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Mechanisms involved in the transcriptional activation of proenkephalin gene expression in bovine chromaffin cells



Mechanisms involved in the transcriptional activation of proenkephalin gene expression in bovine chromaffin cells



Journal of Biological Chemistry 265(31): 19116-19121



Stimulation of cultured bovine chromaffin cells with histamine (10-5 M), nicotine (10-6 M), and veratridine (2 .times. 10-6 M) results in a time-dependent up to 5-fold increase in proenkephalin (Penk) mRNA levels. After an initial lag phase (with no major alterations) Penk mRNA increased markedly between 6 and 12 h followed by a slower, steady increase up to 48 h. The nicotinic receptor antagonist tubocurarine (4 .times. 10-7 M) and the Ca2+ channel blocker D600 (10-5 M) prevent the subsequent rise of Penk mRNA levels after challenge with nicotine, when given within the lag phase (0-6 h), suggesting the need of continuous receptor occupation and Ca2+ entry for induction of gene expression. Similarly, incubation of chromaffin cells with cycloheximide (10-6 M), given at 0-6 h, blocks the increase in Penk mRNA after stimulation with histamine and nicotine indicating that ongoing protein synthesis is necessary for the delayed rise of Penk mRNA. Nuclear run-off experiments revealed high transcription levels of the Penk gene (3-fold at 2 h) and the tyrosine hydroxylase gene (7-fold at 20 min) following stimulation with histamine, which was not observed in the presence of cycloheximide (10-5 M). A more rapid induction of transcription was measured for the c-fos gene after histamine stimulation (high levels after 12 min) followed by c-fos mRNA accumulation (about 20-fold after a 1-h stimulation), which was superinduced when cells were pretreated with cycloheximide. The half-life of Penk mRNA levels (about 12 h), however, seems not to be affected by histamine as suggested by measurement of the subsequent decay of Penk mRNA levels after addition of .alpha.-amanitin or .alpha.-amanitin and cycloheximide. Thus, activation of Penk gene expression upon neurotransmitter challenge is suggested to be due to an enhanced transcriptional activity of the gene mediated by de novo synthesized protein(-like) factors.

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Accession: 007541299

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PMID: 2229066


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