Fig. 5.
CDK11p110 upregulates CBFβ expression directly by associating with its promoter. a Schematic representation of potential CDK11 binding sites in the CBFβ promoter and primer sets (p1, p2, p3, p4) indicating amplified regions encompassing the four primer sites along with the transcription start site (TSS). Chromatin immunoprecipitations were analyzed using a 2% input of KHOS sample treated with CDK11 siRNA by PCR. PCR products were only observed with p3 and p4 primer. b ChIP analysis was performed by CDK11 antibodies or 2% input sample and by measuring enrichment at p3 in human CBFΒ promoter by RT-PCR. The amount of immunoprecipitated DNA by CDK11 antibodies are represented as ratio of input DNA (1:50) and presented as mean of three independent experiments (n = 3, mean ± SD). *P < 0.05; **P < 0.01, Student’s t-test. c Electrophoretic mobility shift assay of CDK11- CBFβ binding activity in nuclear extracts from different cell lines. Metastatic cell lines MNNH/HOS and 143B demonstrated notable high binding activity (lane 3 and 4, purple arrow) compared with KHOS and U-2OS non-metastatic cell lines. d The formation of CDK11-DNA complexes was determined by incubation with labeled CBFβ. Decreased CDK11 DNA-binding activity was present in CDK11 siRNA knockdown KHOS and MNNH/HOS cells (purple arrow)