Fig. 4

HDAC1 and HDAC6 cooperates to enhance RUNX2 expression in TPC1 thyroid cancer cells. Nuclear extract from TPC1 and MDA-MB231 cells were tested for the presence of a multi-protein complex controlling RUNX2 expression. Co-immunoprecipitation experiments were performed to evaluate binding of HDAC6 to HDAC1 (a-b), YAP (c-d), RUNX2 (e-f) and RNA Pol II (i-j). Western blots are representative of two independent experiments. ChIP assay show binding of YAP (g) and RUNX2 (h) to RUNX2 regulatory elements. ChIP experiments were also performed to evaluate levels of RNA Pol II on RUNX2 regulatory elements 48 h after transfection with siRNA specific for HDAC6 (k). Histogram represent the average enrichment of the indicated genomic regions in the immunoprecipitated DNA expressed as percentage of the Input. Data are expressed as mean values +/− SEM of a technical triplicate and are representative of at least two independent experiments. * p < 0.05. A schematic model illustrating how HDAC6 acts on RUNX2 transcription by stabilizing the interaction between the different regulating factors, thus enhancing the activity of the transcriptional complex (l)