , Autoantígenos/química , Autoantígenos/inmunología , Membrana Celular/metabolismo , Niño , Epítopos/análisis , Femenino , Genotipo , Humanos , Masculino , Estructura Terciaria de Proteína , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Adulto JovenRESUMEN
Phogrin/IA-2ß and ICA512/IA-2 are two paralogs receptor-type protein-tyrosine phosphatases (RPTP) that localize in secretory granules of various neuroendocrine cells. In pancreatic islet ß-cells, they participate in the regulation of insulin secretion, ensuring proper granulogenesis, and ß-cell proliferation. The role of their cytoplasmic tail has been partially unveiled, while that of their luminal region remains unclear. To advance the understanding of its structure-function relationship, the X-ray structure of the mature ectodomain of phogrin (ME phogrin) at pH 7.4 and 4.6 has been solved at 1.95- and 2.01-Å resolution, respectively. Similarly to the ME of ICA512, ME phogrin adopts a ferredoxin-like fold: a sheet of four antiparallel ß-strands packed against two α-helices. Sequence conservation among vertebrates, plants and insects suggests that the structural similarity extends to all the receptor family. Crystallized ME phogrin is monomeric, in agreement with solution studies but in striking contrast with the behavior of homodimeric ME ICA512. The structural details that may cause the quaternary structure differences are analyzed. The results provide a basis for building models of the overall orientation and oligomerization state of the receptor in biological membranes.
Asunto(s)
Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Secuencia de Aminoácidos , Sitios de Unión/genética , Cristalografía por Rayos X , Concentración de Iones de Hidrógeno , Modelos Moleculares , Datos de Secuencia Molecular , Pliegue de Proteína , Multimerización de Proteína , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/genética , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/metabolismo , Homología de Secuencia de Aminoácido , Soluciones , Relación Estructura-ActividadRESUMEN
Autoantibodies to IA-2 in type 1 diabetes are associated with HLA-DR4, suggesting influences of HLA-DR4-restricted T cells on IA-2-specific B cell responses. The aim of this study was to investigate possible T-B cell collaboration by determining whether autoantibodies to IA-2 epitopes are associated with T cell responses to IA-2 peptides presented by DR4. T cells secreting the cytokines IFN-γ and IL-10 in response to seven peptides known to elicit T cell responses in type 1 diabetes were quantified by cytokine ELISPOT in HLA-typed patients characterized for Abs to IA-2 epitopes. T cell responses were detected to all peptides tested, but only IL-10 responses to 841-860 and 853-872 peptides were associated with DR4. Phenotyping by RT-PCR of FACS-sorted CD45RO(hi) T cells secreting IL-10 in response to these two peptides indicated that these expressed GATA-3 or T-bet, but not FOXP3, consistent with these being Th2 or Th1 memory T cells rather than of regulatory phenotype. T cell responses to the same two peptides were also associated with specific Abs: those to 841-860 peptide with Abs to juxtamembrane epitopes, which appear early in prediabetes, and those to peptide 853-872 with Abs to an epitope located in the 831-862 central region of the IA-2 tyrosine phosphatase domain. Abs to juxtamembrane and central region constructs were both DR4 associated. This study identifies a region of focus for B and T cell responses to IA-2 in HLA-DR4 diabetic patients that may explain HLA associations of IA-2 autoantibodies, and this region may provide a target for future immune intervention to prevent disease.
Asunto(s)
Autoantígenos/inmunología , Linfocitos B/inmunología , Diabetes Mellitus Tipo 1/inmunología , Epítopos/inmunología , Antígeno HLA-DR4/inmunología , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/inmunología , Linfocitos T/inmunología , Adolescente , Adulto , Alelos , Autoanticuerpos/inmunología , Linfocitos B/metabolismo , Niño , Diabetes Mellitus Tipo 1/genética , Femenino , Antígeno HLA-DR4/genética , Humanos , Inmunofenotipificación , Interleucina-10/biosíntesis , Masculino , Péptidos/inmunología , Fenotipo , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Linfocitos T/metabolismo , Adulto JovenRESUMEN
Cysteines are thought integral to conformational epitopes of islet antigen-2 (IA-2) autoantibodies (IA-2A), possibly through disulfide bond formation. We therefore investigated which cysteines are critical to IA-2A binding in patients with newly diagnosed type 1 diabetes. All 10 cysteines in the intracellular domain of IA-2 were modified to serine by site-directed mutagenesis, and the effects of these changes on autoantibody binding in comparison with wild-type control were investigated by radiobinding assay. Mutation of the protein tyrosine phosphatase (PTP) core cysteine (C909) in IA-2 caused large reductions in autoantibody binding. In contrast, little or no reduction in binding was seen following substitution of the other cysteines. Modification of the core cysteine (C945) in IA-2ß also greatly reduced autoantibody binding. Lysine substitution of glutamate-836 in IA-2 or glutamate-872 in IA-2ß resulted in modest reductions in binding and identified a second epitope region. Binding to IA-2 PTP and IA-2ß PTP was almost abolished by mutation of both the core cysteine and these glutamates. The core cysteine is key to the major PTP conformational epitope, but disulfide bonding contributes little to IA-2A epitope integrity. In most patients, at disease onset, >90% of antibodies binding to the PTP domain of IA-2 recognize just two epitope regions.
