Heterozygous missense mutation in the rod cGMP phosphodiesterase beta-subunit gene in autosomal dominant stationary night blindness.

The locus for autosomal dominant congenital stationary night blindness (adCSNB) has recently been assigned to distal chromosome 4p by linkage analysis in a large Danish family. Within the candidate gene encoding the beta-subunit of rod photoreceptor cGMP-specific phosphodiesterase (beta PDE), we have identified a heterozygous C to A transversion in exon 4, predicting a His258Asp change in the polypeptide. We found a perfect cosegregation (Zmax = 22.6 at theta = 0.00) of this mutation with the disease phenotype suggesting that this missense mutation is responsible for the disease in this pedigree. Homozygous nonsense mutations in the beta PDE gene have been found recently in patients with autosomal recessive retinitis pigmentosa, a common hereditary photoreceptor dystrophy.

Study of Toll-like receptor gene loci in sarcoidosis.

Sarcoidosis is a multi-factorial systemic disease of granulomatous inflammation. Current concepts of the aetiology include interactions of unknown environmental triggers with an inherited susceptibility. Toll-like receptors (TLRs) are main components of innate immunity and therefore TLR genes are candidate susceptibility genes in sarcoidosis. Ten members of the human TLR gene family have been identified and mapped to seven chromosomal segments. The aim of this study was to investigate all known TLR gene loci for genetic linkage with sarcoidosis and to follow positive signals with different methods. We analysed linkage of TLR gene loci to sarcoidosis by use of closely flanking microsatellite markers in 83 families with 180 affected siblings. We found significant linkage between sarcoidosis and markers of the TLR4 gene locus on chromosome 9q (non-parametric linkage score 2.63, P = 0.0043). No linkage was found for the remaining TLR gene loci. We subsequently genotyped 1203 sarcoidosis patients from 997 families, 1084 relatives and 537 control subjects for four single nucleotide polymorphisms of TLR4, including Asp299Gly and Thr399Ile. This genotype data set was studied by case-control comparisons and transmission disequilibrium tests, but showed no significant results. In summary, TLR4 - w ith significant genetic linkage results - appears to be the most promising member of the TLR gene family for further investigation in sarcoidosis. However, our results do not confirm the TLR4 polymorphisms Asp299Gly and Thr399Ile as susceptibility markers. Our results rather point to another as yet unidentified variant within or close to TLR4 that might confer susceptibility to sarcoidosis.

Preimplantation genetic diagnosis: the German situation.

Preimplantation genetic diagnosis makes it possible to detect some genetic disorders in embryos in vitro before they are transferred to the uterus. Using this technique, there is an opportunity for couples who have an increased risk of transmitting severe genetic disorders to their offspring to reduce this risk by >95%. By doing PGD, abortions at a later stage can be avoided.

Spinocerebellar ataxia type 4. Investigation of 34 candidate genes.

The spinocerebellar ataxias (SCAs) with autosomal dominant inheritance are a clinically and genetically heterogeneous group of neurodegenerative disorders. To date 24 different loci have been identified for these conditions. A locus at chromosome 16q22.1 co-segregates with the disease phenotype in families of Scandinavian, Japanese and German origin. The corresponding SCA4 locus was narrowed down to 7.94 Mb for the two European and to 1.25 Mb for Japanese pedigrees. Unfortunately, because of the phenotypic differences between patients from Japan and Europe it is not possible to decide if SCA families linked to chromosome 16q22.1 share a common disease genotype or not. To look for mutations in the German family we screened 34 candidate genes in a 3.69 cM region. With the exception of two cSNPs, no segregation of DNA variations with the disease phenotype was found.

A new technique for cyclic in situ amplification and a case report about amplification of a single copy gene sequence in human metaphase chromosomes through PCR-PRINS.

