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Background: Dyslipidaemia is a well known risk factor for cardiovascular disease (CVD). Lipid metabolism is affected by a range of genes and proteins. This study investigated whether some of these genes are associated with measures of subclinical CVD. Methods: Polymorphisms of paraoxonase 1 and 2, cholesteryl ester transfer protein, hepatic lipase, and lipoprotein lipase were tested for associations with measures of subclinical CVD including carotid intima-media thickness measured by B-mode ultrasound and carotid and coronary arterial calcification measured by computed tomography. Analysis was performed in 620 European American participants in the Diabetes Heart Study, 83% of whom had type 2 diabetes mellitus. Associations of genotypes with subclinical CVD were tested by computing a series of generalised estimating equations. Results: The Q192R variant of paraoxonase 1 and rs285 of lipoprotein lipase were associated with carotid artery calcium (p values = 0.002 and 0.005, respectively). Paraoxonase 2 S311C was associated with coronary artery calcium (p value = 0.037). Conclusions: There is evidence for modest, but significant, association of multiple single nucleotide polymorphisms in lipid genes with measures of subclinical CVD.

Background: CHARGE syndrome has an estimated prevalence of 1/10 000. Most cases are sporadic which led to hypotheses of a non-genetic aetiology. However, there was also evidence for a genetic cause with reports of multiplex families with presumed autosomal dominant, possible autosomal recessive inheritance and concordant twin pairs. We identified a monozygotic twin pair with CHARGE syndrome and a de novo balanced chromosome rearrangement t(8;13)(q11.2;q22). Methods: Fluorescence in situ hybridisation was performed with BAC and PAC probes to characterise the translocation breakpoints. The breakpoint on chromosome 8 was further refined using 10 kb probes we designed and produced using sequence data for clone RP11 33I11, the Primer3 website, and a long range PCR kit. Results: BAC and PAC probe hybridisation redefined the breakpoints to 8q12.2 and 13q31.1. Probe RP11 33I11 spanned the breakpoint on chromosome 8. Using our 10 kb probes we demonstrated that the chromodomain gene CHD7 was disrupted by the translocation between exons 3 and 8. Discussion: Identifying that the translocation breakpoint in our patients occurred between exons 3 and 8 of CHD7 suggests that disruption of this gene is the cause of CHARGE syndrome in the twins and independently confirms the role of CHD7 in CHARGE syndrome.

Background: Cutis laxa is an acquired or inherited condition characterized by redundant, pendulous and inelastic skin. Autosomal dominant cutis laxa has been described as a benign disease with minor systemic involvement. Objective: To report a family with autosomal dominant cutis laxa and a young girl with sporadic cutis laxa, both with variable expression of an aortic aneurysmal phenotype ranging from mild dilatation to severe aneurysm or aortic rupture. Methods and results: Histological evaluation of aortic aneurysmal specimens indicated classical hallmarks of medial degeneration, paucity of elastic fibres, and an absence of inflammatory or atherosclerotic lesions. Electron microscopy showed extracellular elastin deposits lacking microfibrillar elements. Direct sequencing of genomic amplimers detected defects in exon 30 of the elastin gene in affected individuals, but did not in 121 normal controls. The expression of mutant elastin mRNA forms was demonstrated by reverse transcriptase polymerase chain reaction analysis of cutis laxa fibroblasts. These mRNAs coded for multiple mutant tropoelastins, including C-terminally truncated and extended forms as well as for molecules lacking the constitutive exon 30. Conclusions: ELN mutations may cause severe aortic disease in patients with cutis laxa. Thus regular cardiac monitoring is necessary in this disease to avert fatal aortic rupture.

Background: In recent years, non-syndromic idiopathic cardiomyopathies have increasingly been characterised as autosomal dominant conditions caused by single gene mutations. Loci have been identified for hypertrophic and dilated cardiomyopathy, and in some cases the same loci are associated with restrictive cardiomyopathy (RCM). In a kindred with RCM that we previously reported, we ruled out the known cardiomyopathy loci and other candidate genes by linkage analysis and mutation screening. Methods and Results: Here we report a genome-wide analysis in this family that has resulted in linkage to a region on chromosome 10. Conclusions: There are no genes in the interval that are known to cause idiopathic cardiomyopathy, and thus this linkage represents localisation of a new RCM locus.

Background: Autosomal dominant early onset Alzheimer's disease (ADEOAD) is genetically heterogeneous. Mutations of the amyloid precursor protein (APP), presenilin 1 (PSEN1), and presenilin 2 (PSEN2) genes have been identified. Objective: To further clarify the respective contribution of these genes to ADEOAD. Methods: 31 novel families were investigated. They were ascertained using stringent criteria (the occurrence of probable or definite cases of Alzheimer's disease with onset before 60 years of age in three generations). All cases fulfilled the NINCDS-ADRDA criteria for probable or definite Alzheimer's disease. The entire coding regions of PSEN1 and PSEN2 genes and exons 16 and 17 of APP gene were sequenced from genomic DNA Results: PSEN1 mutations, including eight previously unreported mutations, were detected in 24 of the 31 families, and APP mutations were found in five families. In this sample, the mean ages of disease onset in PSEN1 and APP mutation carriers were 41.7 and 51.2 years, respectively. Conclusions: Combining these data with previously published data, yielding 65 ADEOAD families, 66% of the cases were attributable to PSEN1 mutations and 16% to APP mutations, while 18% remained unexplained.

OBJECTIVE: To develop and compare two new technologies for diagnosing a contiguous gene syndrome, the Williams-Beuren syndrome (WBS). METHODS: The first proposed method, named paralogous sequence quantification (PSQ), is based on the use of paralogous sequences located on different chromosomes and quantification of specific mismatches present at these loci using pyrosequencing technology. The second exploits quantitative real time polymerase chain reaction (QPCR) to assess the relative quantity of an analysed locus. RESULTS: A correct and unambiguous diagnosis was obtained for 100% of the analysed samples with either technique (n = 165 and n = 155, respectively). These methods allowed the identification of two patients with atypical deletions in a cohort of 182 WBS patients. Both patients presented with mild facial anomalies, mild mental retardation with impaired visuospatial cognition, supravalvar aortic stenosis, and normal growth indices. These observations are consistent with the involvement of GTF2IRD1 or GTF2I in some of the WBS facial features. CONCLUSIONS: Both PSQ and QPCR are robust, easy to interpret, and simple to set up. They represent a competitive alternative for the diagnosis of segmental aneuploidies in clinical laboratories. They have advantages over fluorescence in situ hybridisation or microsatellites/SNP genotyping for detecting short segmental aneuploidies as the former is costly and labour intensive while the latter depends on the informativeness of the polymorphisms.

Chromosome 22q loss of heterozygosity (LOH) is the most common allelic loss in benign meningioma and is thought to be the earliest initiating event in meningioma formation. We used published data and logistic regression to evaluate the association of 22q LOH with age at diagnosis in 318 transitional, fibroblastic, and meningothelial meningiomas. After adjustment for anatomical location, the odds ratio of 22q LOH per year of age was >1 in each histological type of meningioma, and was significantly >1 in transitional and fibroblastic meningioma. This finding is compatible with involvement of the neurofibromatosis 2 tumour suppressor gene, NF2, on chromosome 22q in the high incidence of benign meningioma in the elderly.

Background: Defects of the mitochondrial genome are recognised as common causes of genetic disease. Sequencing of large portions or even the entire mitochondrial genome is routine in many laboratories for the investigation of mitochondrial disease. However, establishing whether a detected sequence change is polymorphic or pathogenic is still a major difficulty because of its highly polymorphic nature. This has major implications for the patient and the family. Objective: To describe a scoring system for determining the likelihood that a given sequence variant in one of the seven mitochondrially encoded complex I ( MTND ) genes is truly pathogenic. Results: The scoring system was applied to 50 reported MTND mutations. Using this system, 21 of the mutations analysed fell into the group of neutral sequence variants, 10 were classified as possibly pathogenic, three as probably pathogenic, and 16 as almost certainly pathogenic. Conclusions: The proposed scoring system should advance the interpretation of sequence variants and ensure that candidate pathogenic mutations are rigorously investigated.

Background: A myocyte enhancer factor 2A (MEF2A) mutation that segregated with coronary artery disease/myocardial infarction (CAD/MI) in a large family has recently been described. Missense mutations in sporadic coronary artery disease patients were also reported. These data suggest that mutations in exons 7 and 11 of MEF2A cause CAD/MI, though the association was refuted by another study. Objective: To analyse the genetic variation of exons 7 and 11 in a large cohort of Spanish CAD/MI patients and controls. Methods and results: A rare polymorphism, P279L, was detected both in patients and controls. Carriers of the 279Leu allele had a threefold risk of suffering CAD/MI compared with controls (p = 0.009; odds ratio = 3.06 (95% confidence interval, 1.17 to 8.06)). In the controls the allele was found only in those under 50 years of age. Exon 11 showed a high degree of heterogeneity caused by a polyglutamine (CAG)n polymorphism, but no significant differences in genotype or allelic frequencies were found. Conclusions: The 279Leu allele appears to be a genetic risk factor for CAD/MI in the population studied. This effect could be the result of a reduced transcriptional activity on MEF2A with 279Leu.

