Effects of down-Regulating BCL11B Expression on the Proliferation and Apoptosis of Molt-4 Cells by RNA Interference

The B-cell chronic lymphocytic leukemia (CLL)/lymphoma 11B (BCL11B /CTIP2/ RIT1/hRitα) gene, which encodes C₂H₂ zinc-finger transcription factors, plays critical roles during development of T lymphocytes and the central nervous system. Recently, BCL11B gene has been identified to be associated with hematopoietic malignancies, such as T-cell acute lymphoblastic leukemia (T-ALL). However, little is known regarding the molecular mechanism by which BCL11B may contribute as a tumor-suppressor gene. In order to clarify the role of BCL11B gene in T-cell malignancies, we analyzed the effects of BCL11B target-specific small interfering RNA (siRNA) on the proliferation and apoptosis of Molt-4 cells, which is the naturally BCL11B overexpressing cell line. BCL11B-specific siRNA and scrambled non-silencing siRNA control were designed and synthesized by Invitrogen. The siRNAs were transfected into MOLT-4 cells by nucleofector techique. The level of BCL11B mRNA expression was analyzed at 48 and 72 hours after nucleofection by Real-time quantitative PCR with Taqman technique. The cells proliferation rate was assayed by MTT method at 24 and 48 hours after transfection, respectively. Cell morphology was showed by Liu’s Stain. The changes of cell cycle and apoptosis rate were detected by flow cytometry. The results showed that the level of BCL11B mRNA expression was inhibited in siRNA transfected cells at 72 hours after transfection, as compared with scrambled controls. The survival rates of siRNA-transfected cells were significantly decreased at both 24 and 48 hours (P < 0.01). Apoptotic cells could be observed morphologically. The apoptosis rates of Molt-4 cells treated with BCL11BsiRNA were highly increased compared with scrambled control ( P < 0.05). However, the percentage of different phases did not show significantly difference between siRNAtransfected cells and control cells, which was suggested that the apoptosis could be induced all phases of cell cycle. In conclusion, our data indicated that, the survival of Molt-4 cells might be critically dependent on BCL11B gene. Downregulation of BCL11B expression by RNA interference selectively can inhibit the proliferation and induced the apoptosis effectively in transformed Molt-4 cells, farther investigation will be performed in primary T-ALL cells to confirm the effect of presented BCL11B-siRNA on T-ALL.