Since the introduction of PRimed IN Situ labeling (PRINS) as a rapid and extremely sensitive alternative method to conventional fluorescence in situ hybridization (FISH), its application in clinical cytogenetics has been limited to the detection of highly repeated sequences, such as centromeric and telomeric regions. In the original PRINS method, unlabeled oligonucleotide probes are annealed to their repeated complementary target sequences in fixed human metaphase chromosomes on a slide. The probes serve as primers for subsequent in situ chain elongation with Taq DNA polymerase and labeled nucleotides. In contrast to conventional PCR, cyclic in situ amplification of the chromosomal target DNA with paired primers remained both difficult and strictly limited to highly repeated sequences, since the maintenance of constant reaction conditions on the slide during temperature and pressure shifts presents a major problem. We developed a new system for in situ PCR that allows the amplification of target sequences analogous to PCR in the test tube. We applied this method successfully for the detection of highly repeated sequences, for the detection of low copy repeats, and in one case, for the detection of a single-copy DNA sequence. The significance of this development for further in situ PCR applications will be discussed.

Clinical, endocrinological, and cytological characterization of two 46, XX males.

In two 46,XX males, 20 and 21 yr of age, gonadotropins, testosterone, and estradiol were measured in serum and compared to those in a control group of four men. In addition, in one subject, androgen metabolism was measured in biopsied skin and cultured skin fibroblasts. In both XX men, a pulsatile pattern of gonadotropin, testosterone, and estradiol release into serum was observed. The levels of the gonadotropins and estradiol were higher and the levels of testosterone were lower in XX men than in normal men. hCG stimulation resulted in a significant increase in testosterone secretion, and LRH administration caused a more prolonged rise in gonadotropin levels in the XX men. The administration of estradiol caused a positive feedback response in the XX men and resulted in a suppression of gonadotropin secretion in the controls. Finally, the formations of C-19 metabolites from testosterone and estrone from androstenedione were found to be in the same range in skin and skin fibroblasts from the XX men as in those from normal men. It can be concluded that 46,XX men have altered hypothalamic-pituitary and gonadal function compared to normal men.

Eye-tracking dysfunction (ETD) in families with sporadic and familial schizophrenia.

BACKGROUND: Within the field of genetic schizophrenia research, eye-tracking dysfunction can be regarded as a putative trait marker in families with multiple occurrences of the disease (familial schizophrenia). We concentrated on families with single occurrences of schizophrenia (sporadic schizophrenia) to test whether a genetic factor may be present in these families as well. METHODS: Eye movements were recorded using infrared oculography in eight families with sporadic schizophrenia (44 members), eight families with familial schizophrenia (66 members), and nine nonpsychotic families (77 members). Triangle-wave stimuli at 15 degrees /sec and 30 degrees /sec were used, and gains (eye velocity/target velocity), rates, and amplitudes of saccades (classified as catch-up and anticipatory saccades) were determined. RESULTS: 1) In sporadic-schizophrenia families, gain values, saccade rates, and anticipatory saccade amplitudes at 30 degrees /sec differed in a statistically significant fashion from nonpsychotic families, but not from families with multiple occurrences of schizophrenia, and 2) at 30 degrees /sec, a significant effect of target direction on smooth-pursuit maintenance was observed in both sporadic- and familial-schizophrenia families. CONCLUSIONS: Our results support the hypothesis that genetic factors may be present even in sporadic-schizophrenia families and may contribute to a more precise and biologically based definition of the schizophrenia phenotype in future molecular genetic analysis.

Smooth pursuit performance in families with multiple occurrence of schizophrenia and nonpsychotic families.