Background: Most non-syndromic congenital heart defects (CHD) are caused by a complex interaction between maternal lifestyle factors, environmental exposures, and maternal and fetal genetic variants. Maternal periconceptional intake of folic acid containing vitamin supplements is reported to decrease the risk of CHD. The 677C→T and 1298A→C polymorphisms in the methylenetetrahydrofolate reductase (MTHFR) gene decrease enzyme activity. Objective: To examine the relation between CHD and maternal and fetal MTHFR polymorphisms Methods: 375 nuclear families were studied. The transmission/disequilibrium test was used to test for transmission distortion in complete triads. A log-linear approach was used to test for associations between CHD and maternal and offspring polymorphisms, and to estimate independently the contributions of maternal and fetal variants to relative risks. Haplotype frequencies were estimated and a haplotype transmission disequilibrium test carried out. Results: The 1298C allele was transmitted less often than expected (p = 0.0013). There was no distortion in the transmission of the 677T allele, neither was there evidence of a parent of origin effect in the transmission of either of the single nucleotide polymorphisms. The 677C–1298C haplotype was also transmitted less often than expected (p = 0.0020). The relative risk associated with inheriting one copy of the 1298C allele was 0.64 (95% confidence interval, 0.48 to 0.87) and the that associated with inheriting two copies of the 1298C allele, 0.38 (0.21 to 0.70). Conclusions: The apparent protective effect of the MTHFR 1298C allele against CHD could have several explanations and further study is needed.

Background: Primary ciliary dyskinesia (PCD) is a severe inherited disorder characterised by chronic respiratory disease, male infertility, and, in ∼50% of affected individuals, a left-right asymmetry defect called situs inversus. PCD is caused by defects in substructures of the ciliary and flagellar axoneme, most commonly loss of the outer dynein arms. Although PCD is believed to involve mutations in many genes, only three have been identified. Methods: To facilitate discovery of new PCD genes, we have used database searching and analysis to systematically identify the human homologues of proteins associated with the Chlamydomonas reinhardtii outer dynein arm, the best characterised outer arm of any species. Results: We find that 12 out of 14 known Chlamydomonas outer arm subunits have one or more likely orthologues in humans. The results predict a total of 24 human genes likely to encode outer dynein arm subunits and associated proteins possibly necessary for outer arm assembly, plus 12 additional closely related human genes likely to encode inner dynein arm subunits. Conclusion: These genes, which have been located on the human chromosomes for easy comparison with known or suspected PCD loci, are excellent candidates for screening for disease-causing mutations in PCD patients with outer and/or inner dynein arm defects.

Y chromosome microdeletions are the most frequent genetic cause of severe oligozoospermia ( 99.9%) nucleotide identity, organised in massive palindromes. These repeated sequences may undergo genetic exchange through gene conversion—that is, non-reciprocal transfer of sequence information occurring between duplicated sequences within the chromosome, a process that could account for the >99.9% nucleotide identity between the arms of a palindrome. Although this mechanism may serve to preserve Y chromosome genes from the gradual accumulation of deleterious mutations and thus prolong their genetic fitness,6 this peculiar organisation also provides the structural basis for deletions and rearrangements. The classical AZFc deletion, which removes 3.5 Mb between the b2/b4 amplicons, is the most frequent type of deletion. Taking into consideration the Y chromosome structure and the suggested deletion mechanism, a number of other possible partial deletions have been proposed in both the AZFb and AZFc regions.7,8 The frequency and the pathological significance of these partial deletions is not yet clear, although recently a partial deletion termed gr/gr has been described specifically in infertile men with varying degrees of spermatogenic failure.9,10 This deletion removes half the AZFc gene content, including two copies of the major AZFc …

Background: A 3 bp deletion located at the 5′ end of exon 3 of MLH1 , resulting in deletion of exon 3 from RNA, was recently identified. Hypothesis: That this mutation disrupts an exon splicing enhancer (ESE) because it occurs in a purine-rich sequence previously identified as an ESE in other genes, and ESEs are often found in exons with splice signals that deviate from the consensus signals, as does the 3′ splice signal in exon 3 of MLH1 . Design: The 3 bp deletion and several other mutations were created by polymerase chain reaction mutagenesis and tested using an in vitro splicing assay. Both mutant and wild type exon 3 sequences were cloned into an exon trapping vector and transiently expressed in Cos-1 cells. Results: Analysis of the RNA indicates that the 3 bp deletion c.213_215delAGA (gi:28559089, NM_000249.2), a silent mutation c.216T→C, a missense mutation c.214G→C, and a nonsense mutation c.214G→T all cause varying degrees of exon skipping, suggesting the presence of an ESE at the 5′ end of exon 3. These mutations are situated in a GAAGAT sequence 3 bp downstream from the start of exon 3. Conclusions: The results of the splicing assay suggest that inclusion of exon 3 in the mRNA is ESE dependent. The exon 3 ESE is not recognised by all available motif scoring matrices, highlighting the importance of RNA analysis in the detection of ESE disrupting mutations.

The function of BRCA1 is complex and includes roles in DNA damage repair, cell cycle control, regulation of transcription, and X chromosome inactivation.1,2 TSIX is thought to control X chromosome inactivation by blocking the accumulation of XIST on the active X chromosome.3 BRCA1 co-localises with XIST inactive X chromosomes (Xi) and stabilises Xi. Because of this interaction, the loss of BRCA1 is associated with altered Xi chromatin structure and increased expression of silenced Xi genes.1,2 Mouse models have suggested a link between aberrant X chromosome inactivation and sex ratio skewing, with a bias towards male births when TSIX activity is abolished.3 In addition, non-random X chromosome inactivation has been seen in BRCA1 mutation carriers with ovarian cancer.4 With these data as background, a skewed sex ratio in offspring of BRCA1 mutation carriers was reported,5 with a bias towards females, compared with offspring of women with BRCA2 mutations and those without mutations in either gene. Others have not found a difference in the sex ratios of offspring of BRCA1 and BRCA2 mutation carriers.6–8 Here, we analysed the sex ratio of offspring in a large cohort of BRCA1 and BRCA2 mutation carriers, as well as in HBOC families who tested negative for BRCA1 and BRCA2 mutations or were not tested. ### Database We included data from 1276 families evaluated between 1992 and 1994 (University of Michigan) and between 1994 and 2003 (University of Pennsylvania) in breast cancer risk evaluation clinics and considered to be at increased risk for having a breast cancer susceptibility gene mutation. Individuals were either self referred or physician referred for assessment of inherited susceptibility to breast or ovarian cancer. All individuals providing data signed their informed consent before genetic testing and entry into our database, which includes detailed pedigree information. …

ypoparathyroidism is a heterogeneous group of disorders with both acquired and inherited causes, each presenting clinically with hypocalcaemia. Familial cases of hypoparathyroidism may be due to an isolated defect of the parathyroid glands or be a component of a syndrome disorder, examples of which include DiGeorge, hypoparathyroidism-retardation-dysmorphism, and KennyCaffey syndrome. 1 Familial isolated hypoparathyroidism (FIH) is characterised by hypocalcaemia and hyperphosphataemia and may be due to an inherited deficiency or the abnormal activity of parathyroid hormone (PTH). FIH is heterogeneous with X linked, autosomal dominant, and autosomal recessive modes of inheritance reported. 2 Mutations in the PTH gene on chromosome 11p have been described in both autosomal dominant and autosomal recessive forms of the disorder. 34 Mature PTH is generated by cleavage of the signal peptide from the prepro-PTH, followed by successive proteolysis of pro-PTH within the Golgi apparatus. 5 A requirement for normal PTH processing is emphasised by the detection of FIH disease causing mutations, which include two missense amino acid substitutions within the signal region of the prepro-PTH gene, leading to inefficient cleavage by signal peptidases. 35

