Journal of Medical Genetics

MAHER, EAMONN R

doi: 10.1136/jmg.36.10.729pmid: 10539977

Hacia, Joseph G; Collins, Francis S

doi: 10.1136/jmg.36.10.730pmid: 10528850

The development of inexpensive high throughput methods to identify individual DNA sequence differences is important to the future growth of medical genetics. This has become increasingly apparent as epidemiologists, pathologists, and clinical geneticists focus more attention on the molecular basis of complex multifactorial diseases. Such undertakings will rely upon genetic maps based upon newly discovered, common, single nucleotide polymorphisms. Furthermore, candidate gene approaches used in identifying disease associated genes necessitate screening large sequence blocks for changes tracking with the disease state. Even after such genes are isolated, large scale mutational analyses will often be needed for risk assessment studies to define the likely medical consequences of carrying a mutated gene. This review concentrates on the use of oligonucleotide arrays for hybridisation based comparative sequence analysis. Technological advances within the past decade have made it possible to apply this technology to many different aspects of medical genetics. These applications range from the detection and scoring of single nucleotide polymorphisms to mutational analysis of large genes. Although we discuss published scientific reports, unpublished work from the private sector1 2 could also significantly affect the future of this technology.

Burn, John

doi: 10.1136/jmg.36.10.737pmid: 10528851

Narain, Yolanda; Wyttenbach, Andreas; Rankin, Julia; Furlong, Robert A; Rubinsztein, David C

doi: 10.1136/jmg.36.10.739pmid: 10528852

Huntington’s disease (HD) is thought to show true dominance, since subjects with two mutant alleles have been reported to have similar ages at onset of disease compared to heterozygous sibs. We have investigated this phenomenon using a cell culture model. Protein aggregate formation was used as an indicator for pathology, as intraneuronal huntingtin inclusions are associated with pathology in vitro and in vivo. We showed that cytoplasmic and nuclear aggregates are formed by constructs comprising part of exon 1 of huntingtin with 41, 51, 66, or 72 CAG repeats, in a rate that correlates with repeat number. No inclusions were seen with 21 CAG repeat constructs. Mutant and wild type huntingtin fragments can be sequestered into inclusions seeded by a mutant huntingtin. Wild type huntingtin did not enhance or interfere with protein aggregation. The rate of protein aggregation was dose dependent for all mutant constructs tested. These experiments suggested a model for the dominance observed in HD; the decrease in the age at onset of a mutant homozygote may be small compared to the variance in the age at onset for that specific repeat number in heterozygotes. Our experiments also provide a model, which may explain the different repeat size ranges seen in patients and healthy controls for the different polyglutamine diseases.

van Orsouw, Nathalie J; Dhanda, Rahul K; Elhaji, Youssef; Narod, Steven A; Li, Frederick P; Eng, Charis; Vijg, Jan

doi: 10.1136/jmg.36.10.747pmid: 10528853

The hereditary breast and ovarian cancer syndrome is associated with a high frequency of BRCA1mutations. However, the widespread use ofBRCA1 testing has been limited to date by three principal concerns: the fear of loss of health and life insurance, the uncertain clinical value of a positive test result, and the current lack of an inexpensive and sensitive screening test forBRCA1 mutations. We have developed an inexpensive system for gene mutational scanning, based on a combination of extensive multiplex PCR amplification and two dimensional electrophoresis. The efficiency of this system, as a screening test forBRCA1 mutations, was evaluated in a panel of 60 samples from high risk women, 14 of which contained a previously identified mutation in BRCA1. All 14 mutations were identified, as well as an additional five that had previously escaped detection. In addition to the 19 mutations, a total of 15 different polymorphic variants were scored, most of which were recurring. All were confirmed by nucleotide sequencing. The cost of screening per sample was calculated to be approximately US$70 for the manual technique used in this study, and may be reduced to approximately US$10 with the introduction of commercially available PCR robotics and fluorescent imaging. Implementation of this method of mutation screening in the research and clinical setting should permit rapid accrual of quantitative data on genotype-phenotype associations for the evaluation of diagnostic testing.

