Fig. 4.
SphK2 binding to p21 and c-fos promoters enhances acetylation of histone H3. (A and B) MCF-7 cells transfected with vector, V5-SphK2, or V5-SphK2G212E were treated with vehicle or PMA (100 nM, hatched bars) for 3 hours (A) or 30 min (B) and subjected to ChIP analyses with antibodies to V5, HDAC1, H3-K9ac, or normal rabbit IgG, as indicated. The precipitated DNA was analyzed by real-time PCR with primers amplifying the core promoter sequence of the p21 and c-fos genes. Relative binding to the promoter is expressed as the percentage of input. Data are means ± SD. **P < 0.01, relative to vector transfectants, *P < 0.01, relative to PMA-treated vector transfectants, by Student’s t test. (C and D) MCF-7 cells transfected with control siRNA or siSphK2 were treated with vehicle or PMA (100 nM, hatched bars) for 3 hours (C) or 30 min (D) and subjected to ChIP analyses with antibody against H3-K9ac or with normal rabbit IgG. *P < 0.01, relative to PMA-treated siControl. (E) Model for regulation of histone acetylation and gene transcription by nuclear SphK2 and S1P. PMA stimulates nuclear SphK2, which is associated with specific promoter regions, such as those for p21 and c-fos genes, and increases production of S1P. S1P in turn inhibits HDAC1 and HDAC2, which results in increased acetylation (Ac) of histone(s) and leads to enhanced gene transcription.