Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.3.5.5 (CPS)
 1,262 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To study the regulation of transcription of the carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase (CAD) gene from the Syrian hamster, Mesocricetus auratus, we developed a homologous in vitro transcription system on the basis of nuclear extract from Syrian hamster kidney cells. We optimized the reaction temperature and the concentrations of DNA template, KCl, and MgCl2 simultaneously with the response surface method and found an unusually low temperature optimum of 20 degrees C. We therefore investigated whether CAD transcription in vitro depended on a heat-labile component of nuclear extract. Preincubating extract alone at 30 degrees C reduced transcription from the CAD promoter but not from the major late promoter of adenovirus 2. The formation of stable initiation complexes at the CAD promoter was diminished in heat-treated extract; run-off transcripts, however, accumulated at the same rate as in untreated extract. The heat sensitivity of complex formation correlated with the heat sensitivity of DNA binding by transcription factor Sp1, which binds to two sites in the CAD promoter; moreover, both preformed initiation complexes and DNA-bound Sp1 were heat-resistant. Adding purified Sp1 to heat-treated extract restored complex formation. We propose that Sp1 activates CAD transcription by stabilizing initiation complexes at the CAD promoter.
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PMID:Heat sensitivity and Sp1 activation of complex formation at the Syrian hamster carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase promoter in vitro. 134 30

Transcription of the carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase (CAD) gene from the Syrian hamster, Mesocricetus auratus, starts at a single major site. We characterized the cis-acting elements that position RNA polymerase II at the correct start site in the CAD promoter. Sequence alignment showed that the CAD promoter lacks a TATA box, but contains a consensus initiator. Mutational analysis of the CAD promoter demonstrated that the sequences between -81 and +26 were sufficient for accurate and efficient transcription in vitro and in vivo; binding sites for the transcription factor Sp1 around -70 and -49 were necessary for transcriptional activity. The binding site at -49 directed initiation about 50 base pairs downstream. A ubiquitous activator protein, Honk, bound to the CAD promoter between -30 and -12, but did not participate in start site selection. The sequences around +1, which contain the consensus initiator, contributed to promoter activity; however, the presence of a consensus initiator in this region was neither necessary nor sufficient for transcription. We concluded from these results that the Sp1 binding site at -49 substituted for the missing TATA box and played a major role in start site selection at the CAD promoter.
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PMID:Start site selection at the TATA-less carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase promoter. 790