This in turn inhibits the mTOR-dependent translation of mRNA transcripts, including Otub-1, thus maintaining GRAIL expression and inhibiting CD4 T cell proliferation . proliferation was assessed by CFSE staining, and the expression of GRAIL in splenic T cells was measured by real-time PCR, circulation cytometry and Western blot. We found increased GRAIL expression at the early stages of contamination, coinciding with the peak of parasitemia, with these findings correlating with impaired proliferation and poor IL-2 and IFN- secretion in response to plate-bound antibodies. In addition, we showed that this expression of GRAIL E3-ubiquitin ligase in CD4 T cells during the acute phase of contamination was complemented by a high expression of inhibitory receptors such as PD-1 and CTLA-4. We exhibited that GRAIL expression during contamination was Igf1 modulated by the mammalian target of the rapamycin (mTOR) pathway, Piribedil D8 since addition of IL-2 or CTLA-4 blockade in splenocytes from mice 21 days post contamination led to a reduction in GRAIL expression. Furthermore, addition of IL-2 was able to activate the mTOR pathway, inducing Otubain-1 expression, which mediated GRAIL degradation and improved T cell proliferation. Conclusions We hypothesize that GRAIL expression induced by the parasite may be managed by the increased expression of inhibitory molecules, which blocked mTOR activation and IL-2 secretion. Consequently, the GRAIL regulator Otubain-1 was not expressed and GRAIL managed the brake on T cell proliferation. Our findings reveal a novel association between increased GRAIL expression and impaired CD4 T cell proliferation during contamination. Author Summary Chagas disease is usually caused by the protozoan parasite and is endemic in Central and South America, where it affects about 10 million people. In addition, migration has led to the disease being established in non-endemic countries. Contamination involves an acute stage that evolves to a chronic stage where infected individuals may or may not Piribedil D8 show clinical symptoms or suffer progressive heart disease. The relevance of T cells in the control of contamination has been exhibited in human contamination Piribedil D8 and in experimental models. However, the parasite employs different strategies to downregulate the T cell function. These mechanisms can take action at the initial time of T cell activation, leading to a state of anergy where lymphocytes do not respond. However, the molecular components that regulate this process during contamination are not well comprehended. Our findings demonstrate for the first time that this T cell hyporesponsiveness could be linked to an increased expression of GRAIL. We propose that GRAIL expression induced by the parasite could be managed by increased expression of inhibitory molecules, which blocked mTOR activation and IL-2 secretion. GRAIL could then play a key role in downregulating T cell functions by allowing the parasites to establish the chronic disease. Piribedil D8 Introduction Chagas disease, caused by the intracellular protozoa is usually complex, requiring the generation of a substantial antibody response and the activation of both CD4 and CD8 T cell responses. Even in cases in which these responses are sufficiently stimulated to be able to control the acute contamination, is not completely eradicated, but instead persists in infected hosts for decades . employs a variety of strategies to evade the immune system and remain in the infected host. The main method entails the inhibition of specific T-cell responses, and consequently, can favor the establishment of chronic infections [4,5,6,7,8]. Related to this, a number of both host-dependent and parasite-induced mechanisms have been previously shown to impact immune regulation [9,10]. Moreover, T cells from infected hosts are largely unresponsive to antigens and mitogens, resulting in reduced IL-2 synthesis . IL-2 production initiates proliferation, effector functions, and clonal growth via IL-2 receptor (IL-2R)-mediated signaling . In the absence of a strong activation initiated by TCR and CD28 signaling, CD4 T cells fail to proliferate or to produce.