Functional properties of an alternative, tissue-specific promoter for human arylamine N-acetyltransferase 1.
Variable expression of human arylamine N-acetyltransferase 1 (NAT1) due to genetic polymorphism, gene regulation or environmental influences is associated with individual susceptibility to various cancers. Recent studies of NAT1 transcription showed that most mRNAs originate at a promoter, P1, located 11.8 kb upstream of the single open reading frame (ORF) exon. We have now characterized an alternative NAT1 promoter lying 51.5 kb upstream of the NAT1 ORF. In the present study, analysis of human RNAs representing 27 tissue types by reverse transcriptase-polymerase chain reaction (RT-PCR) and quantitative RT-PCR showed the upstream 51.5 kb promoter, designated P3, to be most active in specific tissues, including kidney, liver, lung, and trachea. All NAT1 P3 mRNAs included 5'-untranslated region (5'-UTR) internal exons of 61 and 175 nucleotides in addition to the 79 nucleotide 5'-UTR exon present in P1 mRNA. CAP-dependent amplification of 5'-P3 mRNA termini defined an 84 bp transcription start region in which most start sites are centrally clustered. The hepatoma-derived HepG2 cell line expressed a high level of P3 mRNA with the same spliced structure and start site pattern as found in normal tissues. A 435-bp minimal promoter was defined by transfection of HepG2 with luciferase expression constructs containing genomic fragments from the P3 start region. These findings imply a fundamental role for P3 in NAT1 regulation and define additional regions for genetic polymorphisms associated with enhanced cancer risk.
Barker
David F
DF
Department of Pharmacology and Toxicology and James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY 40292, USA.
Husain
Anwar
A
Neale
Jason R
JR
Martini
Benjamin D
BD
Zhang
Xiaoyan
X
Doll
Mark A
MA
States
J Christopher
JC
Hein
David W
DW
D001191:Arylamine N-Acetyltransferase / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics*; D001483:Base Sequence; D045744:Cell Line, Tumor; D005091:Exons; D005260:Female; D005786:Gene Expression Regulation; D020022:Genetic Predisposition to Disease; D006801:Humans; D007527:Isoenzymes / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics*; D008297:Male; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000235:genetics; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic
D007527:Isoenzymes; D012333:RNA, Messenger; D001191:Arylamine N-Acetyltransferase; C089384:N-acetyltransferase 1