CD4+ T cells (CD4+CD62LloCD44hi) and CD4+ICOS+ cells (Fig. 1, C–E; and not depicted). A significant accumulation of CXCR5+PD-1hiBcl-6hiCD4+ T cells and of a distinct popu- lation of CXCR5-/loPD-1hi CD4+ T cells expressing inter- mediate levels of Bcl-6 could also be observed (unpublished data). The number of regulatory T cells (CD4+CD25+Foxp3+) was also increased in the DKO female mice (unpublished data). Female mice deficient in DEF6 alone exhibited increased numbers of effector CD4+ T cells and CD4+ICOS+ cells, but the increase was more modest than that observed in the DKO female mice (Fig. 1, D and E). No T cell aberrancies were detected in Swap-70/ female mice (Fig. 1, D and E). Thus, the absence of both DEF6 and SWAP-70 results in the aber- rant accumulation of classical Tfh cells (CXCR5+PD-1hiBcl-6hi), and of a population that may represent extrafollicular T helper cells (CXCR5/loPD-1hiBcl-6int). Female DKO mice also exhibited marked aberrancies within the B cell compartment, which were characterized by an increased number of B cells (Fig. 1 F), an expansion of GC B cells (B220+PNA+), an increased number of switched IgG1+ B cells and an accumulation of plasma cells (B220loCD138+) in the spleen but not in the bone mar- row (Fig. 1, F–H; and not depicted). Slight increases in GC B cells and plasma cells were observed in mice deficient in DEF6 alone. Consistent with previous results (Quemeneur et al., 2008), in comparison to WT mice, mice lacking only SWAP-70 demonstrated a slightly increased frequency of plasma cells but decreased numbers of GC B cells (Fig. 1, G and H). Immunofluorescence studies confirmed the striking expansion of GCs and plasma cells in the spleens of the DKO female mice (Fig. 1 I). Collectively these studies indicate that the simultaneous absence of DEF6 and SWAP-70 results in the spontaneous accumulation of GC B cells and plasma cells. Lymphoproliferation is often associated with the devel- opment of autoantibodies and lupus-like disease. An exten- sive serologic and histopathologic analysis of the different groups of mice at one year of age was thus performed. Consis- tent with the finding that only DKO female mice developed lymphoproliferation, anti–double-stranded DNA (dsDNA) autoantibodies were detected exclusively in DKO female mice (Fig. 2 A). Autoantibody production in DKO female mice was associated with an increase in total serum IgG1 lev- els, but not in other IgG isotypes or in IgM and IgA produc- tion (unpublished data). Evaluation of sera from four WT and four DKO female mice with an autoantigen microarray (Fig. 2 B; and not depicted) confirmed these findings and demonstrated that the sera from DKO female mice reacted strongly against several nuclear antigens. 1-yr-old DKO fe- male mice were also found to exhibit glomerular deposition A r t i c l e of IgG and C3 and to develop glomerulonephritis (Fig. 2, C–E). Collectively, these results indicate that the concomitant absence of DEF6 and SWAP-70 results in the development of a lupus-like syndrome, which selectively affects the female gender. Remarkably, this occurs on a C57BL/6 background, a strain relatively resistant to the development of lupus. Young DKO female mice exhibit abnormal responses to T cell–dependent antigens Given the profound deregulation in humoral responses ob- served in the older DKO female mice, we sought to assess the responses of young DKO female mice to immuniza- tion with a T-dependent antigen, NP-KLH. Immunization with NP-KLH elicited a similar number of B220+NP+ cells in all four sets of mice (Fig. 3 A). As previously observed (Quemeneur et al., 2008), the lack of SWAP-70 resulted in a decreased number of NP+PNA+ B cells when compared with WT mice, whereas the absence of DEF6 led to a higher number of NP+PNA+ B cells (Fig. 3 B; and not depicted). The concomitant absence of DEF6 and SWAP-70 resulted in an increase in NP+PNA+ B cells comparable in magnitude to what was detected in the absence of DEF6 alone (Fig. 3 B; and not depicted). ELISPOT assays for total and high affinity NP-specific antibody forming B cells (AFCs) demonstrated that, as previously observed (Quemeneur et al., 2008), the lack of SWAP-70 resulted in an increased number of plasma cells when compared with WT mice, an effect that was asso- ciated with higher serum titers of anti–NP-specific IgG1 and IgG2a (Fig. 3, C–F). Interestingly, the lack of both DEF6 and SWAP-70 resulted in a more dramatic increase in plasma cells compared with mice lacking either SWAP-70 or DEF6 alone (Fig. 3, C–F). Similar results were also obtained upon immu- nization with SRBCs (unpublished data). Collectively, these data suggest that the lack of DEF6 may be primarily responsi- ble for the enhanced GC formation observed in the DKO mice, whereas the absence of both DEF6 and SWAP-70 contributes to the aberrant plasma cell formation detected in these mice. Aberrant production of IL-21 by DEF6-deficient T cells plays a key role in the humoral abnormalities observed in mice lacking DEF6 To further dissect the contribution of DEF6 and SWAP-70 to the aberrant humoral responses observed in the DKO fe- male mice, we set out to separately evaluate the effects of the lack of DEF6 and SWAP-70 in T and B cells. Because we had previously observed that the lack of DEF6 leads to the aberrant production of IL-17 and IL-21 when CD4+ T cells are cultured under neutral conditions (Chen et al., 2008; similar manner by the different sera (left dendrogram). (C, left) Representative H&E-stained sections from the kidneys of 1-yr-old female mice. Photomicro- graphs were taken at a magnification of 400×. Deposition of immunoglobulin complexes in the glomeruli was detected by immunofluorescence with anti-IgG (middle) and anti-C3 (right) staining. Magnification of 400× is shown. Results are representative of 5–7 mice/group. Bar, 250 µm. (D) The graph shows the renal histopathologic score of the indicated mice (n = 5–8/group). Data are representative of two independent experiments. (E) The graph on the left repre- sents the IgG deposition scores as detected by immunofluorescence, and the graph on the right represents the C3 deposition scores in the glomeruli of the indicated mice (n = 3–7/group). Data are representative of two independent experiments. ***, P < 0.005. JEM Vol. 209, No. 3 585