We note that while our studies are under revision, another recently published report indicates that Dlg1 is not required for T-cell activation [30]. However, our study for the first time examines the requirement for Dlg1 in functional regulation of T cells with both TCR-fixed and

polyclonal (endogenously generated) T-cell repertoires by employing several experimental approaches in vivo. First, we tested the requirement for Dlg1 during Ag-driven T-cell clonal expansion in vivo. Using this immunization-based approach we found no evidence for Dlg1 involvement in T-cell activation Carfilzomib mw and clonal expansion by cognate Ag in vivo. We also tested if Dlg1 is required for homeostatic proliferation of T cells in lymphopenic hosts. This process is regulated by signals emanating from cytokine receptors and TCR upon its ligation with MHC/self peptide complexes in vivo [31]. IL-7 is produced abundantly in lymphopenic hosts and can drive homeostatic expansion of both naïve CD4+ and CD8+ T cells. In this context, the expansion of CD8+ T cells has been found to be more robust,

as compared with that see more of CD4+ T cells, presumably due to differential expression of IL-7R components [32]. Consistent with this view, homeostatic expansion of OT1 T cells in our experiments was markedly more robust as compared with OT2 T cells, however in both cases, we found no evidence for involvement of Dlg1 in homeostatic expansion of T cells. Thus, taken together, our in vitro and in vivo studies with TCR-transgenic

T cells do not implicate Dlg1 in TCR activation or T-cell proliferation of primary T cells. We also addressed the potential requirement for Dlg1 in the generation of memory T-cell subsets in vivo. Here, we focused on the endogenous polyclonal T-cell response, because the use of TCR-transgenic mouse models in studies of the Liothyronine Sodium kinetics of memory T-cell induction is thought to be nonphysiological. Thus, TCR-transgenic models can give biased results due to the high frequency of responding Ag-specific T cells and the abundance of Ag [33]. However, our analyses of polyclonal T-cell responses demonstrate significantly increased frequencies of IL-2 producing T cells upon boost immunization in KO mice, as compared with WT mice, although we can not rule out that Dlg1 may also be involved in T-cell migration and/or homing in vivo. However, we observe alterations in the frequencies of effector and central CD4+ T-cell memory subsets indicating that Dlg1 function may be cell autonomous. Given that previous studies have identified central memory CD4+ T cells as significant producers of IL-2 [34], the increased IL-2 production observed in our system most likely derives from Tcm cells.