Retinitis pigmentosa (RP) is the most prevalent hereditary retinal degenerative disease. To date, approximately 40 loci and mutations in more than 25 genes have been identified as the cause of various types of RP.1 The gene for human oxygen regulated photoreceptor protein ( RP1 ) encodes a protein of 2156 amino acids that is localised in the connecting cilia of both rod and cone photoreceptors.2 The RP1 protein is required for the morphogenesis of the outer segments of the photoreceptor cells.3,4 Several laboratories have found mutations in the RP1 gene to be the cause of autosomal dominant retinitis pigmentosa (adRP).5–7 However, to our knowledge, association of the RP1 gene with recessive RP has never been reported. The aim of the present study was to map the disease locus for three consanguineous Pakistani families suffering from RP. We present mapping of these autosomal recessive RP families to the 8q11 locus. The results show, for the first time, that in these families a form of arRP is caused by homozygous mutations of the RP1 gene. Although these mutations were found in the parents and some of the siblings who had normal vision (carriers) in a heterozygous state, they were not found in any of a panel of 100 normal controls. We studied three consanguineous Pakistani families (442RP, 452RP, and 336RP) suffering from autosomal recessive RP (fig 1). The patients had night blindness since early childhood and progressive deterioration of vision with age. All the patients were completely blind by the age of 12–15 years in the case of the 442RP and 452RP families and 17–18 years in the 336RP family. Fundoscopic examination and electroretinographic (ERG) analyses were carried out on all the patients, their heterozygous parents and siblings, and unaffected normal siblings. Figure 1 Pedigrees of autosomal recessive RP families …

Juvenile myoclonic epilepsy (JME; OMIM 606904) is a subtype of common idiopathic generalised epilepsy (IGE) and affects up to 26% of all individuals with IGE.1–3 JME is characterised by the onset in adolescence of bilateral myoclonic jerks usually affecting the upper limbs.1,4 Patients also have generalised tonic-clonic seizures and about one third experience absence seizures. Genetic factors are known to play an important role in the etiology of JME.3,4 While identification of genes underlying predisposition to JME has been relatively slow due to clinical and genetic heterogeneity,5,6 progress made so far in the isolation and characterisation of genes associated with other monogenic subtypes of IGE, provides evidence that most idiopathic epilepsy syndromes are caused by mutations in genes encoding ion channels.7 The implications of these findings in monogenic subtypes of the disorder for the complex polygenic subset are now being increasingly appreciated.8 Two types of voltage gated potassium channels have been associated with seizure disorders, the KCNQ channels and the Kv channels. Loss of function mutations for the potassium channels KCNQ2 and KCNQ3 have been identified in families with a rare autosomal dominant subtype of IGE called benign familial neonatal convulsions (BFNC).9,10 Allelic association of JME with KCNQ3 has been suggested in a South Indian cohort of JME patients.11 Mutation in human KCN1A predisposes to episodic ataxia and partial epilepsy.12 These findings emphasise the importance of potassium channels in controlling neuronal excitability and thus make potassium channel genes potentially interesting candidates for idiopathic epilepsy syndromes. The calcium activated potassium channels are an interesting class of potassium channels that regulate neuronal excitability.13,14 These are gated by intracellular calcium ions and their activity is responsible in part for the afterpolarisation that follows a single action …

Introduction: Clinical variability associated with the common 22q11.2 microdeletion is well known, and has led to a broad application of FISH diagnostics with probes for loci TUPLE1 or D22S75 (N25), although, rarely reported atypical deletions associated with the same phenotypic spectrum would not be discovered by these probes. As most types of 22q11.2 deletions occur between low copy repeats within the region (LCR22), we assumed that atypical deletions should be more common than has been reported. To address this question and the possibility of a deletion size related genotype-phenotype correlation, we systematically assessed the frequency of typical and atypical 22q11.2 deletions in a large cohort of patients. Methods: We used a set of 10 fluorescent in situ hybridisation (FISH) DNA probes, capable of detecting all reported and hypothetical deletions between the LCR22, and analysed 350 patients. Deletion sizes in atypical deletions were established by use of further FISH probes. Frequency of certain atypical deletions was analysed in controls by FISH and quantitative PCR. Results: Patients with conotruncal heart defects (ctCHD) and with typical VCFS phenotype showed the common 3 Mb or nested 1.5 Mb deletions (in 18.5% and 78.6%, respectively), but no atypical deletion, while 5% (3/63) of patients with a mildly suggestive, atypical phenotype showed atypical distal deletions, which were not detected in patients with mental retardation of unknown origin or in healthy controls. Discussion: These statistically significant differences demonstrate that atypical distal 22q11.2 deletions are very uncommon in patients with ctCHDs, while atypical congenital heart defects and mild dysmorphism are recognisable feature of atypical distal deletions. Further phenotype-genotype analysis disclosed association of significant developmental delay with the distal part of the common deletion region, and choanal atresia and atypical CHDs with the adjacent distal deletion region.

2 many companies also offer to deduce ''ancestry''. As the number of markers used in these tests increases, so does the probability of inadvertently diagnosing male infertility through the detec- tion of Y chromosomal deletions. Using commercially typed Y markers, we here report the ascertainment of such deletions in general population samples. METHODS Samples were collected with informed consent and relevant ethical approval from the Leicestershire Research Ethics Committee (ref. 5796) and the Committee for Scientific Investigations in Greenland (ref. 505-16). Deletion analysis was carried out using standard PCR techniques; primer sequences and conditions are given in original references cited for markers in the text below.

Autosomal recessive polycystic kidney disease (ARPKD; MIM 263200) is an important childhood nephropathy, occurring in 1 in 20 000 live births.1 The clinical phenotype is dominated by dilatation of the renal collecting ducts, biliary dysgenesis, and portal tract fibrosis. Affected children often present in utero with enlarged, echogenic kidneys, as well as oligohydramnios secondary to poor urine output. Approximately 30% of affected neonates die shortly after birth as a result of severe pulmonary hypoplasia and secondary respiratory insufficiency. Those who survive the perinatal period express widely variable disease phenotypes with systemic hypertension, renal insufficiency, and portal hypertension due to portal tract fibrosis as the most common clinical features.2 Linkage analysis indicates that mutations in a single locus on chromosome 6p12 are responsible for all typical forms of ARPKD.3,4 Two groups working independently have identified PKHD1 (MIM 606702) as the locus responsible for ARPKD and have demonstrated that this novel gene is among the largest in the human genome, extending over at least 470 kb and including a minimum of 86 exons.5,6 Both PKHD1 and its mouse orthologue ( Pkhd1 ) encode a complex and extensive array of splice variants, with most abundant transcriptional expression in fetal and adult kidney and weaker expression in other tissues including liver and pancreas.5,7 The longest PKHD1 transcript includes 67 exons with an open reading frame (ORF) composed of 66 exons that encode a 4074 amino acid protein, polyductin/fibrocystin.5,6 The full length protein is predicted to have several immunoglobulin-like, plexin, transcription factor (IPT) domains and multiple parallel beta-helix 1 (PbH1) repeats in its approximately 3860 amino acid extracellular amino terminus; a single transmembrane (TM) spanning domain; and a short, cytoplasmic carboxyl terminus with potential phosphorylation sites. Alternatively spliced transcripts are predicted to fall into two broad groups. …

Hirschsprung disease (HSCR) is a complex disorder with traditional germline mutations in RET in up to 30% of familial cases and in 3% of sporadic cases in a population-based series. We have previously demonstrated that an ancestral haplotype at the 5' end of RET (haplotype 0) was strongly associated with a large subset of isolated HSCR cases and that a putative low penetrance susceptibility locus was encompassed within this ancestral haplotype, anchored by exon 2 SNP A45A.To determine the 5' extent of the HSCR-associated ancestral haplotype by defining the linkage disequilibrium breakpoint in search for the low penetrance susceptibility locus.Systematic screening of the region upstream of the anchoring A45A SNP, comprising RET intron 1, exon 1, and promoter in 117 population-based HSCR cases and 100 controls. Dual luciferase assay to determine differential activities between SNP combinations near a transcription start site.New SNP's were found which formed upstream haplotypes, anchored by A45A, in linkage disequilibrium with HSCR (2 = 76.96, p0.00000001). Linkage disequilibrium appeared to break at the -1249C/T SNP. Further, the HSCR-associated genotype (00) was found in60% of HSCR but only 2% of controls. Because only 2 variants, -200AG and -196CA, lie within the promoter region and are in proximity to the transcriptional start site (at -195), we modelled these combinations into constructs for luciferase reporter assay. The HSCR-associated SNP combination showed the lowest activity and the control-associated combination, the highest.Our observations seem to discard the existence of a HSCR-causing mutation as it is conceived in the traditional sense, but strengthen the idea of a specific combination of variants conferring susceptibility to the disease in a low penetrance fashion. The data derived from our functional "in vitro" studies would suggest that the HSCR-associated haplotype 0 may result in a lower level of expression of the RET gene [corrected]

Background: SPG4 encodes spastin, a member of the AAA protein family, and is the major gene responsible for autosomal dominant spastic paraplegia. It accounts for 10–40% of families with pure (or eventually complicated) hereditary spastic paraparesis (HSP). Objective: To assess the frequency of SPG4 mutation in patients with spastic paraplegia but without family histories. Methods: 146 mostly European probands with progressive spastic paraplegia were studied (103 with pure spastic paraplegia and 43 with additional features). Major neurological causes of paraplegia were excluded. None had a family history of paraplegia. DNA was screened by DHPLC for mutations in the 17 coding exons of the SPG4 gene. Sequence variants were characterised by direct sequencing. A panel of 600 control chromosomes was used to rule out polymorphisms. Results: The overall rate of mutations was 12%; 19 different mutations were identified in 18 patients, 13 of which were novel. In one family, where both parents were examined and found to be normal, the mutation was transmitted by the asymptomatic mother, indicating reduced penetrance. The parents of other patients were not available for analysis but were reported to be normal. There was no evidence for de novo mutations. The mutations found in these apparently isolated patients were mostly of the missense type and tended to be associated with a less severe phenotype than previously described in patients with inherited mutations. Conclusions: : The unexpected presence of SPG4 gene mutations in patients with sporadic spastic paraplegia suggests that gene testing should be done in individuals with pure or complicated spastic paraplegia without family histories.

