Lys63-polyubiquitination by the E3 ligase casitas B-lineage lymphoma-b (Cbl-b) modulates peripheral regulatory T cell tolerance in patients with systemic lupus erythematosus

Abstract

T cells from systemic lupus erythematosus (SLE) patients display a wide array of anomalies in peripheral immune tolerance mechanisms. The role of ubiquitin ligases such as Cbl-b has been described recently in these phenomena. However, its role in resistance to suppression phenotype in SLE has not been characterized, which was the aim of the present study. Thirty SLE patients (20 with active disease and 10 with complete remission) and 30 age- and sex-matched healthy controls were recruited. Effector (CD4⁺CD25^–) and regulatory (CD4⁺CD25⁺) T cells (T_regs) were purified from peripheral blood mononuclear cells (PBMCs) by magnetic selection. Suppression assays were performed in autologous and allogeneic co-cultures and analysed by a flow cytometry assay. Cbl-b expression and lysine-63 (K63)-specific polyubiquitination profile were assessed by Western blotting. We found a defective Cbl-b expression in T_regs from lupus patients in contrast to healthy controls (1·1 ± 0·9 versus 2·5 ± 1·8, P = 0·003), which was related with resistance to suppression (r = 0·633, P = 0·039). Moreover, this feature was associated with deficient K63 polyubiquitination substrates and enhanced expression of phosphorylated signal transducer and activation of transcription 3 (pSTAT-3) in T_regs from lupus patients. Our findings support that Cbl-b modulates resistance to suppression by regulating the K63 polyubiquitination profile in lupus T_regs. In addition, defective K63 polyubiquitination of STAT-3 is related to increased pSTAT-3 expression, and might promote the loss of suppressive capacity of T_regs in lupus patients.