Our previous studies have shown that Peyer's patches (PPs) play a key role in the induction of oral tolerance. Therefore, we hypothesized that PPs are an important site for Transforming Growth Factor (TGF)- β signaling and sought to prove that this tissue is of importance in oral tolerance induction. We found that expression of TGF-β type II receptor (TGFβRII) by CD4 + T cells increases and persists in the PPs of normal C57BL/6 mice after either high- or low-dose feeding of OVA when compared to mesenteric lymph nodes (MLNs) and spleen. Approximately one-third of these TGFβRII + CD4 + T cells express the transcription factor Foxp3. Interestingly, the number of TGFβRII + CD4 + T cells in PPs decreased when OVA-fed mice were orally challenged with OVA plus native cholera toxin (CT). In contrast, numbers of TGFβRII + CD4 + T cells were increased in the intestinal lamina propria (iLP) of these challenged mice. Further, these PP CD4 + TGFβRII + T cells upregulated Foxp3 within 2 hours after OVA plus CT challenge. Mice fed PBS and challenged with OVA plus CT did not reveal any changes in TGFβRII expression by CD4 + T cells. In order to test the functional property of TGFβRII in the induction of oral tolerance, CD4dnTGFβRII transgenic mice, in which TGFβRII signaling is abrogated from all CD4 + T cells, were employed. Importantly, these mice could not develop oral tolerance to OVA. Our studies show a critical, dose-independent, role for TGFβRII expression and function by CD4 + T cells in the gut-associated lymphoid tissues, further underlining the vital role of PPs in oral tolerance. © 2011 Gilbert et al.