BACKGROUND: Eye tracking dysfunction (ETD) has been put forward as a trait marker for biological susceptibility to schizophrenia with the hope of identifying a link to specific cerebral lesions. METHODS: Eye movements were recorded using infrared oculography in 8 families (67 members) showing multiple occurrence of schizophrenia and in 9 nonpsychotic families (80 members). Triangle wave stimuli at 15 degrees/s and 30 degrees/s were used and gains (eye velocity/target velocity), rates and amplitudes of different saccade categories (catch-up, back-up, anticipatory saccades, and squarewave-jerks) were determined. RESULTS: In the relatives, the same deficit in maintenance of smooth pursuit performance was found as was seen in the schizophrenic patients. This deficit, which was not observed in the nonpsychotic families, consisted of lower gains for leftward as compared to rightward pursuit. This was emphasized most clearly at 30 degrees/s and was associated with an excess of catch-up saccades in the schizophrenic patients, whereas in the relatives a tendency to exhibit more and larger anticipatory saccades was observed. CONCLUSIONS: The results confirm the hypothesis that eye-tracking dysfunction is a phenotypic marker for genetic liability to schizophrenia. Neurophysiologically, a cerebral dysfunction which includes one or more of the oculomotor centers can be assumed in subjects who carry a genetic susceptibility to schizophrenia.

Extension of the mutation spectrum in Friedreich's ataxia: detection of an exon deletion and novel missense mutations.

Friedreich's ataxia (FRDA), the most common autosomal recessively inherited ataxia, is due to a homozygous GAA triplet repeat expansion in the first intron of the FRDA gene in about 96% of patients. Approximately 4% of FRDA patients are compound heterozygotes with a GAA repeat expansion in one allele and a point mutation in the coding region of the second allele. To reinvestigate the mutation spectrum, we searched for mutations including exon deletions in six patients heterozygous for the GAA repeat expansion and found two unknown missense mutations, p.Asn146Lys and p.Leu186Arg, in trans to the expanded FRDA allele. Interestingly, we detected a heterozygous 2776 bp deletion including exon 5a in one of our patients. This deletion removes 50 of the 210 residues of the frataxin. Furthermore, since no FRDA case with two-point mutations is known, we screened eight patients with FRDA phenotype but GAA alleles within the normal range but did not reveal a mutation within the FRDA gene. In addition, DNA polymorphisms have been found in four out of 100 control individuals in this study.

SCA17 caused by homozygous repeat expansion in TBP due to partial isodisomy 6.

An expanded polyglutamine domain in the TATA-binding protein (TBP) has been described in patients with spinocerebellar ataxia type 17 (SCA17) characterized by cerebellar ataxia associated with dementia. TBP is a general transcription initiation factor that regulates the expression of most eukaryotic genes transcribed by RNA polymerase II. SCA17, as an autosomal dominantly inherited progressive neurodegenerative disorder, is caused by heterozygous expansion of a CAG repeat coding for glutamine. Alleles with 27 to a maximum of 44 glutamine residues were found as the normal range, whereas expansions above 45 repeat units were considered pathological. Here, we present a patient with a very severe phenotype with a late onset but rapidly progressing ataxia associated with dementia and homozygous 47 glutamine residues caused by an apparent partial isodisomy 6. This extraordinary case has important implications for the insights of TBP and SCA17. The expanded polyglutamine domain in both TBP copies is not correlated with embryonic death indicating that the normal function of the protein is not disrupted by this kind of mutation but may account for the dementia seen in this patient.

Different types of repeat expansion in the TATA-binding protein gene are associated with a new form of inherited ataxia.

A novel neurological syndrome has recently been described to be associated with an expanded polyglutamine domain. The expansion results from partial duplication within the TATA-binding protein (TBP). By investigation of 604 sporadic and familial cases with various forms of neurological syndromes and 157 unaffected individuals, we found repeat expansions in the TBP in four patients of two families with autosomal dominant inheritance of ataxia, dystonia, and intellectual decline. Two different genotypes for the repetitive sequence could be demonstrated which led to elongated polyglutamine stretches between 50 and 55 residues, whereas normal alleles with 27 to a maximum of 44 glutamine residues were found in this study. The expansion to 50 or more glutamine residues results in a pathological phenotype and confirms the report of a new polyglutamine disease.

Evaluation of 50 probands with early-onset Parkinson's disease for Parkin mutations.

