DIFFERENTIAL REGULATION OF PITUITARY-SPECIFIC GENE-EXPRESSION BY INSULIN-LIKE-GROWTH-FACTOR-1 IN RAT PITUITARY GH4C1 AND GH3 CELLS

We have compared the influence of insulin-like growth factor 1 (IGF-1) on pituitary gene expression in the rat cell lines GH4C1 and GH3. Inc ubation with IGF-1 increased PRL messenger RNA (mRNA) levels in GH4C1 cells by 4- to 5-fold but decreased the levels of PRL transcripts in G H3 cells. In addition, the levels of GH-mRNA that were not affected by IGF-1 in GH4C1 cells were significantly inhibited by the growth facto r in GH3 cells. IGF-1 also decreased PRL and GH-mRNA response to T-3, retinoic acid, and Fk in GH3 cells. Stability of PRL or GH transcripts was not altered by IGF-1 in GH3 cells, suggesting that the inhibitory effect is exerted at a transcriptional level. The pituitary-specific transcription factor GHF-1/Pit-1 activates both the GH and PRL promote rs. As analyzed by Western blot, IGF-1 did not alter GHF-1/Pit-1 prote in levels in GH4C1 cells but reduced the levels of the transcription f actor in GH3 cells. This decrease is secondary to a reduction of GHF-1 /Pit-1 transcripts in IGF-1-treated GH3 cells. Thus, a different effec t of IGF-1 on the expression of GHF-1/Pit-1 in GH3 and GH4C1 cells is likely involved in the different regulation of GH and PRL gene in both cell types. IGF-I increases the activity of the PRL promoter in trans ient transfection assays in GH4C1 cells by a Ras-dependent mechanism. Expression of oncogenic Ras(Val12) mimics the effect of IGF-1, and the dominant negative Ras(Asn17) blocks IGF-l-mediated stimulation of the PRL promoter in GH4C1 cells. Although IGF-1 did not stimulate the PRL promoter in GH3 cells, Ras(Val12) strongly activated the promoter in these cells. Hence, the machinery to activate Ras-dependent signaling is intact in GH3 cells. Moreover, IGF-1 stimulates the mitogen-activat ed protein kinase in GH3 cells, showing that the components linking th e IGF-1 receptor to Ras are also active. These results suggest that, i n addition to the Ras/mitogen-activated protein kinase pathway, IGF-1 could activate a different pathway and that the combination of both is required to elicit PRL gene expression by the growth factor. This sec ond pathway may be defective in GH3 cells that respond to Ras but not to IGF-1.