Proinsulin degradation and presentation of a proinsulin B-chain autoantigen involves ER-associated protein degradation (ERAD)-enzyme UBE2G2.

Type 1 diabetes (T1D) is characterized by HLA class I-mediated presentation of autoantigens on the surface of pancreatic ß-cells. Recognition of these autoantigens by CD8+ T cells results in the destruction of pancreatic ß-cells and, consequently, insulin deficiency. Most epitopes presented at the surface of ß-cells derive from the insulin precursor molecule proinsulin. The intracellular processing pathway(s) involved in the generation of these peptides are poorly defined. In this study, we show that a proinsulin B-chain antigen (PPIB5-14) originates from proinsulin molecules that are processed by ER-associated protein degradation (ERAD) and thus originate from ER-resident proteins. Furthermore, screening genes encoding for E2 ubiquitin conjugating enzymes, we identified UBE2G2 to be involved in proinsulin degradation and subsequent presentation of the PPIB10-18 autoantigen. These insights into the pathway involved in the generation of insulin-derived peptides emphasize the importance of proinsulin processing in the ER to T1D pathogenesis and identify novel targets for future T1D therapies.

Associaçäo entre auto-anticorpos anti-pro-insulina e anti-insulina com diabetes mellitus do tipo I / The association between insulin and proinsulin autoantibodies with tipe I Diabetes Mellitus

O presente trabalho compara a incidência de auto-anticorpos anti-insulina (IAA) e anti-pró-insulina (PAA) em diabéticos do tipo I de início recente e em seus parentes de primeiro grau. Foram estudados 33 indivíduos normais (grupo I), 16 diabéticos do tipo I de início recente (grupo II) e 141 parentes em primeiro grau de diabéticos do tipo I (grupo III). Os IAA e PAA foram determinados pelo método de radioensaio, sendo considerados anormais níveis de IAA acima de 0,584 pmol/L e de PAA acima de 0,441 pmol/L. Näo foram observadas diferenças significantes quanto a idade e sexo entre os 3 grupos. Nos indivíduos normais os níveis de PAA foram significantemente menores do que os de IAA. Entre os diabéticos de início recente foi encontrada uma incidência de IAA de 37,5 por cento e de PAA de 25,0 por cento, näo ocorrendo, entretanto, diferenças significantes entre os níveis destes dois anticorpos. Entre os parentes em primeiro grau a incidência de IAA foi de 3,5 por cento e de PAA de 7,8 por cento, näo ocorrendo também diferenças entre os dois testes. Houve uma correlaçäo significante entre os níveis dos IAA e dos PAA no grupo de diabéticos de início recente (r=0,64; p < 0,05). Os IAA e PAA parecem estar dirigidos contra o mesmo determinante antigênico e, portanto, têm o mesmo valor preditivo para o DM tipo I

A specific and highly sensitive time-resolved fluoroimmunoassay for human proinsulin

We describe a time-resolved fluoroimmunoassay specific for human proinsulin using a combination of two high-affinity monoclonal antibodies, one against insulin and the other specific for intact proinsulin and for split 65-66 and des 64-65 proinsulin forms. The assay employs only 200 micro liters of serum, with a detection limit of 0.1 pmol/l. The intra-assay variation coefficient was less than 3 percent between 3 and 1000 pmol/l. There was 0 percent cross-reaction with insulin, C-peptide, split 32-33 and des 31-32 proinsulin. Serum concentration of proinsulin was analyzed in 50 subjects during an oral glucose tolerance test (10 non-obese controls, 10 obese controls, 10 subjects with impaired glucose tolerance, 10 patients with type II diabetes meIlitus (DM) and fasting blood glucose (FBG) <140 mg/dl, and 10 patients with type II DM and FBG >150 mg/dl). Mean fasting serum proinsulin levels measured by this assay in non-obese controls (0.84 +/-0.90 pmol/l; 0.1-2.4 pmol/l) were lower than the results reported by her investigators. There was an increase of proinsulin related to obesity and increased glucose levels, suggesting that proinsulin levels increase with insulin resistance.