Hereditary motor and sensory neuropathies, commonly referred to as Charcot-Marie-Tooth disease (CMT), are among the most common inherited neurological diseases, with an overall prevalence of about 1–4/10 000.1 Clinically, the hereditary motor and sensory neuropathies are characterised by progressive muscular and sensory loss in the distal extremities with chronic distal weakness, deformation of the feet (pes cavus), and loss of deep tendon reflexes.2 Two main subgroups have been defined on the basis of electrophysiological and histopathological characteristics: the demyelinating form (CMT1) and the axonal form (CMT2). CMT1 can be distinguished from CMT2 by measuring motor nerve conduction velocities in the median nerve: patients affected by CMT1 show reduced velocities (

Genetic susceptibility to the autoimmune B cell destruction that leads to Type 1 diabetes mellitus (T1D) is a complex trait.1 In recent years, many T1D associations have been reported, but only three (major histocompatibility complex, insulin, and cytotoxic T lymphocyte associated protein 4) have been confirmed in several independent studies.2,3 Independent confirmation is essential to eliminate artefacts of publication bias, multiple hypothesis testing, and, in findings of case–control studies, population stratification.4 Bottini et al recently found, by a case–control design in two independent populations, a novel association of T1D with a single nucleotide polymorphism (SNP) that caused a R620W aminoacid substitution (dbSNP rs2476601) in the lymphoid protein tyrosine phosphatase, non-receptor type 22 ( PTPN22 ) gene.5 PTPN22 encodes LYP, a non-receptor tyrosine phosphatase involved in lymphocyte function. This paper leaves two potential questions unanswered. Firstly, results of case–control studies are potentially artefacts of population stratification, no matter how well matched are the two groups. Secondly, although Bottini et al show that the T allele encodes a protein unable to bind to its important Csk partner, and postulate this as a very attractive candidate mechanism for the genetic effect, they did not address the question of the haplotype structure of the locus and the possibility that the association is due to linkage disequilibrium (LD) with another variant. Here we present the results of a study that confirms this association in a design impervious to population stratification, and in a preliminary step towards addressing the second question, we define the LD block that encompasses the PTPN22-R620W SNP and present a computationally generated list of potentially functional SNPs within the block. ### Subjects Genomic DNA was obtained after informed consent from 588 nuclear families with at least one T1D affected child and two parents. The research ethics board of the Montreal Children’s …

Alpha1-antitrypsin deficiency (AATD) is a genetic condition associated with an increased risk of developing chronic obstructive pulmonary disease (COPD) early in life and, to a lesser extent, liver disease.1 Significant advances have been made during the last decades in understanding its epidemiology and it has been recently suggested that AATD is one of the commonest inherited disorders not only in Caucasians but also among other ethnic groups worldwide.2,3 Although AAT is a highly pleomorphic glycoprotein, with approximately 100 variants having being identified,4 two major deficient variants, namely Z and S, account for most cases of AATD, since the vast majority of such individuals carry the PI*ZZ or PI*SZ genotype, coding for approximately 15% and 25%, respectively, of normal AAT plasma levels. The establishment of international registries, including large series of AATD individuals,5,6 has allowed not only better definition of the epidemiology of AATD, but also more precise definition of the associated clinical phenotypes.5,7,8 Nevertheless, there are at least 30 AAT alleles other than the PI*Z and the PI*S alleles which are associated with significantly reduced or absent plasma AAT levels.9 Given the extreme rarity of such variants, often described in the literature as single case reports, little is known about their epidemiology and even less is known about the associated clinical phenotypes. The Italian Registry for Severe AATD was established in 1996 as a result of a nationwide screening programme sponsored by the two major Italian scientific respiratory societies.10 Although Italy is considered a country with a medium-low prevalence of AATD (mean PI*Z gene frequency: 0.0013),11 the programme succeeded in identifying a relatively large cohort of AATD individuals. During the development of the screening program, we noticed that, in addition to the groups of AATD individuals …

Oral clefts are the most common congenital malformations worldwide. Cleft palate can be non-syndromic (MIM 119540) or it can appear as a part of a syndrome or recurrence pattern. Non-syndromic cleft palate and non-syndromic cleft lip with or without cleft palate (CL/P) are considered to be separate entities, on the basis of different embryonic timing and epidemiology. However, in some syndromes, both of these cleft types segregate in the same pedigree, suggesting that they might share a common genetic background. Oral clefts manifest in over 300 different syndromes, and in some of these syndromes the gene defect is already known (Online Mendelian Inheritance in Man database 2004, http://www.ncbi.nlm.nih.gov/Omim/). Identified mutations in cleft syndromes have shown that functionally and structurally very distinct types of genes have an effect on palatogenesis. Recently, a mutation in interferon regulatory factor 6 (IFR6) was found to cause van der Woude syndrome (VWS) (MIM 119300),1 which is one of the most common cleft syndromes. Mutations in very different type of genes can lead to cleft palate in mice. These genes encode growth factors, receptors, transcription regulators, and enzymes for signalling molecule synthesis. Cleft palate, in addition to other congenital malformations, is found in ∼70 knock out mice strains (The Transgenic/Targeted Mutation Database, http://tbase.jax.org/). Other anomalies occur frequently, and therefore no exact model for non-syndromic cleft palate exists. Usually the penetrance is not complete, but Msx-1 knock outs result in 100% cleft palate.2 It has been suggested that, in humans, ∼50% of cases of cleft palate are non-syndromic.3 The etiology and pathogenesis of non-syndromic cleft palate—and also of all other clefts—are poorly understood. Extrinsic factors, such as maternal smoking4 with a particular allele in TGFα locus,5–7 maternal alcohol consumption,8,9 maternal intake of drugs during the first trimester,10 and …

Psoriasis is a chronic, inflammatory, hyperproliferative disease of the skin, scalp, nails, and joints, with a prevalence of up to 2% in Caucasians1,2 but well under 1% in the Mongoloid races of the Far East.3 The disease varies in severity. Some patients display mild disease with isolated scaling erythematous plaques on the elbows or knees, whereas for others most of their cutaneous surface can be affected. At the cellular level, psoriasis is characterised by markedly increased epidermal proliferation and incomplete differentiation, elongation, dilation, and leakiness of the superficial plexus of dermal capillaries, and by a mixed inflammatory and immune cell infiltrate of the epidermis and papillary dermis.1,2 Dermal infiltrates comprised of T cells and macrophages typically appear in early lesions before epidermal changes.4 The therapeutic effect of immunosuppressive agents suggests psoriasis has a primary immune pathogenic basis.5 Susceptibility to the development of psoriasis is likely to have a significant genetic component. Accumulating evidence is consistent with the idea that psoriasis is a multifactorial disorder caused by the concerted action of multiple disease genes in a single individual and triggered by environmental factors.6 Some of these genes control the severity of a variety of diseases, via their regulation of the inflammatory and immune processes (severity genes), whereas others are unique to psoriasis (specific genes). A number of genetic studies have sought to identify the psoriasis susceptibility loci. Associations between psoriasis and human lymphocyte antigen alleles were first described in 1990.7 Subsequently, genome-wide linkage scans have mapped psoriasis to several chromosomal regions including PSORS1 at 6p21,8,9 PSORS2 at 17q,8–10 PSORS3 at 4q,11 PSORS4 at 1q,12 PSORS5 at 3q,13 PSORS6 at 19p,14 and PSORS7 at 1p.15 Recently, the International Psoriasis Genetics Consortium reassessed these candidate …

56 which encodes a protein composed of 2843 amino acids and is formed by 14 small exons, and a large exon 15 that extends over three quarters of the coding sequence. To date, more than 500 different APC germline mutations have been reported in FAP patients (see Human Gene Mutation Database and references therein). Most of the germline mutations reported so far are localised in the 59 half of the gene and lead to premature truncation due to single base substitutions or small insertions/deletions, resulting in nonsense or frameshift mutations and rarely in splice site mutations. In a small number of cases, single base substitu- tions within exonic sequences predicted to result in missense or silent variants lead to aberrant splicing. 78 Most APC

