Background/Purpose:  Repeated immunization with exogenous antigen such as ovalbumin (OVA) induces SLE in mice otherwise not prone to spontaneous autoimmune diseases, in which autoantibody-inducing CD4 (ai CD4) T cell that had undergone de novo T cell receptor (TCR) revision was generated. The ai CD4 T cells induced varieties of autoantibody including anti-dsDNA antibody and matured CD8 T cells into effector cytotoxic T lymphocyte (CTL) via antigen cross-presentation in dendritic cell (DC), after which they caused tissue injuries (Tsumiyama K et al. PLoS ONE 4(12): e8382, 2009; J Immunol. 191: 91, 2013). We here studied the molecular mechanism of antigen cross-presentation to show that accumulation of cross-presentable antigen in the cytoplasm viaSec61, translocation channel of protein involved in ER-associated degradation (ERAD), finally induces lupus tissue injury in an experimental model based on our ‘self-organized criticality theory’ explaining the cause of SLE.

Methods:  Bone marrow-derived DC (BMDC) was generated from BALB/c mice. BALB/c mice were repeatedly immunized with ovalbumin (OVA) to induce SLE, and splenic DC (spDC) was isolated from these mice. DCs were cultured with fluorescent-labeled OVA, an inhibitor of Sec61 Exotoxin A and/or an inducer of ER stress Tunicamycin. Immunofluorescent staining, immunoprecipitation and immunoblotting were performed to detect early endosome antigen 1 (EEA1), calnexin, Sec61, OVA and unfolded protein response (UPR)-related molecules. OVA/MHC class I complex was detected under flow cytometry.

Results:  Engulfed OVA was co-localized with endosomal marker EEA1, which was then separated from EEA1 in DCs. OVA never co-localized with ER marker calnexin, whereas OVA was co-localized and co-precipitated with translocon Sec61. OVA in the cytoplasm of BMDC became undetectable when co-cultured with the inhibitor of Sec61 Exotoxin A, and thus, antigen was exported directly from endosome to cytoplasm via Sec61. When the amount of OVA accumulated in the cytoplasm was decreased dose-dependently by inhibiting Sec61 using Exotoxin A, the amount of OVA/MHC class I complex expressed on BMDC was significantly decreased. Instead, the amount of endosomal Sec61 and cytoplasmic OVA was both increased upon co-culture with an inducer of ER stress Tumicamycin in BMDC. Similarly, in the mice with lupus kidney disease generated upon repeated immunization with OVA, both endosomal Sec61 and cytoplasmic OVA were up-regulated in their spDC. Further, expression of UPR-related molecules including Bip, IRE1, XBP1 and PERK, and phosphorylated eIF2α was significantly up-regulated, which indicated that ER stress increases endosomal Sec61 thereby increasing the amount of antigen accumulated in the cytoplasm. The cytoplasmic antigen which is cross-presentable finally augments lupus tissue injury.

Conclusion: We found in the first that antigen was transported from endosome to cytoplasm via Sec61 for antigen cross-presentation. Second, antigen cross-presentation was promoted in proportion to the amount of antigen accumulated in cytoplasm of DC. Third, ER stress increased endosomal Sec61 thereby contributing to the induction of lupus tissue injury by facilitating antigen cross-presentation.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/endoplasmic-reticulum-stress-induces-lupus-kidney-disease-by-facilitating-antigen-cross-presentation-via-the-increase-of-endosomal-sec61/