Histidine decarboxylase (HDC) is highly expressed in enterochromaffin-like (ECL) cells, and carries out the conversion of L-histidine to histamine. In preliminary studies, we have demonstrated that the HDC gene is regulated both transcriptionally and post-transcriptionally by gastrin in transfected gastric cell lines. The promoter elements mediating gastrin responsiveness in the HDC gene have been identified as two novel cis-acting elements (GAS-RE1 and GAS-RE2) which are located in tandem downstream of the transcriptional start site, and bind to 52 kD and 35 kD proteins, respectively. The activation of HDC transcription by gastrin occurs through a MAP kinase-dependent pathway leading to increased promoter binding by GAS-REBP1 and GAS-REBP2. A candidate for GAS-REBP2 has been cloned from a human stomach cDNA library, and appears to represent a novel 35 kD DNA binding protein. Gastrin stimulation also leads to post-transitional HDC enzymatic activity, most likely through increases in translation. HDC activation also occurs through post-translational cleavage of the 74 kDa HDC precursor protein to a more active 54 kD enzymatic form. In preliminary studies, we have localized this cleavage site to an 8 amino acid region, and demonstrated that deletion of this region inhibits cleavage and enzymatic activation. Further, we have shown that expression of HDC protein leads to feedback inhibition of HDC promoter activity, and that this inhibition is independent of enzyme activity and mediated by N-terminal peptide sequences. Finally, we have shown that 4.8 kb of the mouse chromogranin A promoter are sufficient for targeting ECL cells and mediating gastrin responsiveness in transgenic mice. The proposed studies are aimed at investigating further the regulation of the HDC gene, using both in vitro and in vivo approaches. (1) The candidate GAS-REBP2 protein will be further characterized, and its role in HDC gene regulation investigated. (2) The HDC enzymatic cleavage site will be precisely defined, and translational regulation of HDC by gastrin will be analyzed. (3) The N-terminal HDC sequences mediating transcriptional inhibition will be mapped, and mechanisms of inhibition explored. (4) Finally, the cis-acting DNA sequences which are necessary for ECL cell-specific expression will be analyzed for the HDC gene using HDC-GFP reporter gene constructs. Overall, these studies will provide greater understanding of the mechanisms involved in the regulation of HDC gene expression and enzymatic activity by gastrin.
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