The RNA-binding protein, CUG-binding protein 1 (CUGBP1), regulates gene expression at the levels of alternative splicing, mRNA degradation, and translation. We used RNA immunoprecipitation followed by microarray analysis to identify the cytoplasmic mRNA targets of CUGBP1 in resting and activated primary human T cells and found that CUGBP1 targets were highly enriched for the presence of GU-rich elements (GREs) in their 3′-untranslated regions. The number of CUGBP1 target transcripts decreased dramatically following T cell activation as a result of activation-dependent phosphorylation of CUGBP1 and decreased ability of CUGBP1 to bind to GRE-containing RNA. A large percentage of CUGBP1 target transcripts exhibited rapid and transient up-regulation, and a smaller percentage exhibited transient down-regulation following T cell activation. Many of the transiently up-regulated CUGBP1 target transcripts encode important regulatory proteins necessary for transition from a quiescent state to a state of cellular activation and proliferation. Overall, our results show that CUGBP1 binding to certain GRE-containing target transcripts decreased following T cell activation through activation-dependent phosphorylation of CUGBP1.