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XBP-1-deficient plasmablasts show normal protein folding but altered glycosylation and lipid synthesis.

ABSTRACT: The accumulation of misfolded secreted IgM in the endoplasmic reticulum (ER) of X-box binding protein 1 (XBP-1)-deficient B cells has been held responsible for the inability of such cells to yield plasma cells, through the failure to mount a proper unfolded protein response. LPS-stimulated B cells incapable of secreting IgM still activate the XBP-1 axis normally, as follows: XBP-1 is turned on by cues that trigger differentiation and not in response to accumulation of unfolded IgM, but the impact of XBP-1 deficiency on glycoprotein folding and assembly has not been explored. The lack of XBP-1 compromised neither the formation of functional hen egg lysozyme-specific IgM nor the secretion of free kappa-chains. Although XBP-1 deficiency affects the synthesis of some ER chaperones, including protein disulfide isomerase, their steady state levels do not drop below the threshold required for proper assembly and maturation of the Igalpha/Igbeta heterodimer and MHC molecules. Intracellular transport and surface display of integral membrane proteins are unaffected by XBP-1 deficiency. Given the fact that we failed to observe any defects in folding of a variety of glycoproteins, we looked for other means to explain the requirement for XBP-1 in plasma cell development. We observed significantly reduced levels of phosphatidylcholine, sphingomyelin, and phosphatidylinositol in total membranes of XBP-1-deficient B cells, and reduced ER content. Terminal N-linked glycosylation of IgM and class I MHC was altered in these cells. XBP-1 hence has important roles beyond folding proteins in the ER.

SUBMITTER: McGehee AM

PROVIDER: S-EPMC4053221 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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XBP-1-deficient plasmablasts show normal protein folding but altered glycosylation and lipid synthesis.

McGehee Annette M AM Dougan Stephanie K SK Klemm Elizabeth J EJ Shui Guanghou G Park Boyoun B Kim You-Me YM Watson Nicki N Wenk Markus R MR Ploegh Hidde L HL Hu Chih-Chi Andrew CC

Journal of immunology (Baltimore, Md. : 1950) 20090826 6

The accumulation of misfolded secreted IgM in the endoplasmic reticulum (ER) of X-box binding protein 1 (XBP-1)-deficient B cells has been held responsible for the inability of such cells to yield plasma cells, through the failure to mount a proper unfolded protein response. LPS-stimulated B cells incapable of secreting IgM still activate the XBP-1 axis normally, as follows: XBP-1 is turned on by cues that trigger differentiation and not in response to accumulation of unfolded IgM, but the impac ...[more]

PMID: 19710472

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Project description:BackgroundThe polymeric immunoglobulin receptor (pIgR) maintains the integrity of epithelial barriers by transporting polymeric antibodies and antigens through the epithelial mucosa into the lumen. In this study, we examined the role of pIgR in maintaining gut barrier integrity, which is important for the normal development in mice.MethodsCohorts of pIgR-/- mice and their wildtype controls were housed under Specific Pathogen Free (SPF) conditions and monitored for weight gain as an indicator of development over time. The general physiology of the gastrointestinal tract was analysed using immunohistochemistry in young (8-12 weeks of age) and aged mice (up to 18 months of age), and the observed immunopathology in pIgR-/- mice was further characterised using flow cytometry. Urinary metabolites were analysed using gas chromatography-mass spectrometry (GC-MS), which revealed changes in metabolites that correlated with age-related increase in gut permeability in pIgR-/- mice.ResultsWe observed that pIgR-/- mice exhibited delayed growth, and this phenomenon is associated with low-grade gut inflammation that increased with ageing. The gross intraepithelial lymphocytic (IEL) infiltration characteristic of pIgR-/- mice was redefined as CD8α+αβ+ T cells, the majority of which expressed high levels of CD103 and CD69 consistent with tissue resident memory T cells (TRM). Comparison of the urinary metabolome between pIgR-/- and wild-type mice revealed key changes in urinary biomarkers fucose, glycine and Vitamin B5, suggestive of altered mucosal permeability. A significant increase in gut permeability was confirmed by analysing the site-specific uptake of sugar probes in different parts of the intestine.ConclusionOur data show that loss of the secretory antibody system in mice results in enhanced accumulation of inflammatory IELs in the gut, which likely reflects ongoing inflammation in reaction to gut microbiota or food antigens, leading to delayed growth in pIgR-/- mice. We demonstrate that this leads to the presence of a unique urinary metabolome profile, which may provide a biomarker for altered gut permeability.

Project description:Aims/hypothesisIndividual variation in plasma N-glycosylation has mainly been studied in the context of diabetes complications, and its role in type 1 diabetes onset is largely unknown. Our aims were to undertake a detailed characterisation of the plasma and IgG N-glycomes in patients with recent onset type 1 diabetes, and to evaluate their discriminative potential in risk assessment.MethodsIn the first part of the study, plasma and IgG N-glycans were chromatographically analysed in a study population from the DanDiabKids registry, comprising 1917 children and adolescents (0.6-19.1 years) who were newly diagnosed with type 1 diabetes. A follow-up study compared the results for 188 of these participants with those for their 244 unaffected siblings. Correlation of N-glycan abundance with the levels and number of various autoantibodies (against IA-2, GAD, ZnT8R, ZnT8W), as well as with sex and age at diagnosis, were estimated by using general linear modelling. A disease predictive model was built using logistic mixed-model elastic net regression, and evaluated using a 10-fold cross-validation.ResultsOur study showed that onset of type 1 diabetes was associated with an increase in the proportion of plasma and IgG high-mannose and bisecting GlcNAc structures, a decrease in monogalactosylation, and an increase in IgG disialylation. ZnT8R autoantibody levels were associated with higher IgG digalactosylated glycan with bisecting GlcNAc. Finally, an increase in the number of autoantibodies (which is a better predictor of progression to overt diabetes than the level of any individual antibody) was accompanied by a decrease in the proportions of some of the highly branched plasma N-glycans. Models including age, sex and N-glycans yielded notable discriminative power between children with type 1 diabetes and their healthy siblings, with AUCs of 0.915 and 0.869 for addition of plasma and IgG N-glycans, respectively.Conclusions/interpretationWe defined N-glycan changes accompanying onset of type 1 diabetes, and developed a predictive model based on N-glycan profiles that could have valuable potential in risk assessment. Increasing the power of tests to identify individuals at risk of disease development would be a considerable asset for type 1 diabetes prevention trials.

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XBP-1-deficient plasmablasts show normal protein folding but altered glycosylation and lipid synthesis.

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XBP-1-deficient plasmablasts show normal protein folding but altered glycosylation and lipid synthesis.

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