Transforming Growth Factor-β Directly Induces p53-up-regulated Modulator of Apoptosis (PUMA) during the Rapid Induction of Apoptosis in Myc-driven B-cell Lymphomas

c-Myc transformed human Burkitt's lymphoma (BL) cells are highly sensitive to TGF-beta-induced apoptosis. Previously we demonstrated that TGF-beta-mediated cell death in BL cells is regulated via the mitochondrial intrinsic apoptosis pathway, which is dependent on the activation of BAX and/or BAK. TGF-beta directly induces transcription of the BH3-only protein BIK and represses expression of the pro-survival factor BCL-X-L but has no effect on the direct BAX/BAK activators BIM or BID (tBID). Here we show that TGF-beta induces the BH3-only activator PUMA to aid induction of the intrinsic cell death pathway. TGF-beta also induced PUMA in normal germinal center CD77-positive centroblasts isolated from human tonsil tissue. PUMA was a direct TGF-beta target gene in B-cells, and we identify a putative Smad-binding region within the human PUMA promoter that recruits Smad3 and Smad4 in cells in response to TGF-beta signaling. Constitutive activity of the isolated Smadbinding region in luciferase reporter assays was dependent on Smad consensus sequences and was partially dependent on endogenous TGF-beta signaling and Smad4. Knockdown ofPUMA in BL cells using lentiviral shRNA resulted in slower kinetics of the TGF-beta -mediated apoptotic response. Analysis of E mu-Myc cell lines demonstrated that c-myc-driven murine lymphomas are also sensitive to TGF-beta-mediated apoptosis. Moreover, Puma(-/-) E mu-Myc lines demonstrated significantly delayed kinetics of the apoptotic response when compared with wild type lymphomas. TGF-beta therefore induces a polygenic response in Myc-driven lymphomas involving transcription of PUMA, which is necessary for the rapid induction of cell death.