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J Schmidtke

Publications and source records attributed to J Schmidtke.

At least 55 records · Page 3Linked to original sources
PubMed1998

Organization and expression of rat Tspy.

We have isolated both a full-length rat Tspy cDNA from testicular mRNA by rapid amplification of cDNA ends (RACE) and RT-PCR and a full-length rat Tspy gene from genomic DNA by PCR. In contrast to the mouse, where Tspy is present in a single copy and is apparently functionless, and to man and cattle, where TSPY is organized in a moderately repetitive cluster, the rat Tspy locus apparently consists of one complete functional and one truncated, probably nonfunctional, copy, coherently localized on Yp, as revealed by FISH analysis.

Animals↗
PubMed1998

A Commentary on the NIH Consensus Development Statement 'Genetic Testing for Cystic Fibrosis'.

A NIH Consensus Development Statement recommends implementation of a cystic fibrosis carrier screening in the general pregnant and pre-pregnant population. This suggestion is discussed in the light of missing or insufficient data about the reasons for high uptake rates in pilot projects, psychological and social risks, and optimal educational and counselling settings. It is concluded that the recommendation is not defendable and, at best, premature.

Journal Article↗
PubMed1998

Mutational analysis of PHEX gene in X-linked hypophosphatemia.

Hypophosphatemic rickets is commonly an X-linked dominant disorder (XLH or HYP) associated with a renal tubular defect in phosphate transport and bone deformities. The XLH gene, referred to as PHEX, or formerly as PEX (phosphate regulating gene with homologies to endopeptidases on the X-chromosome), encodes a 749-amino acid protein that putatively consists of an intracellular, transmembrane, and extracellular domain. PHEX mutations have been observed in XLH patients, and we have undertaken studies to characterize such mutations in 46 unrelated XLH kindreds and 22 unrelated patients with nonfamilial XLH by single stranded conformational polymorphism and DNA sequence analysis. We identified 31 mutations (7 nonsense, 6 deletions, 2 deletional insertions, 1 duplication, 2 insertions, 4 splice site, 8 missense, and 1 within the 5' untranslated region), of which 30 were scattered throughout the putative extracellular domain, together with 6 polymorphisms that had heterozygosity frequencies ranging from less than 1% to 43%. Single stranded conformational polymorphism was found to detect more than 60% of these mutations. Over 20% of the mutations were observed in nonfamilial XLH patients, who represented de novo occurrences of PHEX mutations. The unique point mutation (a-->g) of the 5'untranslated region together with the other mutations indicates that the dominant XLH phenotype is unlikely to be explained by haplo-insufficiency or a dominant negative effect.

Amino Acid Sequence↗
PubMed1998

A new missense substitution at a mutational hot spot of the androgen receptor in siblings with complete androgen insensitivity syndrome.

Several mutations have been described in the human androgen receptor gene including constitutional mutations in androgen insensitivity syndrome, somatic mutations in prostate cancer and triplet expansions in Kennedy's disease (Gottlieb et al. 1997). Here we report on two siblings with complete androgen insensitivity and a novel missense mutation, D695V, in their androgen receptor gene. The two XY females are siblings of German descent and presented at the ages of 23 and 19 years, respectively, with typical clinical features of complete androgen insensitivity. We found both siblings to be hemizygous for a new adenine to thymine transversion at the second nucleotide of codon 695 within the fourth exon of the human androgen receptor gene. The resulting missense mutation D695V is located at the amino-terminal border of the ligand-binding domain of the androgen receptor. The aspartic acid residue at this position is highly conserved in the steroid binding domains of other members of the nuclear receptor family and has already been found to be the site of two other missense mutations associated with androgen insensitivity syndrome (Ris Stalpers et al. 1991, Hiort et al. 1996). Three of four reported subjects showed the complete androgen insensitivity phenotype, in accordance with the two siblings in our study. We suggest that the existence of three pathological amino acid substitutions for aspartic acid 695 most likely reflects the essential role of this residue for normal androgen receptor function in male sexual differentiation.

Adult↗
PubMed1997

A paternity case with apparently conflicting multilocus and single-locus DNA typing results.

A paternity case is presented in which DNA typing with thirteen variable number of tandem repeat (VNTR) systems was in apparent conflict with an initial multilocus DNA analysis using oligonucleotide probe (CAC)5. A dual approach was necessary, because the multilocus analysis yielded a proportion of aberrant nonmaternal offspring bands far too small and level of band-sharing too high for non-paternity. The case could finally be solved by reference to the joint likelihood ratio combining both approaches. Arguments are presented that summation of log likelihoods is indeed valid even if no information is available regarding linkage between the VNTR loci and the loci contributing to a multilocus DNA fingerprint.

DNA↗
PubMed1997

A frequent polymorphism of the gene mutated in ataxia telangiectasia.

