Figure 6.

Activation of myc's transactivating activity induces ODC and cyclin D2, but not D1, and inhibition of c-myc expression correlates with inhibition of D2. BAC1.2F5 cell lines stably expressing myc-ER fusion proteins (as schematically shown in A) were established (see MATERIALS AND METHODS) and proliferation (clone 4) was studied (see Figure 4). Individual clones were isolated and tested for the myc-ER expression by anti-ER antibody immunoblot (B). Clone 4 was selected and either untreated or treated with CSF-1 (4.4 nM) or 4-HT (250 nM). Total RNA was isolated and subjected to Northern blot analyses (30 μg/lane). (C) ODC gene expression; (D) D2 gene expression; (E) D1 gene expression; and (F) GAPDH control. This is a representative experiment of three independent experiments by using the clone 4, 2, and 5. (G–K) BAC1.2F5 cells were treated with or without rottlerin (1 μM) in the presence or absence of CSF-1 (4.4 nM, 15 min for fos; 45 min for myc; and 5 h for D1 and D2), and total RNA was isolated and analyzed for expression of myc (G), D1 (H), fos (I), D2 (J), and GAPDH (K, GAPDH for D1 and D2). Similar levels of GAPDH control were obtained for fos and myc. The fold increases for c-fos and c-myc were calculated based on their own GAPDH levels.