The regulation of CCR6 (chemokine receptor 6) expression during B-cell ontogeny and antigen-driven B-cell differentiation was analyzed. None of the CD34⁺Lin⁻ hematopoietic stem cell progenitors or the CD34⁺CD19⁺ (pro-B) or the CD19⁺CD10⁺ (pre-B/immature B cells) B-cell progenitors expressed CCR6. CCR6 is acquired when CD10 is lost and B-cell progeny matures, entering into the surface immunoglobulin D⁺ (sIgD⁺) mature B-cell pool. CCR6 is expressed by all bone marrow–, umbilical cord blood–, and peripheral blood–derived naive and/or memory B cells but is absent from germinal center (GC) B cells of secondary lymphoid organs. CCR6 is down-regulated after B-cell antigen receptor triggering and remains absent during differentiation into immunoglobulin-secreting plasma cells, whereas it is reacquired at the stage of post-GC memory B cells. Thus, within the B-cell compartment, CCR6 expression is restricted to functionally mature cells capable of responding to antigen challenge. In transmigration chemotactic assays, macrophage inflammatory protein (MIP)-3α/CC chemokine ligand 20 (CCL20) induced vigorous migration of B cells with differential chemotactic preference toward sIgD⁻ memory B cells. These data suggest that restricted patterns of CCR6 expression and MIP-3α/CCL20 responsiveness are integral parts of the process of B-lineage maturation and antigen-driven B-cell differentiation.