Wilms tumour (WT) is an embryonal tumour of the kidney that occurs in 1 in 10 000 children. Familial clusters are rare and account for only 1–3% of cases. Mutations in WT1 account for a minority of WT families and two autosomal dominant familial WT predisposition genes have been mapped to chromosomes 17q21 and 19q13.1–3 However, a considerable proportion of familial WT pedigrees are not attributable to any of these loci.4 Fanconi anaemia (FA, MIM 227650) is a rare autosomal recessive condition affecting ∼1 in 300 000 children. FA is characterised by variable congenital abnormalities, short stature, bone marrow failure, hypersensitivity to DNA crosslinking agents, and a predisposition to haematological malignancies such as acute myeloid leukaemia in childhood.5 FA is heterogeneous and consists of at least 11 complementation groups, A, B, C, D1, D2, E, F, G, I, J, and L.6–8 Eight FA genes have been cloned and at least six FA proteins, FANCA, FANCC, FANCE, FANCF, FANCG, and FANCL, form a nuclear complex required for monoubiquitination of FANCD2. This modification promotes translocation of FANCD2 to DNA repair foci that also contain BRCA1, BRCA2, and RAD51.9 In 2002, Howlett and colleagues reported biallelic BRCA2 mutations in individuals with Fanconi anaemia D1 (FA-D1). Subsequently, additional FA-D1 and unclassified FA cases were examined and cases with BRCA2 mutations and WT and/or brain tumours were reported.10–13 These data prompted us to investigate BRCA2 in a familial WT pedigree, WILMS2, which includes siblings with both WT and brain tumours.4,14,15 ### Family report WILMS2 was ascertained as part of our research on susceptibility to WT, which is approved by the London Multicentre Research Ethics Committee. The family includes two affected brothers with WT. The elder child first came to attention when his cryptorchidism was corrected at …

The term hereditary spastic paraplegia (HSP) is used to describe a group of clinically and genetically heterogeneous disorders in which the defining clinical feature is progressive spasticity and weakness of the lower limbs. The phenotype is traditionally classified as “pure” when symptoms and signs are generally confined to those of a progressive spastic paraparesis, or “complicated” when associated with additional neurological or other clinical features.1 Inheritance may be autosomal dominant, autosomal recessive, or rarely X linked. Overall autosomal dominant inheritance is most commonly associated with pure forms of the disease, whereas autosomal recessive HSP shows greater phenotypic variability, including several well defined syndromes.2,3 To date nine autosomal recessive HSP loci have been identified and causative mutations found in three genes: SPG7 (paraplegin), SPG20 (spartin), and SPG21 (maspardin). SPG7 encodes paraplegin, a mitochondrial protein, which is a member of the AAA protein superfamily (ATPase associated with diverse cellular activities) and is homologous to a number of yeast mitochondrial metalloproteases.4 SPG7 mutations may result in either pure or complicated HSP phenotypes.4,5 Muscle biopsy analysis of patients with SPG7 mutations may show histological evidence of mitochondrial dysfunction4,6 and recently biochemical studies have shown specific defects in mitochondrial respiratory chain function.5,7 Mutations …

It is generally accepted that multiple factors play a role in gastric carcinogenesis, including diet,1,2 lifestyle,3 infectious agents such as Helicobacter pylori ( H. pylori ),4,5 and genetic factors as well as gene-environment interactions.6,7 Although the continuous decrease in the incidence of gastric cancer in recent decades can be explained by changing lifestyle, diet habits, and reduced H. pylori infection, the fact that some individuals develop gastric cancer while others do not under similar environmental exposures suggests that genetic predisposition plays an important role in the pathogenesis of this disease. In addition, familial clustering of cases of gastric cancer, a higher incidence of the disease among the relatives of patients than among matched controls, and a more common coincidence of the disease in homozygotic than in heterozygotic twins, also supports genetic links with gastric cancer.8,9,10 In contrast with the vast amount of accumulated information on environmental factors, little is known about the genetic factors linked to gastric cancer. Several studies have reported disease susceptible loci for gastric cancer, based on association analysis of candidate genes involved in DNA repair, detoxification, or the immune system.11 Studies on familial clustering of gastric cancer revealed germline mutations in CDH1, ATM, and MET.12–14 In addition, gastric cancer frequently occurs in family members with monogenic diseases such as hereditary non-polyposis colorectal cancer, Li-Fraumeni syndrome, familial adenomatous polyposis, and Peutz-Jeghers syndrome.15–18 However, these polymorphisms and/or mutations cannot explain the overall genetic components of gastric cancer. It is now feasible to screen the entire human genome for genes exerting a major effect on susceptibility to multifactorial diseases. Because gastric cancer, like many complex diseases, exhibits genetic heterogeneity, we carried out a genome-wide linkage study to search for possible susceptibility loci related to gastric …

Bacterial, environmental, population related, and individual host factors are major determinants of the outcome of H pylori infection.1,2 Many bacterial virulence genes—including the pathogenicity island cagA , the s1m1 vacA alleles, babA2 , sabA , and oipA —have been associated with a higher degree of gastric mucosal inflammation, intestinal metaplasia, gastric or duodenal ulcer, gastric adenocarcinoma, and MALToma.1,3–7 H pylori triggers and maintains gastric mucosal inflammation by different mechanisms, which are partly strain dependent and partly strain independent.1 T and B lymphocyte activation and infiltration of the gastric mucosa depend on H pylori antigen processing. The number of infiltrating polymorphonuclear cells varies depending on the virulence of the infecting strain, being much greater when infections are caused by cagA positive strains.3,5,7–9 The inflammatory cells infiltrating H pylori infected gastric mucosa produce a pattern of proinflammatory cytokines.10,11 High mucosal levels of mononuclear cytokines (IL8, IL6, IL1β, tumour necrosis factor α (TNFα), and interferon γ (IFNγ)) and lymphocytic derived cytokines (IL2, IL2R) have been described in H pylori infected patients.10–13 H pylori infection also induces the production of IL12,14–16 a heterodimeric proinflammatory protein that triggers the production of IFNγ and favours the differentiation of T helper 1 (Th1) cells,17,18 which, in H pylori infected mucosa, prevail over Th2 cells.15,16,19 The ability of IL12 to induce Th1 is one of the biological bases of the importance of this cytokine in resisting most bacteria, including H pylori , and also intracellular protozoa and fungal pathogens.18,20,21 Cellular sources of IL12 in response to infections are mainly dendritic cells and phagocytes.16–21 The two subunits of IL12—p35 and p40—are encoded by different genes, named IL12A and IL12B respectively, which are unrelated …

Charcot-Marie-Tooth (CMT) disease is a motor and sensory neuropathy with clinical and genetic heterogeneity. Patients usually present in the first or second decade of life with distal muscle atrophy in the legs, areflexia, foot deformity (mainly pes cavus), and steppage gait. In most cases, hands are also involved as the disease progresses. CMT is the most frequent inherited neuropathy, with a prevalence in Spain of 28 in 100 000.1 Based on electrophysiological studies and histopathologic findings in nerve biopsies, CMT has been subcategorised into two main and distinct neuropathies: (i) demyelinating CMT (CMT1, MIM 118200) associated with reduction in a nerve conduction velocities (NCVs) in all nerves and segmental demyelination and remyelination (“onion bulbs”); and (ii) axonal CMT (CMT2, MIM 118220) associated with normal or almost normal NCVs and loss of myelinated axons. Other phenotypes are associated with motor and sensory nerve involvement: Dejerine-Sottas neuropathy (DSN, MIM 145900) is a severe demyelinating neuropathy with onset in infancy, delayed motor milestones, and NCVs less than 10 m/s; congenital hypomyelinating neuropathy (CHN, MIM 605253) is a dysmyelinating neuropathy characterised by infantile hypotonia, distal muscle weakness, and marked reduction of NCVs; hereditary neuropathy with liability to pressure palsies (HNPP, MIM 162500) is a milder sensory and motor neuropathy with periodic episodes of numbness, muscular weakness, and atrophy.2 Genetic heterogeneity is characteristic of the disease not just because of the large number of genes and loci associated with CMT (currently 21 genes),3–5 but also because the disease may segregate with different Mendelian patterns. The most frequent pattern of inheritance is autosomal dominant, but autosomal recessive and X linked segregation are also observed. The relationship of the type of CMT, demyelinating or axonal, with specific genes is not perfect. For instance, MPZ mutations are usually manifested clinically as an autosomal dominant …

