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B Horsthemke

Publications and source records attributed to B Horsthemke.

At least 73 records · Page 4Linked to original sources
PubMed1997

The tricho-rhino-phalangeal syndromes: frequency and parental origin of 8q deletions.

The tricho-rhino-phalangeal syndromes type I (TRPS I) and type II (TRPS II) result from the deletion of overlapping sets of genes within the Langer-Giedion syndrome chromosomal region (LGCR) on chromosome 8. In contrast to TRPS I patients, most TRPS II patients have cytogenetically visible deletions and are often mentally retarded. Using Southern blot and fluorescence in situ hybridization analysis, we searched for submicroscopic deletions in 12 patients with TRPS I and an apparently normal karyotype. One patient of normal intelligence was found to have a deletion of approximately 5 Mb. This suggests that mental retardation in TRPS is caused by genes outside the 5-Mb region. Using three LGCR microsatellite markers, we determined the parental origin of this TRPS I deletion and of eight TRPS II deletions. In six patients, the deletion was of paternal origin and in three patients it was of maternal origin.

Blotting, Southern↗
PubMed1997

Isolation of DNA from the centromere of human chromosome 7 by microdissection.

Centromeres remain the least characterized regions of human chromosomes because they have a very high content of repetitive DNA. Here, we describe a microdissection library from the centromeric region of human chromosome 7 and its use for generating sequence tagged sites (STSs). The library contains about 1500 clones with an average insert size of 150 bp and only about 15% of the clones harbour repetitive human DNA. Seven clones hybridizing to alphoid DNA were found to correspond to a fragment of the D7Z2 alphoid array on chromosome 7, thus confirming the origin of the library. A number of clones not containing known repetitive DNA were used to generate STSs that identified yeast artificial chromosomes (YACs) and in turn allowed the STSs to be placed on the physical map. One STS is located between the two Genethon genetic markers closest to the centromere on the q side. Another STS was located 3-4cM away in 7q11.2, while a third identified YACs containing both low-copy and alphoid sequences that are not yet mapped but are clearly centromeric. The library therefore comprises a collection of sequences from the centromeric region of chromosome 7 that can be used to generate STSs and to map the entire centromeric region.

Amniotic Fluid↗
PubMed1997

Different mechanisms and recurrence risks of imprinting defects in Angelman syndrome.

Angelman syndrome (AS) is a neurogenetic disorder that appears to be caused by the loss of function of an imprinted gene expressed from maternal chromosome 15 only. Approximately 6% of patients have a paternal imprint on the maternal chromosome. In the few cases, this is due to an inherited microdeletion, in the 15q11-q13 imprinting center (IC), that blocks the paternal-->maternal imprint switch in the maternal germ line. We have determined the segregation of 15q11-q13 haplotypes in nine families with AS and with an imprinting defect. One family, with two affected siblings, has a microdeletion affecting the IC transcript. In the other eight patients, no mutation was found at this locus. In two families, the patient and a healthy sibling share the same maternal alleles. In one of these families and in two others, grandparental DNA samples were available, and the chromosomes with the imprinting defect were found to be of grandmaternal origin. These findings suggest that germ-line mosaicism or de novo mutations account for a significant fraction of imprinting defects, among patients who have an as-yet-undetected mutation in a cis-acting element. Alternatively, these data may indicate that some imprinting defects are caused by a failure to maintain or to reestablish the maternal imprint in the maternal germ line or by a failure to replicate the imprint postzygotically. Depending on the underlying cause of the imprinting defect, different recurrence risks need to be considered.

Angelman Syndrome↗
PubMed1997

Constitutional RB1-gene mutations in patients with isolated unilateral retinoblastoma.

In most patients with isolated unilateral retinoblastoma, tumor development is initiated by somatic inactivation of both alleles of the RB1 gene. However, some of these patients can transmit retinoblastoma predisposition to their offspring. To determine the frequency and nature of constitutional RB1-gene mutations in patients with isolated unilateral retinoblastoma, we analyzed DNA from peripheral blood and from tumor tissue. The analysis of tumors from 54 (71%) of 76 informative patients showed loss of constitutional heterozygosity (LOH) at intragenic loci. Three of 13 uninformative patients had constitutional deletions. For 39 randomly selected tumors, SSCP, hetero-duplex analysis, sequencing, and Southern blot analysis were used to identify mutations. Mutations were detected in 21 (91%) of 23 tumors with LOH. In 6 (38%) of 16 tumors without LOH, one mutation was detected, and in 9 (56%) of the tumors without LOH, both mutations were found. Thus, a total of 45 mutations were identified in tumors of 36 patients. Thirty-nine of the mutations-including 34 small mutations, 2 large structural alterations, and hypermethylation in 3 tumors-were not detected in the corresponding peripheral blood DNA. In 6 (17%) of the 36 patients, a mutation was detected in constitutional DNA, and 1 of these mutations is known to be associated with reduced expressivity. The presence of a constitutional mutation was not associated with an early age at treatment. In 1 patient, somatic mosaicism was demonstrated by molecular analysis of DNA and RNA from peripheral blood. In 2 patients without a detectable mutation in peripheral blood, mosaicism was suggested because 1 of the patients showed multifocal tumors and the other later developed bilateral retinoblastoma. In conclusion, our results emphasize that the manifestation and transmissibility of retinoblastoma depend on the nature of the first mutation, its time in development, and the number and types of cells that are affected.

