Table S2 summarizes all the individual peptides identified in the MHC-II peptidome of the blood leukocytes isolated from 40 NOD mice after glucose challenge

Table S2 summarizes all the individual peptides identified in the MHC-II peptidome of the blood leukocytes isolated from 40 NOD mice after glucose challenge. from NOD mice without glucose challenge (control). JEM_20202530_TableS4.xlsx (21K) GUID:?7BA2D8D0-8715-4B92-B408-B29643FE4E90 Data Availability StatementThe MS data are available via ProteomeXchange with the dataset identifier PXD024400. Abstract Assessing the self-peptides presented by susceptible major histocompatibility complex (MHC) molecules is crucial for evaluating the pathogenesis and therapeutics of tissue-specific autoimmune diseases. However, direct examination of such MHC-bound peptides displayed in the target organ remains largely impractical. Here, we demonstrate that the blood leukocytes from the nonobese diabetic (NOD) mice presented peptide epitopes to autoreactive CD4 T cells. These peptides were bound to the autoimmune PEG6-(CH2CO2H)2 class II MHC molecule (MHC-II) I-Ag7 and originated from insulin B-chain and C-peptide. The presentation required a glucose challenge, which stimulated the release of the insulin peptides from the pancreatic islets. The circulating leukocytes, especially the B cells, promptly captured and presented these peptides. Mass spectrometry analysis of the leukocyte MHC-II peptidome revealed a series of cellCderived peptides, with identical sequences to those previously identified in the islet MHC-II peptidome. Thus, the blood leukocyte peptidome echoes that found in islets and serves to identify immunogenic peptides in an otherwise inaccessible tissue. Introduction Recognition of tissue-specific antigens is critical for initiating T cell responses driving autoimmunity. In type 1 diabetes (T1D), pancreatic islets are targeted by finely programmed autoimmune responses, leading to selective demise of the insulin-producing cells (Anderson and Bluestone, 2005). The primary genetic susceptibility of this disease is conferred by variants of the MHC-II alleles that bind peptide antigens derived from cells (Todd et al., 1987; Acha-Orbea and McDevitt, 1987; Miyazaki et al., 1990). These peptideCMHC-II complexes form the substrates for immune recognition by autoreactive CD4 T cells, resulting in their activation. Examining the antigenic entities displayed in the islets has substantially facilitated our understanding of the autoimmune process and provided insights into the development of targeted immunotherapies (Clemente-Casares et al., 2012; Unanue, 2014). Yet probing these tissue-derived antigens is limited by their inaccessibility. Recent investigations examined how cells convey their immunological information to the adaptive immune system (Vomund et al., 2015; Wan et al., 2018). Examining the non-obese diabetic (NOD) mouse that spontaneously develops autoimmune diabetes confirmed two sites for presentation of diabetogenic antigens: the peripheral lymphoid system and the islet (Wan et al., 2020). Secondary lymphoid tissues, particularly the pancreatic draining LN, are critical for the priming of the diabetogenic T cells (H?glund et al., 1999; Gagnerault et al., 2002; Levisetti et al., 2004); the local presentation in islets PEG6-(CH2CO2H)2 further enhances T cell pathogenicity (Melli et al., 2009; Ferris et al., 2014; Carrero et al., 2017). Peripheral lymphoid tissues are consistently sensitized by antigenic products secreted PEG6-(CH2CO2H)2 from the cells obeying glucose stimulation (Wan et al., 2018). These materials (referred to as secretome) contain catabolized peptide fragments generated in the cell crinosomes (Wan et al., 2018), a set of lysosomal vesicles that degrades excessive insulin granules to maintain cellular homeostasis (Smith and Farquhar, 1966; Halban and Wollheim, 1980; Orci et al., 1984). We have identified immunogenic epitopes from insulin, a prime antigen required for initiation of diabetes in NOD mice (Nakayama et al., 2005), in the crinosomes and the secretome (Wan et al., 2018). Diabetes development in the NOD mouse depends on the I-Ag7 MHC-II molecule (Lund et al., 1990; Singer et al., 1998; Gioia et al., 2019), a structural homologue to the human autoimmune HLA-DQ8 (Corper et al., 2000; Latek et al., 2000; Lee et al., 2001). Both molecules select a similar repertoire of self-peptides (Suri et al., 2005). We performed an unbiased analysis of the immunopeptidome of islets isolated from NOD mice and identified cellCspecific peptides bound to I-Ag7 (Wan et al., 2020). The major peptides that gave rise to autoreactivity derived from the insulin B-chain (InsB) and the C-peptide (InsC), which Rabbit polyclonal to ZNF131 were also identified in the MHC-II peptidomes of the pancreatic draining LN and spleen (Wan et al., 2020). The InsB-derived sequences were highly compatible with those that activated HLA-DQ8Crestricted CD4 T cells isolated from the peripheral blood or the islets in patients with T1D (Yang et al., 2014; Michels et.