JunB Oncogenic Activity in Anaplastic Large Cell Lymphoma
V Atsaves, L Lekakis, M Feretzaki, V Leventaki, C Liakou, E Drakos, FX Claret, D Jones, LJ Medeiros, E Patsouris, GZ Rassidakis. The University of Texas M.D. Anderson Cancer Center, Houston, TX; National and Kapodistrian University of Athens, Athens, Greece
Background: JunB is a member of the jun family of AP-1 transcription factors involved in cell proliferation and survival. Recent evidence suggests that JunB is overexpressed in CD30+ lymphomas and positively regulates CD30 gene expression at the transcriptional level in Hodgkin lymphoma and anaplastic large cell lymphoma (ALCL) cells.We hypothesized that JunB operates as an oncogene in ALCL and may contribute to ALCL oncogenesis, at least in part, through activation of CD30 signalling leading to uncontrolled cell cycle progression.
Design: To investigate the underlying mechanism of JunB overexpression in ALCL, we used real-time quantitative PCR to assess JunB gene copy number in primary tissue samples obtained prior to treatment from 6 patients with ALCL. Transfection experiments were performed in human embryonic kidney (HEK) cell line 293T and in 2 ALCL cell lines, Karpas 299 and SU-DHL1. Protein expression, promoter activity and DNA binding were assessed by Western blot analysis, luciferase assay and EMSA method, respectively. BrdU incorporation and cell cycle were analyzed by flow cytometry.
Results: We show significant JunB gene amplification in the majority of ALCL tumor samples. Forced expression of JunB gene in HEK 293T resulted in CD30 upregulation. Inversely, silencing of JunB using siRNA in ALCL cell lines resulted in decreased CD30 expression and CD30 promoter activity. Silencing of JunB also resulted in decreased AP-1 binding activity in ALCL cells and significant decrease of the S-phase fraction of cell cycle. These changes were associated with increased levels of the CDK inhibitors p21 and p14 with simultaneous decrease of the unphosphorylated fraction of Rb and expression levels of cyclins A, D2 and D3. In addition, silencing of CD30 using siRNA in ALCL cells led to decreased AP-1 binding activity assessed by EMSA, which was associated with cell cycle arrest linked to decreased levels of p21 and p14 similar to those seen after JunB silencing.
Conclusions: Our data suggest that JunB functions as an oncogene in ALCL. JunB gene amplification might represent the initial genetic event that triggers uncontrolled cell cycle progression, at least in part, through activation of CD30 signaling and its downstream effectors in ALCL.
Wednesday, March 11, 2009 9:30 AM
Poster Session V # 184, Wednesday Morning