Congenital disorders of glycosylation type Ia or CDG-Ia (MIM 212065) is the most common type of a group of recessive disorders characterised by deficient glycosylation.1 The disease is caused by mutations in the PMM2 gene, coding for a phosphomannomutase (PMM). PMM converts mannose-6-phosphate to mannose-1-phosphate, a precursor of the mannosyl donor in N-and O-glycosylation and the synthesis of GPI anchors. PMM deficiency leads to underglycosylation and altered processing of the N-glycans in serum proteins of CDG-Ia patients.2,3 A plethora of different mutations, mostly missense mutations, results in a clinical spectrum ranging from mild to very severe with neonatal death.4,5 A founder effect for the mutation F119L (c.357C→A) in the Scandinavian population results in a more homogenous group of patients in these countries.6 Around 37% of CDG-Ia patients are heterozygous for the missense mutation R141H (c.422G→A), which has never been observed in the homozygous state.7,8 The very low residual (

Developmental dyslexia is diagnosed as a specific impairment in reading ability, despite adequate intelligence and educational opportunity,1 that affects approximately 5% of schoolchildren.2 Much evidence has been accumulated from twin and family based studies to indicate that dyslexia can have a hereditary basis, but that the genetic aetiology is complex, involving multiple risk factors.1–3 Linkage analysis has identified numerous genomic regions that may harbour susceptibility genes influencing dyslexia, including on chromosomes 1, 2, 3, 6, 15, and 18, with varying degrees of reproducibility.1,2 The first of these linkages was reported two decades ago,4 to the centromere of chromosome 15. Although subsequent studies failed to replicate linkage to this specific region,5 there is evidence for linkage elsewhere on chromosome 15, particularly at 15q21 ( DYX1 , OMIM 127700).6,7 Recently, DYX1C1 (also known as EKN1 ) was proposed as the gene underlying the putative effect on 15q21.8 This was initially based on studies of a balanced translocation, t(2;15)(q11;q21), co-segregating with reading problems within a single nuclear family from Finland.9 The 15q21 breakpoint in this family directly disrupts DYX1C1 , in an interval that includes exons 8 and 9 (fig 1). Investigation of DYX1C1 in individuals from 20 additional Finnish families with multiple cases of dyslexia led to the identification of eight single nucleotide polymorphisms (SNPs). Two of these SNPs were found to associate with dyslexia in these families, and in additional Finnish affected cases and controls. It was proposed that these two associated SNPs altered the expression or function of DYX1C1 , one by altering a transcription factor binding site, the other as a result of a premature truncation of the protein product by four amino acids, thereby leading to increased risk of developing dyslexia.8 Figure 1 Location of DYX1C1 on chromosome 15. Top: …

The dopamine D3 receptor is a member of the D2 family of dopamine receptors. Several investigators have suggested that D3 receptors are involved in the regulation of locomotion,1 in the pathophysiology of schizophrenia,2 and in drug abuse.3 Dopamine enhances basal and stimulus evoked release of arachidonic acid, as well as the production of one of its metabolites, prostaglandin E2 (PGE2), in CHO (Chinese hamster ovary) cells transfected with the D2 receptor.4–6 In contrast, dopamine does not influence basal arachidonic acid release7–9 but inhibits stimulus evoked release in cells transfected with the rat D3 receptor.10 On the other hand, with respect to cAMP production, the effect of dopamine is similar in CHO cells transfected with either D2 or D3 receptors, both receptor subtypes mediating a reduction in forskolin induced cAMP formation.11,12 A single nucleotide polymorphism (SNP) in the first exon of the dopamine D3 receptor gene, the ser9gly polymorphism, leads to a serine to glycine amino acid substitution in the N -terminal extracellular domain of the receptor protein.13 An association between this polymorphism and schizophrenia has been suggested, but also questioned,14,15 a meta-analysis suggesting the effect to be small but real in white subjects.16 Other investigators have suggested that the gly-9 allele may be associated with an increased risk of developing antipsychotic induced tardive dyskinesia rather than with schizophrenia per se17; this finding also gained support from a report containing both pooled analyses of original data and a meta-analysis,18 and also from a recent primate study19 (see also a recent report by Lohmueller and coworkers20). In addition, the ser9gly polymorphism has been reported to be associated with therapeutic response to atypical antipsychotic agents, …

The melanocortin-4 receptor gene ( MC4R ) is involved in central energy homeostasis and body weight regulation. Both endogenous anorexigenic and orexigenic ligands bind to the receptor.1 Under normal conditions, the anorexigenic tone prevails as revealed by the fact that Mc4r knock-out mice2 develop elevated body weight. Mc4r−/− mice show higher food intake but a similar metabolic rate and similar decreased physical activity compared to wild type (WT) mice of the same strain.2–4 In comparison to a standard low fat diet, this deviant regulation of energy homeostasis is even more pronounced upon intake of a moderately fat diet,5 which leads to an even higher body mass. In all studies, the effect on body weight is smaller in heterozygous than in homozygous knockout mice, but the exact degree of dominance is not clear. In heterozygous Mc4r+/− animals, body mass is increased on average by about 7–45% and in homozygous Mc4r−/− by 50–100% compared to WT2–5 with substantial overlap between groups. The mutations might have a sex dependent effect, but the results are contradictory. In one study, the effect in males was only about half of that in females.2 However, two studies did not detect a sex by genotype interaction in this Mc4r−/− strain.4,5 One study in a different knockout line of the same inbred strain found a sex by genotype interaction in the opposite direction, with only a marginal effect in heterozygous females whereas heterozygous males had a body weight intermediate between WT and homozygous knockouts.3 The first mutations in the human MC4R gene were reported in extremely obese probands.6–8 Since then, several other studies investigated the association of different MC4R mutations with obesity. According to a recent overview,9 at least 34 putatively functionally relevant …

Complex I is the largest of the mitochondrial respiratory chain enzyme complexes, consisting of at least 46 subunits, seven of which are encoded by mtDNA. Deficiency of complex I is the most common respiratory chain defect, and can be caused by mutations in both nuclear and mtDNA encoded genes. It has a wide range of clinical presentations, from lethal infantile mitochondrial disease to isolated myopathy.1–3 Mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) is one of the syndromes associated with complex I deficiency and in approximately 80% of cases is caused by a mutation, 3243A→G, in the mitochondrial tRNALeu(UUR) gene ( MTTL1 ). Other mutations in MTTL1 and other transfer RNA genes ( MTTF , MTTV , MTTQ ) account for most of the remainder of cases.4 However, a number of mutations in the mitochondrial MTND subunit genes of complex I have also been reported to cause MELAS, most notably in MTND5 5 and to a lesser extent in MTND6 .6 In stark contrast, there are presently no mutations in the MTND1 subunit gene associated with MELAS. There are several mutations in MTND1 associated with Leber’s hereditary optic neuropathy (LHON) which may be pathogenic, but only one, the 3460G→A mutation, that has robust evidence, including cell biology studies, for pathogenicity.7–9 Here we report three unrelated patients with MELAS and isolated complex I deficiency in skeletal muscle and cultured fibroblasts due to previously unreported mutations in the MTND1 gene. Evidence confirming the pathogenic nature of these mutations includes data from cell fusion experiments and blue native polyacrylamide gel electrophoresis (BN-PAGE), the latter confirming a crucial role for the ND1 subunit in the assembly of complex I holoenzyme.10 ### Patient 1 Patient 1, a white male, presented at 4 years of age with a 3 month history of increasing tiredness, clumsiness, …

Idiopathic epilepsies have a genetic basis and are characterised by the absence of an overt underlying neurological abnormality. Most idiopathic epilepsies are complex diseases with considerable clinical and genetic heterogeneity and an unclear inheritance pattern because of genetic and environmental factors. Families in which the disease segregates as an autosomal dominant trait with reduced disease penetrance have been identified occasionally. In some of these families, a single gene defect was identified as the cause of epilepsy. To date, mutations in 13 genes have been identified for distinct epilepsy types. Most genes encode subunits of ion channels.1,2 In addition, the gene remains to be identified for 21 mapped loci for epilepsy, which highlights the genetic heterogeneity of the idiopathic epilepsy syndromes.3,4 Familial temporal lobe epilepsy (MIM 608096) was first described by Berkovic et al . and was recognised as a distinct epileptic syndrome by the International League Against Epilepsy.5 It is defined by familial occurrence of simple partial seizures, complex partial seizures, and secondarily generalised seizures of temporal lobe origin.6 Two genetically distinct autosomal dominant familial temporal lobe epilepsy syndromes have been reported. Autosomal dominant lateral temporal lobe epilepsy (MIM 600512), or autosomal dominant partial epilepsy with auditory features, was described first by Ottman et al . ,7 and recently, mutations in the leucine rich glioma inactivated 1 ( LGI1 ) gene on chromosome 10q24 were identified.8,9 Auras that present as auditory and visual hallucinations are a clinical hallmark of this syndrome. The other variant of familial temporal lobe epilepsy is characterised clinically by onset in teenage years or early adulthood, absence of antecedent factors, low frequency of deja vu, and a usually good prognosis. This variant, which still can be heterogeneous genetically, is not mapped yet. In a large family with febrile seizures …