Actuarial Analysis↗
PubMed1997

Balanced translocation 46,XY,t(2;15)(q37.2;q11.2) associated with atypical Prader-Willi syndrome.

The lack of normally active paternal genes in 15q11-q13, as an outcome of either a paternal deletion or maternal disomy, accounts for >95% of all patients with Prader-Willi syndrome. Other mechanisms, including imprinting mutations and unbalanced translocations involving pat 15q11-q13, have been described elsewhere. In this study, we present a patient with a rare balanced, de novo translocation-46,XY,t(2;15)(q37.2;q11.2)-involving breakage within the Prader-Willi/Angelman syndrome region of the paternal homologue, without an apparent deletion. The patient demonstrated several manifestations of the Prader-Willi syndrome but was clinically atypical. Cytogenetic and molecular studies of this case demonstrated the translocation breakpoint to be between SNRPN and IPW, with mRNA expression of SNRPN and PAR-5 but absence of IPW and PAR-1 expression. These results suggest that disruption of either IPW expression or a nearby gene by an upstream break may contribute to the Prader-Willi syndrome phenotype and that expression of SNRPN or other upstream genes is responsible for other aspects of the classical Prader-Willi syndrome phenotype.

Animals↗
PubMed1997

Imprinted segments in the human genome: different DNA methylation patterns in the Prader-Willi/Angelman syndrome region as determined by the genomic sequencing method.

A deletion of 15q11-q13 and uniparental disomy 15 lead to Prader-Labhart-Willi syndrome (PWS) or Angelman syndrome (AS) because this region contains genes expressed exclusively from the paternal (PWS) or maternal (AS) chromosome, respectively. DNA methylation plays a role in the control of imprinted gene expression, but so far only a few 5'-CG-3' dinucleotides within the recognition sites of the methylation sensitive enzymes have been studied. As part of a study on DNA methylation patterns in the human genome, we have applied the bisulfite protocol of genomic sequencing to study all 5'-CG-3' dinucleotides around exon 1 of SNRPN and at the D15S63 locus, which contains a start site for alternative SNRPN transcripts possibly involved in imprint switching during gametogenesis. At least 17 PCR products derived from single chromosomes of normal individuals as well as PWS and AS patients have been sequenced. We have found that cytosine residues outside 5'-CG-3' dinucleotides are always unmethylated. However, > 96% of all of the 23 5'-CG-3' dinucleotides around SNRPN exon 1 are methylated on the maternal chromosome and completely devoid of methylation on the paternal chromosome. This finding is in contrast to the D15S63 locus, where only the two Cfol/Hhal sites are methylated on the maternal chromosome at the same frequency as seen for the SNRPN segment. At the other five 5'-CG-3' dinucleotides, differential methylation is less pronounced, i.e. 45-70% on the maternal chromosome and 5-14% on the paternal chromosome. The differences between SNRPN and D15S63 methylation may reflect different biological functions of the alternative SNRPN transcripts. The systematic investigation of 5'-CG-3' methylation patterns as reported here will provide the basis for a PCR-based methylation test to diagnose PWS and AS.

5-Methylcytosine↗
PubMed1997

The murine Ext1 gene shows a high level of sequence similarity with its human homologue and is part of a conserved linkage group on chromosome 15.

We have cloned and sequenced the murine homologue of the human EXT1 gene. At the protein level, these genes show almost complete identity as divergence is limited to only 5 amino acid positions that are scattered about the whole sequence. In addition, similarity searches identified a protein from chromosome III of C. elegans that shows significant similarity to the human and murine EXT/Ext genes. Using high resolution backcross mapping, the murine Ext1 was mapped at 26.55 cM between D15Mit143 and D15Mit153 on mouse chromosome 15. Therefore, Ext1 is part of an evolutionarily conserved linkage group including SDC2/Hspg1, TRHR/Trhr, EXT1/Ext1, MYC/Myc, and TG/Tgn.

Amino Acid Sequence↗
PubMed1996

Validation studies of SNRPN methylation as a diagnostic test for Prader-Willi syndrome.

