Arylamine N-acetyltransferases are xenobiotic metabolising enzymes, involved in metabolic pathways leading to either detoxification or bio-activation of drugs and carcinogens. In humans, polymorphisms of the two functional NAT genes (NAT1 and NAT2} have been implicated to cancer susceptibility. The genomic region of the NAT1 loci, 8p22, is often deleted in tumours and is believed to contain tumour suppressor genes. Two previously developed cosmid probes, carrying either NAT1 or NAT2, were tested as probes for FISH analysis using nuclei from healthy lymphocytes and the RT112 urothelial carcinoma cell line (Chapter 3). FISH analysis of cells obtained from barbotage samples of bladder cancer patients was performed and deletion of the NAT genomic region was a common observation (Chapter 6). To allow refined mapping of the NAT region on 8p22, a series of PAC clones carrying the NAT loci were isolated by PCR screening of a genomic library (Chapter 3). Furthermore, the NAT1 and NAT2 isoenzymes were studied in healthy intestinal and mammary tissue, as a first step towards understanding their potential involvement in these two types of cancer (Chapter 6). The mouse has been extensively used as a model for studying NAT. Three functional Nat genes are present in the mouse genome. A fine restriction map was generated for the Nat1 and Nat2 genomic region in mice, using Nat-positive genomic DNA plasmid clones previously isolated from the fast acetylator Balb/c and 129/Ola strains. The Nat1 and Nat2 genes were mapped 9.4kb apart and no polymorphisms were detected between the two strains for the restriction enzymes used for mapping (Chapter 4). The 129/Ola restriction map has since been the basis for the production of a construct for Nat2 knockout and transgenic mouse strains. PAC clones positive for Nat were isolated from a mouse genomic library and used as FISH probes to map the three murine Nat genes on chromosome 8, band B3.1-3.3. One PAC clone contained all three Nat loci, establishing co-localisation of Nat3 with the other two Nat genes in mice. The minimum distance of Nat3 from Nat1 and Nat2 was estimated to be 22kb, while the three loci are within 130kb. YAC clones carrying the Nat genes were also isolated to facilitate physical mapping of the Nat cluster in mice (Chapter 5). This will allow integration of cytogenetic, physical and previous genetic data for comparative studies.</p
