We have previously shown that the RNA binding protein, polypyrimidine tract-binding protein (PTBP1) plays a critical role in regulating the expression of CD40L in activated CD4 T cells. This is achieved mechanistically through message stabilization at late times of activation as well as by altered distribution of CD40L mRNA within distinct cellular compartments. PTBP1 has been implicated in many different processes, however whether PTBP1 plays a broader role in CD4 T cell activation is not known. To examine this question, experiments were designed to introduce shRNA into primary human CD4 T cells to achieve decreased, but not complete ablation of PTBP1 expression. Analyses of shPTB-expressing CD4 T cells revealed multiple processes including cell proliferation, activation-induced cell death and expression of activation markers and cytokines that were regulated in part by PTBP1 expression. Although there was an overall decrease in the steady-state level of several activation genes, only IL-2 and CD40L appeared to be regulated by PTBP1 at the level of RNA decay suggesting that PTBP1 is critical at different regulatory steps of expression that is gene-specific. Importantly, even though the IL-2 protein levels were reduced in cells with lowered PTBP1, the steady-state level of IL-2 mRNA was significantly higher in these cells suggesting a block at the translational level. Evaluation of T cell activation in shPTB-expressing T cells revealed that PTBP1 was linked primarily to the activation of the PLCγ1/ERK1/2 and the NF-κB pathways. Overall, our results reveal the importance of this critical RNA binding protein in multiple steps of T cell activation.