The principal objective of this proposal is to elucidate transcriptional mechanisms involved in the regulation of neuroendocrine gene expression by neurotrophic, hormonal, and other stimuli. The gene encoding the neuroendocrine peptides neurotensin and neuromedin (NT/N gene) is expressed widely in the central nervous system, particularly in limbic regions, and is subject to strict multi-hormonal regulation in the catecholamine-producing PC12 cell line. NT has been co-localized to catecholamine-producing neurons and adrenal medullary cells, and increasing evidence suggests that NT modulates dopaminergic functions in the brain, Nerve growth factor acts cooperatively with adenylate cyclase activators and glucocorticoids to induce NT/N gene expression in PC12 cells, and experiments with different inducer combinations suggest that NGF plays a permissive role in the induction. Surprisingly, lithium can substitute for NGF in this permissive role. Transient transfection experiments indicate that both NGF and lithium influence NT/N gene expression principally through an AP-1 site located within a promoter proximal 200 bp region, but by distinct mechanisms which are likely to provide insights into novel modes of AP-1 regulation. The AP-1 site was originally identified in the metallothionein promoter and SV40 enhancer and is the target of a family of transcription factors that are structurally related to the proteins encoded by the c-fos and c-jun proto-oncogenes. Intensive investigation of these genes has provided evidence that the control of their expression is a principle means of regulating AP-1 activity. Lithium does not induce c-fos, jun-B, or c-jun gene expression, although the available evidence suggests that lithium may act through the induction of one or more AP-1 genes. NGF's permissive effect does not require new protein synthesis suggesting that NGF action results in the post-translational modification of preexisting AP-1 proteins. A central focus of this proposal is the identification and characterization of the AP-1 proteins involved in these responses. The ability of lithium to activate NTIN gene expression in PC12 cells is intriguing in view of its importance as therapeutic agent for the treatment of manic-depressive illness. The effect of chronic lithium treatment on NTIN gene expression in rats will be investigated to determine whether similar regulation occurs in vivo. Finally, transient transfection experiments will be used to identify cis-regulatory sequences required for constitutive NTIN gene expression in a NT-producing rat medullary thyroid carcinoma cell line.
| Xing, Y; Johnson, C V; Dobner, P R et al. (1993) Higher level organization of individual gene transcription and RNA splicing. Science 259:1326-30 |