Prader-Willi syndrome (PWS) is caused by absence of a paternal contribution of the chromosome region 15q11-q13, resulting from paternal deletions, maternal uniparental disomy, or rare imprinting mutations. Laboratory diagnosis is currently performed using fluorescence in situ hybridization (FISH), DNA polymorphism (microsatellite) analysis, or DNA methylation analysis at locus PW71 (D15S63). We examined another parent-of-origin-specific DNA methylation assay at exon alpha of the small nuclear ribonucleoprotein-associated polypeptide N gene (SNRPN) in patients referred with clinical suspicion of PWS or Angelman syndrome (AS). These included 30 PWS and 17 AS patients with known deletion or uniparental disomy status, and a larger cohort of patients (n = 512) suspected of PWS who had been analyzed previously for their methylation status at the PW71 locus. Results of SNRPN methylation were consistent with known deletion or uniparental disomy (UPD) status as determined by other molecular methods in all 47 cases of PWS and AS. In the larger cohort of possible PWS patients, SNRPN results were consistent with clinical diagnosis by examination and with PW71 methylation results in all cases. These data provide support for the use of SNRPN methylation as a diagnostic method. Because methylation analysis can detect all three major classes of genetic defects associated with PWS (deletion, UPD, or imprinting mutations), methylation analysis with either PW71 or SNRPN is an efficient primary screening test to rule out a diagnosis of PWS. Only patients with an abnormal methylation result require further diagnostic investigation by FISH or DNA polymorphism analysis to distinguish among the three classes for accurate genetic counseling and recurrence-risk assessment.

Autoantigens↗
PubMed1996

Minimal definition of the imprinting center and fixation of chromosome 15q11-q13 epigenotype by imprinting mutations.

Patients with disorders involving imprinted genes such as Angelman syndrome (AS) and Prader-Willi syndrome (PWS) can have a mutation in the imprinting mechanism. Previously, we identified an imprinting center (IC) within chromosome 15q11-ql3 and proposed that IC mutations block resetting of the imprint, fixing on that chromosome the parental imprint (epigenotype) on which the mutation arose. We now describe four new microdeletions of the IC, the smallest (6 kb) of which currently defines the minimal region sufficient to confer an AS imprinting mutation. The AS deletions all overlap this minimal region, centromeric to the PWS microdeletions, which include the first exon of the SNRPN gene. None of five genes or transcripts in the 1.0 Mb vicinity of the IC (ZNF127, SNRPN, PAR-5, IPW, and PAR-1), each normally expressed only from the paternal allele, was expressed in cells from PWS imprinting mutation patients. In contrast, AS imprinting mutation patients show biparental expression of SNRPN and IPW but must lack expression of the putative AS gene 250-1000 kb distal of the IC. These data strongly support a model in which the paternal chromosome of these PWS patients carries an ancestral maternal epigenotype, and the maternal chromosome of these AS patients carries an ancestral paternal epigenotype. The IC therefore functions to reset the maternal and paternal imprints throughout a 2-Mb imprinted domain within human chromosome 15q11-q13 during gametogenesis.

Angelman Syndrome↗
PubMed1996

Prognostic implications of monosomy 3 in uveal melanoma.

BACKGROUND: A high proportion of patients with uveal melanoma die of metastatic disease. In a subgroup of uveal melanomas there is the loss of one chromosome 3. To assess the prognostic implications of this genetic anomaly, we studied 54 patients for a median of 3.4 years. METHODS: 180 patients underwent primary enucleation for malignant uveal melanoma at the Ophthalmology Department of the Universitätsklinikum Essen between 1987 and 1993. Tumour material was available for chromosome analysis and DNA preparation from 69 of these patients (for logistic reasons unlikely, we believe, to introduce bias). 15 patients were excluded from our study: nine because the methods for assessment of monosomy 3 were unsuccessful; five because of insufficient information about their relapse status; one because histopathological data were incomplete. Of the 54 remaining patients, the tumours of 16 were assessed for copy number of chromosome 3 by karyotype analysis, of 30 by comparative genomic hybridisation, and of eight by both techniques. Clinical status was assessed by contact with family doctor or a clinical check up. Statistical analysis was by the log-rank test and Cox proportional-hazard regression. FINDINGS: The tumours of 30 patients had monosomy 3. 17 (57%) of these patients relapsed with metastatic disease, and the 3-year relapse-free survival rate was 50%. By contrast, of the 24 patients whose tumours had retained both chromosomes 3, none developed metastatic disease. In univariate analysis monosomy 3 was the most significant (p < 0.0001) predictor of poor prognosis in uveal melanoma, followed by tumour location (p < 0.0007) and tumour diameter (p < 0.0021). Histopathological subtype, age, sex, extrascleral growth, and tumour thickness had no additional predictive value. INTERPRETATION: In uveal melanoma, monosomy 3 is a significant predictor of both relapse-free and overall survival.

