The helix-loop-helix transcription factor Tall is required for blood cell development and its activation is a frequent event in T-cell acute lymphoblastic leukemia. The Akt (protein kinase B) kinase is a key player in transduction of anti-apoptotic and proliferative signals in T cells. Because Tall has a putative Akt phosphorylation site at Thr90, we investigated whether Akt regulates Tall. Our results show that Akt specifically phosphorylates Thr90 of the Tall protein within its transactivation domain in vitro and in vivo. Coimmunoprecipitation experiments showed the presence of Tall in Akt immune complexes, suggesting that Tall and Akt physically interact. We further showed that phosphorylation of Tall by Akt causes redistribution of Tall within the nucleus. Using luciferase assay, we showed that phosphorylation of Tall by Akt decreased represser activity of Tall on EpB42 (P4.2) promoter. Thus, these data indicate that Akt interacts with Tall and regulates Tall by phosphorylation at Thr90 in a phosphatidylinositol 3-kinase-dependent manner.