Asunto(s)
Autoanticuerpos/inmunología , Diabetes Mellitus Tipo 1/inmunología , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/inmunología , Adolescente , Adulto , Secuencia de Aminoácidos , Anticuerpos Monoclonales/inmunología , Niño , Preescolar , Cisteína/química , Mapeo Epitopo , Femenino , Humanos , Masculino , Datos de Secuencia Molecular , Estructura Terciaria de Proteína , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Relación Estructura-ActividadRESUMEN
The receptor-type protein-tyrosine phosphatase (RPTP) phogrin is localized at the membrane of secretory granules of pancreatic islet ß-cells and, similarly to the closely related ICA512, plays a role in the regulation of insulin secretion, in ensuring proper granulogenesis and stability, and in the regulation of ß-cell growth. The mature membraneproximal ectodomain of phogrin (MPE phogrin) was produced as a recombinant protein and characterized. CD, fluorescence, controlled proteolysis, size-exclusion chromatography, and multi-angle light scattering showed that it is a properlyfolded monomeric domain. Equilibrium experiments, in the presence of guanidinium chloride and thermal unfolding, suggest a two-state mechanism with a ΔG of 2.3-3.3 kcal/mol, respectively. The study establishes common features and differences of MPE phogrin and the homologous ectodomain of ICA512. A homology model of phogrin was built based in the x-ray structure of MPE ICA512. The model is a starting point for modeling the entire receptor and for testing the quaternary structure and interactions of this protein in vivo. A description of the membrane insertion mode and putative interacting surfaces of this large protein is fundamental for the understanding of its biological function.
Asunto(s)
Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/metabolismo , Animales , Dicroismo Circular , Ratones , Modelos Moleculares , Estructura Terciaria de Proteína , Desplegamiento Proteico , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , TermodinámicaRESUMEN
ICA512 (or IA-2) is a transmembrane protein-tyrosine phosphatase located in secretory granules of neuroendocrine cells. Initially, it was identified as one of the main antigens of autoimmune diabetes. Later, it was found that during insulin secretion, the cytoplasmic domain of ICA512 is cleaved and relocated to the nucleus, where it stimulates the transcription of the insulin gene. The role of the other parts of the receptor in insulin secretion is yet to be unveiled. The structures of the intracellular pseudocatalytic and mature extracellular domains are known, but the transmembrane domain and several intracellular and extracellular parts of the receptor are poorly characterized. Moreover the overall structure of the receptor remains to be established. We started to address this issue studying by X-ray crystallography the structure of the mature ectodomain of ICA512 (ME ICA512) and variants thereof. The variants and crystallization conditions were chosen with the purpose of exploring putative association interfaces, metal binding sites and all other structural details that might help, in subsequent works, to build a model of the entire receptor. Several structural features were clarified and three main different association modes of ME ICA512 were identified. The results provide essential pieces of information for the design of new experiments aimed to assess the structure in vivo.
Asunto(s)
Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Sitios de Unión , Calcio/química , Núcleo Celular/metabolismo , Cristalización , Cristalografía por Rayos X/métodos , ADN/metabolismo , Dimerización , Humanos , Concentración de Iones de Hidrógeno , Insulina/química , Modelos Moleculares , Conformación Molecular , Mapeo de Interacción de Proteínas , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Solventes/química , Propiedades de SuperficieRESUMEN
Phogrin, a receptor tyrosine phosphatase-like protein, is localized to dense-core secretory granules (SGs) in various neuroendocrine cells. A previous report showed that the N-terminal luminal domain mediates targeting of this protein to SGs in AtT-20 cells. Here, we show that the luminal domain specifically interacts with carboxypeptidase E (CPE), one of the key proteins involved in peptide hormone sorting, in a weakly acidic condition. The luminal domain consists of pro-sequence domain (pro) and subsequent N-side mature domain and the pro domain was preferentially required for phogrin interaction with CPE and for its targeting to SGs. Small interfering RNA-directed reduction of the CPE protein level resulted in an improper accumulation of phogrin at the trans-Golgi network in AtT-20 cells. This finding indicates that CPE is involved in the sorting process of phogrin to SGs. However, SG localization of CPE was hindered by overexpression of the phogrin mutants that lack the transport motif of binding to clathrin adaptor complexes. Phogrin-depleted AtT-20 cells also exhibited reduced CPE targeting and increased CPE degradation. Our results suggest that the luminal interaction between phogrin and CPE contributes to their targeting to SGs in a cooperative manner in neuroendocrine cells.