Aged↗
PubMed1996

Cytogenetic and age-dependent risk factors associated with uniparental disomy 15.

Prader-Willi syndrome (PWS) results primarily from either a paternal deletion of 15q11-q13 or maternal uniparental disomy (UPD) of chromosome 15. Including the present and published cases, more than 120 patients with maternal UPD of human chromosome 15 have been ascertained. Investigation of chromosome 15 markers indicates that approximately 71 per cent of the additional maternal chromosomes were the result of meiosis I segregation errors, 13 per cent were the result of meiosis II errors, and 16 per cent resulted from post-zygotic duplication of one chromosome 15. An increase in maternal age is associated with UPD cases due to meiotic errors. The age-specific risk for UPD(15) is analysed and shows an exponential increase with maternal age which is similar to that observed for trisomy 21. For women greater than or equal to 40 years of age, the risk for UPD(15) is approximately 1/3400 livebirths. The frequency of chromosome aberrations associated with UPD(15) is also discussed. Two types of aberrations are at significantly increased risk of fetal UPD(15): de novo (or inherited) isochromosome 15 and confined placental mosaicism for trisomy 15. Two additional abnormalities, de novo small marker chromosomes derived from 15, e.g., idic15(pter-q11:q11-pter), and familial Robertsonian translocations involving chromosome 15, appear to have a mildly increased risk of UPD(15).

Adult↗
PubMed1996

Imprint switching on human chromosome 15 may involve alternative transcripts of the SNRPN gene.

Imprinting on human chromosome 15 is regulated by an imprinting centre, which has been mapped to a 100-kb region including exon 1 of SNRPN. From this region we have identified novel transcripts, which represent alternative transcripts of the SNRPN gene. The novel exons lack protein coding potential and are expressed from the paternal chromosome only. We have also identified intragenic deletions and a point mutation in patients who have Angelman or Prader-Willi syndrome due to a parental imprint switch failure. This suggests that imprint switching on human chromosome 15 may involve alternative SNRPN transcripts.

Alternative Splicing↗
PubMed1996

Familial translocations involving 15q11-q13 can give rise to interstitial deletions causing Prader-Willi or Angelman syndrome.

A de novo interstitial deletion of 15q11-q13 is the major cause of Prader-Willi syndrome (PWS) and Angelman syndrome (AS). Here we describe two unrelated PWS patients with a typical deletion, whose fathers have a balanced translocation involving the PWS/AS region. Microsatellite data suggest that the deletion is the result of an unequal crossover between the derivative chromosome 15 and the normal chromosome 15. We conclude that familial translocations involving 15q11-q13 can give rise to interstitial deletions causing PWS or AS and that prenatal diagnosis in such families should include fluorescence in situ hybridisation or microsatellite studies or both.

Angelman Syndrome↗
PubMed1996

A pseudogene for the human ribosomal protein L5 (RPL5P1) maps within an intron of the SNRPN transcription unit on human chromosome 15.

The ribosomal protein genes are members of multigene families, most of which are processed pseudogenes. Here we report that an intronless pseudogene for the L5 ribosomal protein maps within an intron of the SNRPN transcription unit on human chromosome 15. The pseudogene (RPL5P1) shares 95% sequence homology with the L5 mRNA, has a poly(A) stretch and is flanked by direct repeats.

Autoantigens↗
PubMed1996

The spectrum of RB1 germ-line mutations in hereditary retinoblastoma.

We have searched for germ-line RB1 mutations in 119 patients with hereditary retinoblastoma. Previous investigations by Southern blot hybridization and PCR fragment-length analysis had revealed mutations in 48 patients. Here we report on the analysis of the remaining 71 patients. By applying heteroduplex analysis, nonisotopic SSCP, and direct sequencing, we detected germ-line mutations resulting in premature termination codons or disruption of splice signals in 51 (72%) of the 71 patients. Four patients also showed rare sequence variants. No region of the RB1 gene was preferentially involved in single base substitutions. Recurrent transitions were observed at most of the 14 codons within the RB1. No mutation was observed in exons 25-27, although this region contains two CGA codons. This suggests that mutations within the 3'-terminal region of the RB1 gene may not be oncogenic. When these data were combined with the results of our previous investigations, mutations were identified in a total of 99 (83%) of 119 patients. The spectrum comprises 15% large deletions, 26% small length alterations, and 42 % base substitutions. No correlation between the location of frameshift or nonsense mutations and phenotypic features, including age at diagnosis, the number of tumor foci, and manifestation of nonocular tumors was observed.

Adolescent↗