BACKGROUND: Early onset PD has been associated with different mutations in the Parkin gene, including exon deletions and duplications. METHODS: The authors performed an extensive mutational analysis on 50 probands with onset of PD at younger than 50 years of age. Thirteen probands were ascertained from a registry of familial PD and 37 probands by age at onset at younger than 50 years, blind to family history. Mutational analysis was undertaken on the probands and available family members and included conventional techniques (single strand conformation polymorphism analysis and sequencing) and a newly developed method of quantitative duplex PCR to detect alterations of gene dosage (exon deletions and duplications) in PARKIN: RESULTS: Using this new technique, the authors detected eight alterations of gene dosage in the probands, whereas 12 mutations were found by conventional methods among the probands and another different mutation in an affected family member. In total, the authors identified compound heterozygous mutations in 14%, heterozygous mutations in 12%, and no Parkin mutation in 74% of the 50 probands. We expanded the occurrence of Parkin mutations to another ethnic group (African-American). CONCLUSION: The authors systematically screened all 12 Parkin exons by quantitative PCR and conventional methods in 50 probands. Eight mutations were newly reported, 2 of which are localized in exon 1, and 38% of the mutations were gene dosage alterations. These results underline the need to screen all exons and to undertake gene dosage studies. Furthermore, this study reveals a frequency of heterozygous mutation carriers that may signify a unique mode of inheritance and expression of the Parkin gene.

Exon deletions in the GCHI gene in two of four Turkish families with dopa-responsive dystonia.

Most cases of dopa-responsive dystonia (DRD) are thought to be caused by mutations in the GCHI gene; however, by sequencing, mutations are found in only 40% to 60%. Recently, a single report identified, via Southern blot analysis, a large genomic GCHI deletion in a "mutation-negative" case. This report describes four families with DRD, two of which carry large deletions, thus confirming that deletions are an important subtype of GCHI mutations. These deletions were detected by quantitative duplex PCR that is amenable to DNA diagnostics.

A point mutation (W70A) in the rod PDE-gamma gene desensitizing and delaying murine rod photoreceptors.

PURPOSE: To examine the corneal electroretinogram (ERG) of transgenic mice (W70A mice) carrying a point mutation (W70A) in the gene encoding for the gamma-subunit of rod cGMP phosphodiesterase (PDEgamma). METHODS: The ERG of W70A mice was compared with that of normal mice. Cone responses were separated from rod responses by light adaptation, whereas rod sensitivity was assessed by threshold stimulation with dim light. Spectral sensitivity curves of the ERG were obtained using a constant response criterion. RESULTS: The ERG of the W70A mouse has a desensitized, delayed rod b-wave at threshold, and a prolonged rod b-wave at higher flash intensities. The a-wave is absent even at maximal stimulation. The cone ERG of the W70A mouse is indistinguishable from that of normal mice. The spectral sensitivity of the W70A mouse is maximal in the UV spectrum, in contrast to the normal mouse, which is most sensitive in the green region of the spectrum. This supports the interpretation of the results as normal cone and abnormal rod function in the W70A mouse. CONCLUSIONS: The W70A mouse represents new model of stationary nyctalopia that can be recognized by its unusual ERG features.

Fra(X) frequency on the active X-chromosome and phenotype in heterozygous carriers of the fra(X) form of mental retardation.

Female heterozygotes of the fra(X) form of mental retardation show variable degrees of mental impairment and phenotype expression of the disorder. This might be an effect of inactivation of the X-chromosome which carries the fra(X)(q). Prior replication studies in heterozygous carriers gave contradictory results with respect to possible genotype-phenotype correlation. In the interpretation of these studies it is important to understand the effect of BrdU on the fra(X)(q) expression. In a group of 13 hemizygous patients with fra(X)(q) and 7 heterozygous carriers we studied the effect of BrdU on fra(X) expression. In the heterozygous carriers the use of BrdU resulted in a significant suppression of the fra(X)(q), while in hemizygous patients no difference in fra(X)(q) frequency with or without BrdU could be observed. It can be concluded that BrdU suppresses the fra(X)(q) preferentially on the inactive X-chromosome. Thus the fra(X)(q) frequency on the active X-chromosome is of primary importance in phenotype correlation studies among heterozygous carriers. In our group of heterozygous carriers we observed a negative correlation between (IQ) phenotype and fra(X)(q) expression on the active X-chromosome. This suggests that the gene for the fra(X)(q) form of mental retardation is on the X-chromosome and undergoes inactivation.

