UBD, a downstream element of FOXP3, allows the identification of LGALS3, a new marker of human regulatory T cells

Abstract

Here, we report the identification of the ubiquitin-like gene UBD as a downstream element of FOXP3 in human activated regulatory CD4 þ CD25 hi T cells (T reg ). Retroviral transduction of UBD in human allo-reactive effector CD4 þ 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 þ 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 þ CD25 hi derived regulatory T cells.

CD147 (Basigin/Emmprin) identifies FoxP3+CD45RO+CTLA4+-activated human regulatory T cells

“…In addition, we also identified several other proteins that seemed to be expressed at higher levels in CD25 enriched T cells of which some already have been shown to be specifically enriched in Tregs. [19][20][21] CD147 has previously been described to be expressed on activated lymphocyte populations and positively correlate with CD25 expression. 32 Here, we show that CD147 splits the FoxP3 population and directly identifies the CD45RO ϩ subset with a positive correlation with CTLA-4 and FoxP3 itself.…”

“…To increase the resolution and hence the number of proteins identified in our experiments, we performed subcellular fractionation of each T-cell subset before analysis of in-gel tryptic digests by nano-LC-ESI-LTQ Orbitrap mass spectrometry. From this we identified several candidates with a preferential expression on CD4 ϩ CD25 ϩ T cells including galectin-3, TNF receptor superfamily member 6 (CD95), transferrin receptor protein 1 (CD71) and receptor-type tyrosineprotein phosphatase (CD148), proteins that previously have been reported to be either up-regulated in Tregs 19,20 or regulated in a FoxP3-dependent manner. 21 We were also able to identify the specific expression of FoxP3 itself in our nucleic/insoluble cytoskeleton fractions.…”

Using transcriptional profiling to develop a diagnostic test of operational tolerance in liver transplant recipients

“…The expression pattern of a group of 68 target genes and 4 housekeeping genes (18S, GUS, HPRT1, and GAPDH) was measured by qPCR employing the ABI 7900 Sequence Detection System and LDA microfluidic PCR cards (PE Applied Biosystems) on peripheral blood samples obtained from 15 non-TOL, 16 TOL, and 16 CONT individuals. Selected target genes included the 24 genes identified by PAM, 44 genes selected among those most highly ranked in the SAM-derived gene list, and 6 genes (UBD, HLA-DOB, FOXP3, LTBP3, MAN1A1, LGALS3) selected on the basis of previous reports (11,23,26,33,34). To quantify the levels of mRNA, we normalized the expression of the target genes to the housekeeping gene HPRT1 (which was found to be the most stably expressed gene among the 4 housekeeping genes selected) and presented the results as relative expression between cDNA of the target samples and a calibrated sample according to the ΔCt method.…”

Intratumoral FOXP3 expression in infiltrating breast carcinoma: Its association with clinicopathologic parameters and angiogenesis.

“…The activity of these cells is known to be closely associated with the expression of forkhead/ winged helix transcription factor, FOXP3 (foxp3 in mice). Generally, foxp3 has been shown to be expressed exclusively in Tregs and plays a very important role in the development and function of these cells [6]. When foxp3 gene is introduced via retrovirus or enforced transgene expression, naïve CD4 ' CD25 ( T cells transform to Tregs [7].…”