Fig. 1. Selective positive and negative coregulator effects by G9a on Runx2 target genes.
C4-2B/Rx2dox cells were infected with lentiviral vectors encoding shRNA targeting G9a mRNA (shG9a) or a non-specific sequence (shNS), and the infected populations were selected with puromycin. The two infected cell populations were cultured in medium supplemented with charcoal-stripped serum (CSS) containing dox (250 ng/ml) to induce Runx2 expression or equal volume of vehicle (distilled water) for 24 h before harvest. A: Immunoblots (left) were performed using antibodies against G9a, FLAG epitope (to detect Runx2), and actin as a control. mRNA levels were assessed by qRT-PCR (right). B & C: The mRNA levels for the indicated target genes of Runx2 were analyzed by qRT-PCR as described in Materials and Methods. All experiments were repeated at least three times and the representative results are shown. Abbreviations used: PIP, Prolactin-induced protein; MMP13 and MMP9, Matrix metalloproteinsase-13 and -9, respectively; PGC, Progastricsin-C; CSF-2, Colony-stimulating factor-2; SDF-1, Stromal differentiating factor-1; CST7, Cystatin-7.