© 1994 Oxford University Press
research-article |
Biochemical and genetic analysis of two acetyltransferases from hamster tissues that can metabolize aromatic amine derivatives
Department of Chemical Carcinogenesis, Michigan Cancer Foundation Detroit, MI 48201, USA
1To whom correspondence should be addressed
Cytosolic activities in liver, stomach, small intestine, colon, lung, kidney, brain and spleen of hamster have been shown to be capable of N–acetylation, O–acetylation and N,O–acetyltransfer utilizing aromatic amine derivatives as substrates. These activities, which have been implicated in the metabolic activation and carcinogenicity of these compounds, were highest in the liver and small intestine. N–Acetylation could be demonstrated with either acetyl CoA or N–hydroxy–N–acetylaminoarenes serving as acetyl donors. Each of these tissues gave two immunoreactive bands ({tilde}29 and 31 kDa) on Western blots using anti–rat AT monoclonal antibodies for detection. Single copies of two distinct acetyltransferase genes, designated AT1 and AT2, were detected in hamster DNA by Southern blot analysis using gene–specific hybridization probes for the 3' end of the AT coding regions. A third gene with >80% sequence similarity to codons 118–158 of AT2 was also detected. Sequence analysis of the two AT genes showed that both had intronless, 0.87 kb coding regions. One was identical with the ATI reported by Abu-Zeid et al. (Abu-Zeid,M., Nagata,K., Miyata,M., Ozawa,S., Fukuhara,M. and Yamazoe,Y., 1991, An arylamine acetyltransferase (AT-1) from Syrian golden hamster liver: cloning, complete nucleotide sequence, and expression in mammalian cells. MoL Carcinogen., 4, 81-88). The second, AT2 (GenBank accession number L24912
[GenBank]
), coded for a 290 amino acid sequence that was 79% homologous with hamster ATI and had a calculated molecular weight of 33.8 kDa, a theoretical pi of 5.96 and the three cysteines that have been conserved in all known vertebrate ATs at positions 44, 68 and 223. Northern blot hybridization using gene-specific AT probes detected mRNAs for both ATI and AT2 in each of the eight tissues analyzed. Two ATI transcripts, 
1.7 and 23 kb, were found in approximately equal ratios in all eight tissues. Three transcripts for AT2, 1.9, 2.1 and 2.4 kb, were present in approximately equal ratios in the brain, small intestine, lung and colon. The liver, stomach, kidney and spleen had primarily the smaller two AT2 mRNAs. The overall abundance of AT mRNAs was highest in liver, small intestine and colon. The coding sequences of all AT mRNAs analyzed were identical to their corresponding genes, although the lengths of both the 5' and 3' untranslated transcript regions varied. Collectively, these data indicate that two closely-related acetyltransferases differing in substrate specificity are expressed in diverse hamster tissues, but with marked differences in overall activity level that may help govern tissue-specific metabolic activation of aromatic amines.