Villard, Laurent; Briault, Sylvain; Lossi, Anne-Marie; Paringaux, Christine; Belougne, Jérôme; Colleaux, Laurence; Pincus, D R; Woollatt, E; Lespinasse, James; Munnich, Arnold; Moraine, Claude; Fontès, Michel; Gecz, Jozef

Christianson, Arnold L; Stevenson, Roger E; van der Meyden, C H; Pelser, Julie; Theron, Francois W; van Rensburg, Petro L; Chandler, Michael; Schwartz, Charles E

doi: 10.1136/jmg.36.10.759pmid: 10528855

Murray, Anna; Webb, James; Dennis, Nick; Conway, Gerard; Morton, Newton

doi: 10.1136/jmg.36.10.767pmid: 10528856

Genetic causes of premature ovarian failure (POF) include X chromosome deletions and fragile X (FRAXA) premutations. While screening a cohort of women with POF for FRAXA premutations, a more distal trinucleotide repeat, FRAXE, was also tested. We found an unexpected excess of FRAXE alleles with apparently fewer than 11 repeats in the POF group. However, sequence analysis of these alleles showed that the excess was caused by three females who carry cryptic deletions in FMR2, the gene associated with FRAXE. We propose that microdeletions withinFMR2 may be a significant cause of premature ovarian failure, being found in 1.5% of women with the condition, and in only 0.04% of the general female population. The deletions may affect transcription of either FMR2 or an adjacent gene.

Borrego, Salud; Sáez, María Eugenia; Ruiz, Agustín; Gimm, Oliver; López-Alonso, Manuel; Antiñolo, Guillermo; Eng, Charis

doi: 10.1136/jmg.36.10.771pmid: 10528857

Hirschsprung disease (HSCR) is a common genetic disorder presenting with functional intestinal obstruction secondary to enteric aganglionosis. HSCR can be familial or sporadic. Although five putative susceptibility genes have been identified, only germline mutations in the RET proto-oncogene account for a significant minority (up to 50%) of familial HSCR; 3% of sporadic HSCR in a population based series carry RETmutations. From 1998 to February 1999, we prospectively ascertained 64 cases of sporadic HSCR from the western Andalusia region. To determine if polymorphic sequence variants within RETcould act as low penetrance predisposing alleles, we examined allelic frequencies at seven polymorphic loci in this population based series. Whether allele frequencies differed from those in the control population were determined by either chi-squared analysis or Fisher’s exact test. For two sequence variants, A45A (c 135G→A) (exon 2) and L769L (c 2307T→G) (exon 13), the rarer polymorphic allele was over-represented among HSCR cases versus controls (p<0.0006). In contrast, two other polymorphisms, G691S (c 2071C→A) (exon 11) and S904S (c 2712C→G) (exon 15), were under-represented in the HSCR patients compared to controls (p=0.02). Polymorphisms in theRET proto-oncogene appear to predispose to HSCR in a complex, low penetrance fashion and may also modify phenotypic expression.

Manouvrier-Hanu, S; Amiel, J; Jacquot, S; Merienne, K; Moerman, A; Coëslier, A; Labarriere, F; Vallée, L; Croquette, M F; Hanauer, A

doi: 10.1136/jmg.36.10.775pmid: 10528858

An unreported missense mutation of the ribosomal S6 kinase 2 (RSK2) gene has been identified in two male sibs with a mild form of Coffin-Lowry syndrome (CLS) inherited from their healthy mother. They exhibit transient severe hypotonia, macrocephaly, delay in closure of the fontanelles, normal gait, and mild mental retardation, associated in the first sib with transient autistic behaviour. Some dysmorphic features of CLS (in particular forearm fullness and tapering fingers) and many atypical findings (some of which were reminiscent of FG syndrome) were observed as well. The moderate phenotypic expression of this mutation extends the CLS phenotype to include less severe mental retardation and minor, hitherto unreported signs. The missense mutation identified may be less deleterious than those previously described. As this mutation occurs in a protein domain with no predicted function, it could be responsible for a conformational change affecting the protein catalytic function, since a non-polar amino acid is replaced by a charged residue.