Analysis of the promoter region of the gene encoding NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase.
Sequence analysis of the 5'-flanking region of the gene encoding NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase (NMDMC) revealed several putative cis-regulatory elements. To delineate the function of these regulatory elements, various deletion mutants of the 5'-flanking region were connected to the reporter gene chloramphenicol acetyltransferase (CAT) and promoter activity was measured in transient transfection assays. Transfection experiments performed with the sequence extending from -508 to +59 produced a high-level transient expression of the CAT gene in BALB/c 3T3-SV-T2 and NIH 3T3 cells. Removal of the sequence from +16 to +59 which includes the second transcription start point at +43, a TATA-like box and 5'-untranslated sequences abolished the promoter activity. Deletion analysis of 5'-upstream sequences revealed that the region from positions -55 to +59 is sufficient to mediate a high CAT activity comparable to the level obtained with the construct -508/+59. Within this region are found a CAAT box, a TATA-like box and two putative GC boxes. A functional analysis of the promoter showed that the sequence from -55 to +59 is sufficient to respond to stimulation by serum.
Bélanger
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Department of Biochemistry, McGill University, Montréal, Québec, Canada.
Peri
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MacKenzie
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D000619:Aminohydrolases / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D001769:Blood; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004252:DNA Mutational Analysis; D007700:Kinetics; D045064:Methenyltetrahydrofolate Cyclohydrolase; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP) / Q000235:genetics*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D016385:TATA Box; D014162:Transfection
D009097:Multienzyme Complexes; C055342:methylene tetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP); D015500:Chloramphenicol O-Acetyltransferase; D000619:Aminohydrolases; D045064:Methenyltetrahydrofolate Cyclohydrolase