ABSTRACT
Here, we report the identification of the ubiquitin-like gene UBD as a downstream element of FOXP3 in human activated regulatory CD4(+)CD25(hi) T cells (T(reg)). Retroviral transduction of UBD in human allo-reactive effector CD4(+) T helper (T(h)) cells upregulates CD25 and mediates downregulation of IL4 and IL5 expression similar to overexpression of FOXP3. Moreover, UBD impairs T(h) cell proliferation without upregulation of FOXP3 and impairs calcium mobilization. In the presence of ionomycin, overexpression of UBD in T(h) cells leads to the induction of IL1R2 that resemble FOXP3-transduced T(h) cells and naturally derived T(reg) cells. A comparison of the transcriptome of FOXP3- and UBD-transduced T(h) cells with T(reg) cells allowed the identification of the gene LGALS3. However, high levels of LGALS3 protein expression were observed only in human CD4(+)CD25(hi) derived T(reg) cells and FOXP3-transduced T(h) cells, whereas little was induced in UBD-transduced T(h) cells. Thus, UBD contributes to the anergic phenotype of human regulatory T cells and acts downstream in FOXP3 induced regulatory signaling pathways, including regulation of LGALS3 expression. High levels of LGALS3 expression represent a FOXP3-signature of human antigen-stimulated CD4(+)CD25(hi) derived regulatory T cells.
Subject(s)
Forkhead Transcription Factors/immunology , Galectin 3/immunology , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/metabolism , Ubiquitins/immunology , Biomarkers/metabolism , CD4 Antigens/immunology , Calcium/metabolism , Cell Proliferation , Cells, Cultured , Forkhead Transcription Factors/genetics , Galectin 3/genetics , Gene Expression Regulation/drug effects , Humans , Interleukin-4/immunology , Interleukin-4/metabolism , Interleukin-5/immunology , Interleukin-5/metabolism , Ionomycin/pharmacology , Ionophores/pharmacology , Receptors, Interleukin-1/biosynthesis , Receptors, Interleukin-1 Type II , Receptors, Interleukin-2/immunology , Retroviridae/genetics , T-Lymphocyte Subsets/immunology , T-Lymphocytes, Regulatory/drug effects , Transduction, Genetic , Ubiquitins/geneticsABSTRACT
Human HLA-B*3501 binds an antigenic peptide of 14-aa length derived from an alternative reading frame of M-CSF with high affinity. Due to its extraordinary length, the exact HLA binding mode was unpredictable. The crystal structure of HLA-B*3501 at 1.5 A shows that the N and C termini of the peptide are embedded in the A and F pockets, respectively, similar to a peptide of normal length. The central part of the 14-meric peptide bulges flexibly out of the groove. Two variants of the alternative reading frame of M-CSF peptide substituted at P2 or P2 and P9 with Ala display weak or no T cell activation. Their structure differs mainly in flexibility and conformation from the agonistic peptide. Moreover, the variants induce subtle changes of MHC alpha-helical regions implicated as critical for TCR contact. The TCR specifically recognizing this peptide/MHC complex exhibits CDR3 length within the normal range, suggesting major conformational adaptations of this receptor upon peptide/MHC binding. Thus, the potential antigenic repertoire recognizable by CTLs is larger than currently thought.