Asunto(s)
Carboxipeptidasa H/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/metabolismo , Vesículas Secretoras/metabolismo , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Secuencias de Aminoácidos , Animales , Carboxipeptidasa H/química , Línea Celular , Técnicas de Silenciamiento del Gen , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Ratones , Mutación , Células Neuroendocrinas/metabolismo , Hormonas Peptídicas/metabolismo , Unión Proteica , Transporte de Proteínas , ARN Interferente Pequeño/biosíntesis , ARN Interferente Pequeño/genética , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/genética , Vesículas Secretoras/enzimología , Red trans-Golgi/metabolismoRESUMEN
Diabetes mellitus in dogs shares many characteristics with the human type 1 disease and virtually all diabetic dogs require insulin therapy to control hyperglycaemia. Insulin deficiency is suspected to result from immune-mediated destruction of pancreatic beta cells in some cases. Human patients suffering from Type 1A (immune-mediated) diabetes or latent autoimmune diabetes of the adult (LADA) demonstrate circulating autoantibodies against the 65kDa isoform of glutamic acid decarboxylase (GAD65) and/or insulinoma antigen-2 (IA-2). The aims of the current study were to develop radio-immunoassays to detect serum antibodies against recombinant canine GAD65 and IA-2 and to identify diabetic dogs showing serological evidence of autoreactivity to these pancreatic beta cell antigens. Canine GAD65 and the 3' end of IA-2 (coding for amino acids 771-979 of the intracellular domain) were amplified by PCR from cDNA prepared from canine insulinoma tissue and cloned into the pCRII vector. The canine sequences were later confirmed by identifying GAD2 and PTPRN genes from the dog genome assembly. Recombinant (35)S-methionine-radiolabelled canine GAD65 and IA-2 (771-979) proteins were used in radio-immunoprecipitation assays to screen sera from 30 newly diagnosed diabetic dogs and 30 control dogs. Four of 30 canine diabetic patients had significant GAD65 autoreactivity (p<0.01) compared to controls and 3 dogs were positive for autoantibodies to IA-2 (771-979). Two diabetic dogs showed dual autoantigen reactivity. These preliminary data indicate that serological reactivity to GAD65 and IA-2 is present in a proportion of diabetic dogs and suggests that, in some cases, canine diabetes is associated with an autoimmune response to these antigens.
Asunto(s)
Autoanticuerpos , Diabetes Mellitus Tipo 1/veterinaria , Enfermedades de los Perros/inmunología , Glutamato Descarboxilasa/inmunología , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/inmunología , Secuencia de Aminoácidos , Animales , Clonación Molecular , Diabetes Mellitus Tipo 1/inmunología , Perros , Femenino , Expresión Génica , Glutamato Descarboxilasa/química , Masculino , Datos de Secuencia Molecular , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/químicaRESUMEN
IA-2 (insulinoma-associated protein 2) is a protein-tyrosine phosphatase receptor located in secretory granules of neuroendocrine cells. Initially, it attracted attention due to its involvement in the autoimmune response associated to diabetes. Later it was found that upon exocytosis, the cytoplasmic domain of IA-2 is cleaved and relocated to the nucleus, where it enhances the transcription of the insulin gene. A concerted functioning of the whole receptor is to be expected. However, very little is known about the structure and function of the transmembrane and extracellular domains of IA-2. To address this issue, we solved the x-ray structure of the mature ectodomain of IA-2 (meIA-2) to 1.30A resolution. The fold of meIA-2 is related to the SEA (sea urchin sperm protein, enterokinase, agrin)) domains of mucins, suggesting its participation in adhesive contacts to the extracellular matrix and providing clues on how this kind of molecule may associate and form homo- and heterodimers. Moreover, we discovered that meIA-2 is self-proteolyzed in vitro by reactive oxygen species, suggesting the possibility of a new shedding mechanism that might be significant in normal function or pathological processes. Knowledge of meIA-2 structure should facilitate the search of its possible ligands and molecular interactions.
Asunto(s)
Modelos Moleculares , Pliegue de Proteína , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/química , Autoinmunidad/inmunología , Cristalografía por Rayos X , Diabetes Mellitus/genética , Diabetes Mellitus/inmunología , Exocitosis/genética , Exocitosis/inmunología , Matriz Extracelular , Humanos , Especies Reactivas de Oxígeno/química , Especies Reactivas de Oxígeno/inmunología , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/genética , Proteínas Tirosina Fosfatasas Clase 8 Similares a Receptores/inmunología , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Homología Estructural de Proteína , Relación Estructura-ActividadRESUMEN
IA2 and phogrin are important targets of humoral and cell-mediated autoimmunity in type 1 diabetes in man. They belong to a conserved subfamily of transmembrane protein tyrosine phosphatases (PTPs) associated with the regulatory pathway of secretion. To examine potential cross-reactivity between PTP family members we tested sera from T1D patients for reactivity to IA2, and the Drosophila (FLYDA) and C. elegans (IDA) orthologs using radioimmunoprecipitation assays of (35)S Met-labeled in vitro translated products of the cytosolic domains of these proteins. Approximately 80% of sera reacted with at least one probe. Of these, 82.5% showed reactivity to human IA2, 74.1% to FLYDA, and 33.7% to IDA. The majority of sera that bound FLYDA and/or IDA also recognized IA2. This raises the possibility that in some cases reactivity to IA2 may have arisen by molecular mimicry.