Identification of a gastrin response element in the vesicular monoamine transporter type 2 promoter and requirement of 20 S proteasome subunits for transcriptional activity.
Vesicular monoamine transporter type 2 (VMAT2) is crucial for accumulation of monoamine neurotranmitters into neuronal secretory vesicles and histamine into secretory granules of the enterochromaffin-like cell in the acid-secreting gastric mucosa. Gastric VMAT2 expression is regulated by the antral hormone gastrin acting at the CCK(2) receptor. We demonstrate a gastrin response element (-56)ccgccccctc(-47) in the proximal VMAT2 promoter that binds in a gastrin-sensitive manner to nuclear proteins from gastric epithelial cell lines. Mutations within this sequence prevented nuclear protein binding and significantly reduced gastrin-stimulated expression of VMAT2 promoter-reporter constructs in gastric epithelial cells. In a yeast one-hybrid screen of an AR42J cell cDNA library, using the gastrin response element as bait, we identified a beta subunit of the 20 S proteasome, PSMB1, as a potential binding partner. In supershift assays, antibodies to PSMB1 and other proteasome beta subunits disrupted gastrin sensitive nuclear protein binding to the VMAT2 promoter. Moreover, RNA interference of PSMB1 significantly inhibited gastrin-mediated VMAT2 transcription. These data suggest that elements of the 20 S proteasome interact with the VMAT2 promoter to enhance G-protein-coupled receptor-mediated transcription.
Catlow
Krista
K
Physiological Laboratory, School of Biomedical Sciences, University of Liverpool, Liverpool L69 3BX, United Kingdom.
Ashurst
H Louise
HL
Varro
Andrea
A
Dimaline
Rod
R
D001483:Base Sequence; D004247:DNA; D024202:Electrophoretic Mobility Shift Assay; D005755:Gastrins / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011485:Protein Binding; D034622:RNA Interference; D014158:Transcription, Genetic; D050493:Vesicular Monoamine Transport Proteins / Q000235:genetics*
D005755:Gastrins; C493452:SLC18A2 protein, human; D050493:Vesicular Monoamine Transport Proteins; D004247:DNA; D046988:Proteasome Endopeptidase Complex