Journal Article

Functional analysis of arylamine <i>N</i>-acetyltransferase 1 (<i>NAT1</i>) <i>NAT1*10</i> haplotypes in a complete NATb mRNA construct

Lori M. Millner, Mark A. Doll, Marcus W. Stepp, J.Christopher States and David W. Hein

in Carcinogenesis

Volume 33, issue 2, pages 348-355
Published in print February 2012 | ISSN: 0143-3334
Published online November 2011 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/bgr273
Functional analysis of arylamine N-acetyltransferase 1 (NAT1) NAT1*10 haplotypes in a complete NATb mRNA construct

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N-acetyltransferase 1 (NAT1) catalyzes N-acetylation of arylamines as well as the O-acetylation of N-hydroxylated arylamines. O-acetylation leads to the formation of electrophilic intermediates that result in DNA adducts and mutations. NAT1*10 is the most common variant haplotype and is associated with increased risk for numerous cancers. NAT1 is transcribed from a major promoter, NATb, and an alternative promoter, NATa, resulting in messenger RNAs (mRNAs) with distinct 5′-untranslated regions (UTRs). To best mimic in vivo metabolism and the effect of NAT1*10 polymorphisms on polyadenylation usage, pcDNA5/Flp recombination target plasmid constructs were prepared for transfection of full-length human mRNAs including the 5′-UTR derived from NATb, the open reading frame and 888 nucleotides of the 3′-UTR. Following stable transfection of NAT1*4, NAT1*10 and an additional NAT1*10 variant (termed NAT1*10B) into nucleotide excision repair-deficient Chinese hamster ovary cells, N- and O-acetyltransferase activity (in vitro and in situ), mRNA and protein expression were higher in cells transfected with NAT1*10 and NAT1*10B than in cells transfected with NAT1*4 (P < 0.05). Consistent with NAT1 expression and activity, cytotoxicity and hypoxanthine phosphoribosyl transferase mutants following 4-aminobiphenyl exposures were higher in NAT1*10 than in NAT1*4 transfected cells. Ribonuclease protection assays showed no difference between NAT1*4 and NAT1*10. However, protection of one probe by NAT1*10B was not observed with NAT1*4 or NAT1*10, suggesting additional mechanisms that regulate NAT1*10B. The higher mutants in cells transfected with NAT1*10 and NAT1*10B are consistent with an increased cancer risk for individuals possessing NAT1*10 haplotypes.

Journal Article.  5767 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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