Butterfly shaped macular dystrophy was first described by Deutman et al. in 1970.1 It is characterised by bilateral accumulation of pigmented or yellowish material at the level of the retinal pigment epithelium. Lesions consist of 3–5 “wings,” which resemble the wings of a butterfly. Affected patients present with a subnormal electrooculogram and normal or slightly diminished visual acuity. The disease is relatively benign, but it can progress with age to chorioretinal atrophy in the parafoveal and peripapillary regions.2 Butterfly shaped macular dystrophy shares important similarities with age related macular degeneration—the most common cause of blindness in older patients.3,4 In both diseases, abnormal deposition of lipofuscin like material at the level of the retinal pigment epithelium is found, which results in loss of the overlying photoreceptors.2 Butterfly shaped macular dystrophy has an autosomal dominant inheritance pattern. To date, it has been associated only with mutations in the peripherin / RDS gene.5–12 We ascertained members of the family with butterfly shaped macular dystrophy that was described originally by Deutman et al. in 1970,1 and we excluded peripherin / RDS as the causative gene in this family.2 In addition, we excluded the ROM-1 gene; four genes expressed in cone photoreceptors; all known non-syndromic macular, retinal pigment epithelium, and choroidal dystrophy loci; all known Leber congenital amaurosis loci; and all known non-syndromic congenital and stationary retinal disease loci.2 This study aimed to identify the locus responsible for butterfly shaped macular dystrophy in this family with a genomewide linkage scan. ### Patients and ophthalmic examination We previously ascertained 13 members (eight affected and five unaffected) of a Dutch family with butterfly shaped macular dystrophy that was first described in 1970 by Deutman et al.1,2 Ophthalmic examination of the participating family members included best corrected Snellen visual acuity, slit …

Hereditary paraganglioma (PGL) is characterised by slow growing, vascular tumours that can develop in any component of the paraganglia, a neuro-ectodermal system that is distributed from the skull base to the pelvic floor.1 Common tumour sites include the carotid body in the head and neck and adrenal and extra-adrenal paraganglia in the abdomen. Heterozygous germline inactivating mutations in SDHD , SDHC , and SDHB , which encode three of the four subunits of mitochondrial complex II (succinate dehydrogenase), cause hereditary paraganglioma types 1, 3, and 4 (PGL1, PGL3, and PGL4), respectively. Mutations in the fourth subunit of mitochondrial complex II, SDHA , have yet to be demonstrated in hereditary paraganglioma. Germline loss of function mutations followed by somatic loss of non-mutant alleles in the tumours2–4 suggests a tumour suppressor role for mitochondrial complex II in the paraganglia. Over 25 mutations in SDHD and 25 mutations in SDHB have been detected in hereditary paraganglioma, including those reviewed by Baysal1 and the more recent additions of multiple mutations in SDHB 4–6 and SDHD .7–9 All reported mutations are single nucleotide alterations leading to splice site, missense, nonsense, or frameshift mutations, or intra-exonic deletions and insertions of up to four nucleotides, which have been detected through exonic PCR amplifications and sequencing. In contrast to the abundance mutations in SDHB and SDHD , only a single multiply affected family and an isolated case, containing a single nucleotide initiation codon and a splice site mutations in SDHC , respectively, have been described by Niemann et al .3,10 However, analyses of SDHC in four series of patients with paraganglioma or pheochromocytoma 6,8,11,12 yielded no definitive SDHC mutations. These findings indicate that the relative contribution of complex II subunit mutations to hereditary paraganglioma is not …

Wolf-Hirschhorn syndrome is a multiple malformation syndrome with distinct abnormal craniofacial features, prenatal onset growth retardation, failure to thrive, microcephaly, usually severe mental retardation, seizures, and congenital heart malformations. Large variations are observed in phenotypic expression of these features, with mental retardation ranging from severe to mild. There is a one third mortality in the first two years of life. Most patients with Wolf-Hirschhorn syndrome carry 4p terminal deletions. However, the size of these deletions is variable and several phenotypic features have been tentatively mapped within the 4pter region.1–4 Further fine mapping of the different phenotypic features will ultimately lead to a functional understanding of the genes that cause these abnormal phenotypes. The minimal ‘Wolf-Hirschhorn syndrome’ phenotype was defined as the typical facial appearance, congenital hypotonia, mental retardation, growth delay, and seizures.2,4 The Wolf-Hirschhorn syndrome critical region was originally confined to a region of 165 kb and nine transcripts within this region were described.5 A patient with a small intrachromosomal 4p deletion and a partial Wolf-Hirschhorn syndrome phenotype further refined the critical region (WHSCR1).6 Two genes, the Wolf-Hirschhorn Syndrome Candidate genes 1 (WHSC1) and 2 (WHSC2), are located in the region. The expression pattern of WHSC1 colocalises spatially and temporarily with the major Wolf-Hirschhorn syndrome malformations and the gene is homologous with a Drosophila dysmorphology gene.7 WHSC2 is a nuclear protein with a helix-loop-helix motif that is ubiquitously expressed throughout development.8,9 The identification of a Wolf-Hirschhorn syndrome patient with a terminal 1.9 Mb deletion not including this Wolf-Hirschhorn syndrome critical region led Zollino et al4 to postulate a novel critical region distal to the previously defined critical region, which was termed the Wolf-Hirschhorn critical region 2 (WHSCR2). The distal boundary of this region is located within the WHSCR1 and at …

Hutchinson-Gilford progeria syndrome (HGPS) is a rare fatal genetic disorder which is characterised by the very early appearance of accelerated ageing in children. It was first described by Hutchinson1 and Gilford.2 Children with HGPS appear healthy at birth, with distinctive clinical features appearing within the first few years of life, including severe growth retardation, atherosclerosis, incomplete sexual maturation, loss of subcutaneous fat, craniofacial abnormalities, alopecia (hair loss), stiffness of joints, and other skeletal abnormalities including coxa valga (giving a “horse-riding” stance), delayed and abnormal dentition, and scleroderma-like areas of skin, with normal mental and emotional development.3 A specific causal mutation for HGPS in the LMNA gene was identified4,5 and down-regulation of isoforms of the protein arising from the LMNA gene were found in HGPS patients.6 The LMNA gene encodes the A-type lamins, lamins A and C, in which several other laminopathies have their causal mutation.7,8 The heterozygous de novo point mutation is located in exon 11 of the LMNA gene, which is transcribed to lamin A while the shorter lamin C is unaffected by the mutation. The GGC>GGT G608G single-base substitution reveals a cryptic splice site and is proposed to result in a truncated transcript encoding a protein that is 50 amino acid shorter. This mutant protein will be referred to as progerin. A less common mutation in the same codon (GGC>AGC), predicted to have similar effects on lamin A splicing, was also identified.4 Western blots reveal progerin and the normal lamin A protein in HGPS patient cells.5 This hypothesised mechanism raises questions about allelic expression and splicing efficiency. De Sandre-Giovannoli et al 4 postulated that only the mutated allele may be expressed in HGPS cells. We now examine lamin A transcript levels in HGPS cells, identify their allelic sources …

ystic fibrosis is a common lethal autosomal recessive disease affecting whites with an incidence of about 1 in 2500. The median lifespan is approximately 30 years. Chronic obstruction and infection of the respiratory tract, pancreatic insufficiency, elevated sweat electrolyte concentration, and male infertility characterise this disease. Clinical manifestations are attributed to mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, 1 which encodes an epithelial chloride channel. 2 Certain phenotypes of cystic fibrosis, such as pancreatic insufficiency, are highly associated with the CFTR genotype. Pulmonary symptoms are highly variable, even among patients from the same family. 3 Other genetic factors, as well as environmental factors, thus affect the cystic fibrosis disease phenotype. In association studies, evidence has been found that mannose binding lectin (MBL) protein affects cystic fibrosis disease. 45 MBL protein is an important mediator component of the innate immune defense system, which functions as an opsonin and complement activator. It is part of a family of proteins called collectins, because they contain collagen-like regions and lectin domains. Through the lectin domain, they bind to carbohydrate structures presented by a wide range of pathogenic bacteria, viruses, fungi, and parasites. Mannose binding lectin protein is synthesised in the liver by hepatocytes and secreted in the blood, and circulates as dimers or hexamers composed of subunits containing three identical polypeptides. 67

