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Figure 4 | Journal of Hematology & Oncology

Figure 4

From: c-FLIP is involved in tumor progression of peripheral T-cell lymphoma and targeted by histone deacetylase inhibitors

Figure 4

HDACI-mediated c-FLIP downregulation was related to NF-κB inactivation via interrupting p50 interaction with c-FLIP. A, Apoptosis and NF-κB-associated gene expression were detected in Jurkat and H9 cells treated with VPA (0.5 mM) and SAHA (2 μM) for 72 h. The genes were classified as extrinsic apoptosis-related genes, NF-κB members and NF-κB target gene c-FLIP. B, Nuclear P65 and P50 expression, as well as whole cell p-IKKαβ, IKKαβ and p-IκBα expression were detected in Jurkat and H9 cells treated with VPA (0.5 mM) and SAHA (2 μM) for 72 h. C, Nuclear P50 expression was evaluated in Jurkat and H9 cells pre-incubated with TNF-α (10 ng/ml) for 4 h and treated with VPA (0.5 mM) and SAHA (2 μM) for 72 h. D, The activity of P50 binding to the c-FLIP promoter was measured by CHIP assay in Jurkat and H9 cells treated with VPA (0.5 mM) and SAHA (2 μM) for 72 h. DNA-protein complexes from Jurkat and H9 cells were precipitated with anti-P50 antibody and amplified with primers for the region of the binding site using PCR (Upper panel). Antibody against RNA Polymerase II was referred as the positive control. Non-specific Ig and PCR with primers amplifying the distal region of the c-FLIP promoter were referred as the negative control (Lower panel). E, The effect of P50 on transcriptional activity of the c-FLIP promoter was measured by luciferase reporter assay in HEK-293T cells transfected with negative control (P50 NC) or P50-overexpressing vector (P50 Over). *, P < 0.05 comparing with the negative control (NC). F, P50 and c-FLIP expression were detected in Jurkat cells transfected with Con siRNA or P50 siRNA. Cell apoptosis of Jurkat cells transfected with Con siRNA or P50 siRNA was measured. *, P < 0.05 comparing with the negative control (Con siRNA).

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