Figure 1

Lrrc32 mRNA is preferentially expressed in naturally-occurring freshly-isolated non-expanded human T _regs but is not observed on the surface of these cells. a) T cell sorting gates based upon CD25 surface expression. b) Lrrc32 mRNA expression comports with Foxp3 mRNA expression and CD25 surface expression, and Lrrc32 mRNA is preferentially expressed in T_regs, compared to T_effs. Relative expressions of 18SrRNA-normalized Foxp3 and Lrrc32 genes were determined using real-time PCR. Data summarize four independent experiments. Results are expressed as the mean ± SEM. *p = 0.01 compared to CD25^mid cells, **p = 0.005 compared to CD25^mid cells. c) Flow cytometric analysis of freshly-isolated activated CD4⁺ human T cells shows that the CD25^hi subgroup (composed of the CD25hi+ and CD25hi++ subgroups denoted in figure 1b) demarcating T_regs expresses LRRC32 on the cell surface (top right panel) but that the CD25^- subgroup demarcating T_effs expresses negligible amounts of LRRC32 on the cell surface (bottom right panel). Analysis strategy is indicated via the arrows. d) Flow cytometric analysis of sorted resting CD4⁺ human T_regs shows that they lack surface expression of LRRC32 protein (top left panel). However, after permeabilization of the cell membrane, a low expression of LRRC32 protein can be observed intracellularly (top right panel). After activation of the sorted T_regs with a T_reg expansion kit, LRRC32 can be seen on the surface of these activated T_regs (bottom left panel), and evidence of intracellular expression of LRRC32 can also be observed after permeabilization of the cell membrane (bottom right panel).