Muir-Torre syndrome (MTS; MIM 158320) is an autosomal dominant predisposition to skin tumours and various internal malignancies. Clinical criteria for a diagnosis of MTS are the synchronous or metachronous occurrence of at least one sebaceous gland neoplasia and at least one internal neoplasm in a patient (regardless of the family history).1,2 The sebaceous gland neoplasias comprise adenomas, epitheliomas (sebaceomas), and carcinomas. In contrast, the frequent sebaceous gland hyperplasia is not indicative of MTS.2,3 According to Schwartz and Torre,2 the sebaceous neoplasias precede the internal neoplasias or are concurrent with them in 41% of MTS patients. As sebaceous gland neoplasias are rare, MTS should always be suspected when a sebaceous tumour has been diagnosed. Cystic sebaceous neoplasia is probably the most sensitive marker for this tumour predisposition syndrome.2,4–6 Colorectal cancer is by far the most common internal malignancy in MTS patients.7 The spectrum of internal malignancies in MTS is similar to the various tumour entities observed in hereditary non-polyposis colorectal cancer (HNPCC; MIM 114500). HNPCC is an autosomal dominant cancer predisposition syndrome characterised by early onset of colorectal cancer and other associated tumours.8,9 Several genes underlying HNPCC which are involved in DNA mismatch repair (MMR) have been identified within the last decade.10–13 Germline mutations in the DNA MMR genes were detected in a high proportion of MTS patients, demonstrating that MTS most often represents a phenotypic variant of HNPCC.14,15 Due to the underlying genetic mechanisms of tumourigenesis, tumours of these MTS patients exhibit high microsatellite instability (MSI-H), the characteristic feature of HNPCC tumours.16 Microsatellite analysis in tumour tissue of MTS patients therefore provides a useful tool to pre-select patients for mutation analysis in DNA MMR genes.3 Immunohistochemical testing for expression of the MSH2 and …

Neurofibromatosis type 2 (NF2) is an autosomal dominant condition characterised by vestibular schwannomas, schwannomas of other cranial nerves, meningiomas, and other low grade brain malignancies.1 The severity of NF2 is variable, with some patients having early onset disease and more rapidly growing tumours that occur in greater numbers. The NF2 gene is on chromosome 22q12.2,3 The protein product (termed merlin or schwannomin) is a cell cytoskeleton associating protein. Genotype−phenotype correlations have been demonstrated, with missense mutations and large deletions causing mild disease, and nonsense or frameshift mutations causing severe disease.4,5 The current mutation screening techniques of single strand conformation polymorphism analysis (SSCP), protein truncation test, and denaturing gradient gel electrophoresis detect 33–65% of mutations, although adding a deletion strategy increases the proportion to 80%.6 However, deletion testing and chromosome analysis are rarely reported in studies of NF2 mutations. In this study, we present five NF2 patients for whom chromosome analysis, usually following routine molecular screening, revealed the underlying genetic aberration. ### Case 1 A 20 year old female was referred with a large diffuse goitre commencing early in puberty, moderate learning difficulties, and bilateral sensorineural hearing loss, which resembles the phenotype of Pendred’s syndrome. She was born after a normal pregnancy and delivery. Her physical and mental development during early infancy was not remarkable, but developmental delay became apparent in the second year of life. Growth parameters were within the normal range. She began to walk at the age of 3 years. Six months later, audiometric tests showed no hearing impairment although chronic otitis media was diagnosed on one side. According to the parents, her communication had been improving with age, but her speech remained dysarthric and her articulation was very poor. In primary school, she had sub-average grades in all subjects. Her affect was good and …

Evidence of genotype specific selection at the BRCA2 polymorphism N372H was originally reported by Healey et al.1 These workers found significant evidence of a heterozygous excess in women control samples when studying the polymorphism for breast cancer association. They then genotyped a large series of newborn boys and girls to examine if this effect was also seen at birth. The newborn girls were consistent with the women but the newborn boys were significantly different. The genotypes in the boys appeared to demonstrate a significant deficit of heterozygotes. Healey et al suggested that sex differential viabilities resulted in a stable allele frequency, but this was not formally investigated. We wanted to explore further this apparent sex specific selection in two ways; to refine the estimates of fitness, and we examined the mathematical properties of the sex specific selection model to determine if the data were consistent with a stable equilibrium. Three control populations were available for additional genotyping. These were blood donors, colon cancer controls, and mammography screening control women. Anonymous blood donors were recruited from the Sheffield Blood Transfusion Service in 1996, and their ages and sexes were recorded. Blood DNA samples were obtained for the mammography screening series of controls from white women attending for routine mammography screening at the Sheffield Breast Screening Service between September 2000 and August 2002. Women’s DNA samples were included in the study if the mammogram showed no evidence of a breast lesion. All women resident in Sheffield and on the Community Health Index, who are between the ages of 50 and 65, are invited to attend for screening every three years, and the uptake in Sheffield is 81.2%. Ethical approval for this study was obtained from the South Sheffield Research Ethics Committee and informed written consent was obtained from all subjects. Colon …

Serine protease functionality is based on nucleophilic attack of a targeted peptidic bond by a serine. The serine protease superfamily is extremely diverse and includes proteases such as plasminogen, prostatin, hepsin, the kallikrein family ( KLK genes clustered on chromosome 19.13), and a recently discovered cluster of tryptic-like serine proteases located on human chromosome 16p13.1,2 Serine protease mutations have been reported as causative in only a few autosomal recessive human hereditary conditions, which produce diverse pathological conditions.3,4 We report the first human kallikrein mutation and describe its association with a rare autosomal recessive form of amelogenesis imperfecta. The amelogenesis imperfectas are a clinically and genetically heterogeneous group of disorders characterised by faulty development of the tooth enamel due to hypoplasia or hypomineralisation.5 The amelogenesis imperfecta phenotypes vary widely depending on the specific gene involved, the location and type of mutation, and the corresponding putative change at the protein level.6,7 The amelogenesis imperfecta enamel defects can be broadly divided into hypoplastic (enamel crystallites do not grow to the correct length) and hypomineralised (crystallites fail to grow in thickness or width) phenotypes. The prevalence of amelogenesis imperfecta varies in different countries (ranging from 1 in 700 in Sweden to 1 in 14 000 in the United States) suggesting allele frequency differences between populations.8–11 Amelogenesis imperfecta can be inherited as an autosomal dominant, autosomal recessive, or X-linked Mendelian trait. While autosomal dominant amelogenesis imperfecta types are most common in the United States and Europe, autosomal recessive amelogenesis imperfecta types are more common in the Middle East.8,10,11 Dental enamel is the most highly mineralised tissue in the human body with 85% of its volume occupied by highly organised carbonate substituted hydroxyapatite crystals.12 These crystallites are packed into a highly ordered decussating prism …

Cutaneous malignant melanoma (CMM) is a potentially fatal form of skin cancer, whose incidence and mortality are increasing in the Western world.1,2 Approximately 3–12% of cutaneous malignant melanoma develops in families with multiple cases of melanoma.3–5 Worldwide studies of large families prone to melanoma have demonstrated linkage to a locus on chromosome 9p21 (MIM 600160) in the majority of kindreds, and probable linkage to 1p22 6 and 1p36 7,8 in others. About one third of 9p21 linked families carry mutations in the CDKN2A tumour suppressor gene,9,10 which encodes the p16 cell cycle inhibitor. Rare kindreds may also possess mutations of the coding sequences of CDK4 (MIM 12829),11,12 or p14ARF (translated from exons 1β and 2 of CDKN2A ).13–15 More studies are needed to understand the genetic basis of melanoma. In Italy, the melanoma prone families studied to date are mostly from the north16 and northwestern areas,15,17,18 the centre,19 and the Sardinia region.20 These families are generally characterised by small numbers of melanoma cases, and a relatively high frequency of CDKN2A mutations. We studied families prone to melanoma from northeastern Italy, to characterise genetic susceptibility to melanoma in this population. ### Study population The study group was comprised of 55 families: 44 (80%) from southern Emilia Romagna and northern Marche (close to the border between the two regions); 10 families, all with two cases per family, from other areas of Italy (four from the south, three from the centre, two from the north, and one from Sardinia), and one family from Russia. All the families were recruited at the Dermatology Unit of the Bufalini Hospital in Cesena, Italy. Bufalini Hospital’s and the National Cancer Institute’s Ethical Committees approved the study, and written informed consent was obtained …

True hermaphroditism, defined clinically as the presence of both male and female gonadal tissue in the same individual usually accompanied by ambiguous genitalia, is a genetically heterogeneous condition.1–3 Less than 10% of true hermaphrodites with an apparent 46,XX constitution are SRY -positive, usually resulting from translocation between Xp and Yp.4–7 Previously we described two true hermaphrodites (AK and PG) of Polish origin, both of whom carry X/Y translocations with SRY translocated onto distal Xp.6 Here, we show that the same X/Y translocation is also carried by fertile male and female relatives of the hermaphrodite probands, and identify varying sexual phenotype in individuals with the same chromosome abnormality. AK was ascertained aged 17 with ambiguous internal and external genitalia. Biopsy revealed a left ovary and a right testis with no signs of spermatogenesis. PG was ascertained at 6 months of age with ambiguous internal and external genitalia. She was found to have a right ovary and a left ovotestis, which was removed and upon microscopical examination showed signs of dysgenesis in the testicular part. Circulating testosterone levels were