Alan C. Gough

Debrisoquine hydroxylase gene polymorphism and susceptibility to Parkinson's disease

The pathogenesis of Parkinsons disease may be influenced by genetic and environmental factors. Cytochrome P450 mono-oxygenases help to protect against toxic environmental compounds and individual variations in cytochrome P450 expression might, therefore, influence susceptibility to environmentally linked diseases. The frequency of mutant CYP2D6 alleles was studied in 229 patients with Parkinsons disease and 720 controls. Individuals with a metabolic defect in the cytochrome P450 CYP2D6-debrisoquine hydroxylase gene with the poor metaboliser phenotype had a 2.54-fold (95% Cl 1.51-4.28) increased risk of Parkinsons disease. Determination of CYP2D6 phenotype and genotype may help to identify those at greatest risk of Parkinsons disease and may also help to identify the environmental or metabolic agents involved in the pathogenesis of this disease.

A novel human cytochrome P450 gene (P450IIB): chromosomal localization and evidence for alternative splicing.

We have isolated from a single human liver cDNA library two clones which are highly homologous (78% over the coding region) to the major phenobarbital-inducible P450 from rat (P450IIB1). This is the first direct demonstration of the presence of the P450IIB gene subfamily in humans. This subfamily is much less extensive than the rodent homologues, but does appear to contain at least two genes. Of the cDNA clones isolated one is apparently normally spliced, whereas the other lacks exon 8 and retains all or part of intron 5. Both clones contain transcribed Alu sequences. The human P450IIB gene has been located to chromosome 19q12----19q13.2 using a probe derived from intron 5, and is close to the CYP 2A locus encoding cytochrome P450IIA2. Restriction fragment length polymorphisms have been found with the enzymes BamHI and MspI which will enable linkage to be determined between these two loci.

Restriction fragment length polymorphism analysis and assignment of the metalloproteinases stromelysin and collagenase to the long arm of chromosome 11

Collagenase and stromelysin are two metalloproteinases produced mainly by connective tissue cells and involved in the breakdown of the extracellular matrix. cDNA clones for both of these genes have been isolated and sequencing has shown them to be closely related. The collagenase and stromeylsin cDNA clones have been used to assign these genes to the long arm of chromosome 11 in the regions 11q21-22.1 and 11q22.2-22.3, respectively. This has been achieved using somatic cell hybrids and in situ hybridization. In addition a Taq1 restriction fragment length polymorphism has been demonstrated using the stromelysin cDNA.

Localization of the gene for human simple epithelial keratin 18 to chromosome 12 using polymerase chain reaction.

Many human genes encoding keratin intermediate filament proteins are clustered on chromosomes 17 (the type I genes) and 12 (the type II genes). Some have not yet been localized, notably the genes for the primary embryonic keratins 8 and 18, normally expressed in simple epithelia: this is because the numerous pseudogenes for these keratins have made it difficult to identify the true functional gene in each case. Through the use of human-specific primers from within introns of the published gene sequence for human type I keratin 18, human genomic DNA has been specifically amplified using the polymerase chain reaction. A single reaction product was obtained. DNA from a characterized series of mouse-human somatic cell hybrid lines was tested for the presence of sequences able to initiate the chain reaction from these primers, and the presence or absence of this genomic DNA PCR product allowed us to assign a gene for human keratin 18 to chromosome 12 unambiguously. This differs from the location of other human type I keratins on chromosome 17 and may indicate the early divergence of the genes for stratifying cell keratins from that of simple, or embryonic, keratin 18.

The human cytochrome P450 CYP3 locus: assignment to chromosome 7q22-qter

SummaryThe cytochrome P-450s are a large multigene family of enzymes involved in the metabolism and detoxification of drugs and chemicals. Using a full length cDNA clone for the human nifedipine oxidase gene (CYP3) and a panel of human-rodent somatic cell hybrids, we have assigned this gene family to chromosome 7q22-qter. A search for RFLPs using this probe yielded no results.

Cloning of the mouse homologue of the yeast cell cycle control gene cdc2

The cdc2 cell cycle control genes of yeast and man encode 34 kDa protein kinases. Using the human CDC2 gene as a DNA probe we have isolated cDNA clones corresponding to the mouse cdc2 gene. Sequencing of the mouse clones show 96% identity at the amino acid level to the human CDC2 gene and detects a 2 kb RNA product indistinguishable to that seen in human cells. The mouse gene should be useful in studying the functional control of the cell cycle in mouse cells.

Evolutionary conservation of the human homologue of the yeast cell cycle control gene cdc2 and assignment of Cd2 to chromosome 10.

SummaryThe human homologue of the fission yeast Schizosaccharomyces pombe cell cycle control gene cdc2 has been assigned to chromosome 10. DNA hybridization reveals that this gene is highly conserved in vertebrates. The human CDC2 gene probe detects a simple two-allele polymorphism in Taq1-digested DNA.

Chromosome assignment of the human glutathione S-transferase μ3 gene (GSTM3) to chromosome 1 by gene specific polymerase chain reaction

Oligonucleotide primers specific for exons 4 and 5 sequences were used to amplify a unique 199-bp fragment in the human GSTM3 gene. Using DNA from a panel of somatic cell hybrids we assigned the GSTM3 locus to chromosome 1p.