Clinical diagnosis of partial duplication 7q.

We report on two sibs with partial dup (7q), a retarded 9-month-old boy and an aborted fetus of 17 weeks' gestational age. Besides minor anomalies, the boy had frontal bossing, macrocephaly with hydrocephaly, a high forehead, and a large fontanelle. GTG banded chromosomes showed a 14p+ abnormality. Because his mother carries a balanced, de novo translocation with a breakpoint in band 7q33, the boy has a duplication of the distal portion of band 7q33 and the segment 7q34----qter. Our findings suggest that the phenotype in terminal duplications of 7q may, in some patients, be recognized clinically.

Folic acid treatment in males and females with fragile-(X)-syndrome.

Ten males with the fragile X (fra(X] syndrome were treated with folic acid (10 mg/day) for 4 months in a double-blind design study. To eight heterozygotes with mental impairment and fra(X), folic acid was given for 4 months (10 mg/day) in an effort to study possible beneficial effects of folic acid. Psychological and cytogenetic testing were carried out during the trial. There was no improvement in concentration, fine motor co-ordination, or comprehension in the adult male and female patients of the study. One patient showed improvement under a control medications. In the females, improvement was seen only in the youngest patient, a 5-year-old girl. Folate treatment does not seem to be effective in fra(X) adults, but may have some effect in children of both sexes with the disorder. Cytogenetic studies using peripheral lymphocytes showed that the fra(X) frequency decreased significantly (t = 0.00856; 1% level) only in cells cultured in a folic acid-free medium but not in cells cultured in a medium with added antifolate (methotrexate). This shows a "contamination effect" of folate-free culture medium after oral folic acid treatment of these patients. The decrease of fra(X) involves primarily the early-replicating X when culturing with folic acid-free medium. A synergistic suppression effect of "external folate" and BrdU is the most likely explanation of this phenomenon.

VACTERL with hydrocephalus and branchial arch defects: prenatal, clinical, and autopsy findings in two brothers.

VACTERL association is defined as a combination of vertebral, anal, cardiac, tracheoesophageal, renal and limb anomalies, in particular radial defects. In recent years hydrocephalus was observed in patients with apparent VACTERL association. This particular condition was recognized as a hereditary entity with poor prognosis. Both autosomal recessive and X-linked forms were described. Here we report prenatal, clinical and autopsy findings in 2 brothers with this syndrome, who had, in addition, branchial arch anomalies. The recurrence in this family suggests X-linked inheritance. Branchial arch defects have so far not been described as part of the VACTERL+H syndrome. This observation further supports that a variety of brain anomalies including hydrocephalus associated with VACTERL anomalies represents separate entities with a considerable recurrence risk. The use of the term VACTERL "association" for these conditions is misleading and is discouraged.

Deletion of the Hunter gene and both DXS466 and DXS304 in a patient with mucopolysaccharidosis type II.

Hunter syndrome is an X-linked mucopolysaccharidosis due to deficiency of the lysosomal enzyme iduronate-2-sulfatase (IDS). A cDNA clone containing the entire coding region of the human IDS gene, mapped in Xq28, has been used as molecular probe to study a patient with Hunter syndrome. A submicroscopic deletion has been detected that spans the IDS gene as well as DXS466 and DXS304, 2 loci mapped probably not more than 900 kb from the IDS locus. A detailed clinical description of the patient is provided and his phenotype is compared to that of other patients with IDS deletion described recently. By following the segregation of a restriction fragment length polymorphism at the IDS locus in the patient's family, our data suggest that the deletion occurred in the germ cells of the patient's grandfather.