CCR6⁺ Treg cells exhibit the phenotype of effector-memory T cells and have a high turnover rate in vivo. (A) Phenotypic characterization. Lymph node cells of naive mice were stained with αCD4, αCD25, and αCCR6 together with antibodies or fusion proteins, directed either against CCR7, CD44, CD11a, CD54, CD5, βTCR, CD69, or CD62L. Histograms are shown for the 4 subsets of CD25⁺CCR6⁺, CD25⁺CCR6^-, CD25^-CCR6⁺, and CD25^-CCR6^-CD4⁺ T cells and were generated by gating on the respective markers. Staining for CCR7 was carried out using a CCL19-Ig-fusion protein. Vertical bars are inserted as reference for relative shifts. One of 4 independent experiments each with cells pooled from 2 to 6 animals is shown. (B) In vivo turnover rate. FACS analysis of lymph node cells is shown for populations stained with αCCR6 (left panels) or αCD44 (right panels). Proliferating cells were labeled by providing BrdU with the drinking water for a period of 7 days (left columns) or 14 days (right columns). Cells were analyzed by FACS immediately after this period. Histograms of the BrdU-specific staining shown are gated on CD4⁺ cells. The percentages of BrdU⁺ cells are indicated. One of 2 independent experiments is shown.