Multiple mechanisms of growth hormone-regulated gene transcription.

Diverse physiological actions of growth hormone (GH) are mediated by changes in gene transcription. Transcription can be regulated at several levels, including post-translational modification of transcription factors, and formation of multiprotein complexes involving transcription factors, co-regulators and additional nuclear proteins; these serve as targets for regulation by hormones and signaling pathways. Evidence that GH regulates transcription at multiple levels is exemplified by analysis of the proto-oncogene c-fos. Among the GH-regulated transcription factors on c-fos, C/EBPbeta appears to be key, since depletion of C/EBPbeta by RNA interference blocks the stimulation of c-fos by GH. The phosphorylation state of C/EBPbeta and its ability to activate transcription are regulated by GH through MAPK and PI3K/Akt-mediated signaling cascades. The acetylation of C/EBPbeta also contributes to its ability to activate c-fos transcription. These and other post-translational modifications of C/EBPbeta appear to be integrated for regulation of transcription by GH. The formation of nuclear proteins into complexes associated with DNA-bound transcription factors is also regulated by GH. Both C/EBPbeta and the co-activator p300 are recruited to c-fos in response to GH, altering c-fos promoter activation. In addition, GH rapidly induces spatio-temporal re-localization of C/EBPbeta within the nucleus. Thus, GH-regulated gene transcription mediated by C/EBPbeta reflects the integration of diverse mechanisms including post-translational modifications, modulation of protein complexes associated with DNA and re-localization of gene regulatory proteins. Similar integration involving other transcription factors, including Stats, appears to be a feature of regulation by GH of other gene targets.

Endogenous CCAAT/enhancer binding protein beta and p300 are both regulated by growth hormone to mediate transcriptional activation.

The regulation of c-fos transcription by GH involves multiple factors, including CCAAT/enhancer binding protein (C/EBP) beta. Knockdown of C/EBPbeta by RNA interference prevents stimulation of endogenous c-fos mRNA by GH, indicating a key role for C/EBPbeta in GH-stimulated c-fos transcription. GH rapidly increases the occupancy of both endogenous C/EBPbeta and p300 on the c-fos promoter in 3T3-F442A preadipocytes as indicated by chromatin immunoprecipitation. The transient occupancy of p300 on c-fos and the presence of p300 in the anti-C/EBPbeta immunoprecipitate coincide with the transient increase in c-fos transcription with GH, suggesting that a nuclear complex containing both p300 and C/EBPbeta occupies the c-fos promoter in response to GH. Expression of p300 with C/EBPbeta markedly increases c-fos promoter activity when neither alone is effective, indicating that p300 coactivates C/EBPbeta-mediated c-fos promoter activation. Such coactivation can determine a baseline for c-fos activation by GH. Furthermore, the occupancy of phosphorylated murine C/EBPbeta (T188) on c-fos upon GH treatment is simultaneous with increased occupancy by p300, suggesting that phospho-C/EBPbeta recruits p300 in response to GH. Thus, endogenous C/EBPbeta and p300 on c-fos are dynamically regulated by GH to determine transcriptional activation. Phosphorylated C/EBPbeta and p300 appear to function as part of a regulated complex that mediates GH-stimulated transcription.