The gene mutated in ataxia telangiectasia (AT), the ATM gene, has recently been cloned and shown to encode a multi-domain protein with homology to cell-cycle regulators. We report here an intragenic restriction fragment length polymorphism of the ATM gene that is common in the German population with allele frequencies of 0.56 and 0.44, respectively. Observed heterozygosity approximated 50% in healthy individuals as well as in 152 randomly selected breast cancer patients. We present a simple and rapid polymerase chain reaction (PCR)-based assay for this dimorphism that should be useful to trace the inheritance of ATM alleles in ataxia telangiectasia and breast cancer families and to study loss of heterozygosity in primary tumours.

Alleles↗
PubMed1997

Organization and expression of bovine TSPY.

We have isolated genomic sequences as well as transcripts from the bovine homolog of the human testis-specific protein, Y-encoded, TSPY which-in both species-is located on the Y Chromosome (Chr), organized as a gene family with a variable number of members, and expressed exclusively in the testis. 1266 bp of bovine TSPY specific sequence have been isolated from a testis cDNA library, by RT-PCR analyses and by Rapid Amplification of cDNA Ends (RACE). A bovine TSPY gene 4 is organized in seven exons, and transcripts are polyadenylated at various 3' ends. Consensus polyadenylation signals AAUUAAA are missing. Microheterogeneous sequence variation is found between TSPY family members. In addition, homologies to other Y-located repeated sequence families, BRY, have been discovered; these sequences are presumably derived from ancient members of the TSPY cluster, now forming a separate, probably nonfunctional subfamily. Bovine TSPY is subject to differential splicing. In the adult, it is expressed in early germ-cell stages, and expression could also be detected in fetal testis. Comparison with the human homolog shows the highest degree of similarity in the coding regions of exons 2, 3, and 4, which are also precisely conserved regarding their length.

Age Factors↗
PubMed1997

Possible association of the allele status of the CS.7/HhaI polymorphism 5' of the CFTR gene with postnatal female survival.

Cystic fibrosis (CF) patients show a high degree of linkage disequilibrium between the CF transmembrane conductance regulator (CFTR) gene and polymorphisms 5' of that gene. To determine whether the region 5' of CFTR contains biologically important sequences, the allele frequencies of six CFTR-linked polymorphisms (metH/MspI, XV-2c/TaqI, CS.7/HhaI, KM19/PstI, MP6d9/MspI, J44/XbaI) were determined in 417 randomly selected elderly individuals (over 75 years of age) from the Czech population. The elderly individuals were considered "escapees" of strong selective pressures that had operated during their lifetime, prior to the introduction of modern health care since 1950. The pooled allele frequencies of the analyzed marker polymorphisms in the elderly did not significantly differ from published data. However, when analyzed by ex, the allele frequencies of markers CS.7/HhaI and KM19/PstI differed significantly (P < 0.05) between elderly females and males. The allele frequencies of the six polymorphisms were then determined in 646 newborns and 345 young adults of reproductive age; these individuals were selected in a similar manner and drawn from the same population. In these control groups, the studied marker polymorphisms exhibited no statistically significant differences between sexes and/or between individuals of the same sex, only between different age groups. A gradual relative increase in the frequency of allele "2" of marker CS.7/HhaI was observed from newborn females to elderly women, the overall difference in allele frequencies of this marker polymorphism between newborn females and elderly women reaching statistical significance (P < 0.05). Interestingly, allele "2" is the major constituent of the extended "B-haplotype", which is in strong linkage disequilibrium with common CF alleles. Taken together, our data suggest that the region spanning markers CS.7 and KM19 is associated with a genetic factor that influences postnatal female survival, providing a possible mechanism for increasing the frequency of particular mutations in the adjacent CFTR gene.

Adult↗
PubMed1997

Distinct spectrum of CFTR gene mutations in congenital absence of vas deferens.

Congenital absence of the vas deferens (CAVD) is a frequent cause for obstructive azoospermia and accounts for 1%-2% of male infertility. A high incidence of mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene has recently been reported in males with CAVD. We have investigated a cohort of 106 German patients with congenital bilateral or unilateral absence of the vas deferens for mutations in the coding region, flanking intron regions and promotor sequences of the CFTR gene. Of the CAVD patients, 75% carried CFTR mutations or disease-associated CFTR variants, such as the "5T" allele, on both chromosomes. The distribution of mutation genotypes clearly differed from that observed in cystic fibrosis. None of the CAVD patients was homozygous for delta F508 and none was compound heterozygous for delta F508 and a nonsense or frameshift mutation. Instead, homozygosity was found for a few mild missense or splicing mutations, and the majority of CAVD mutations were missense substitutions. Twenty-one German CAVD patients were compound heterozygous for delta F508 and R117H, which was the most frequent CAVD genotype in our study group. Haplotype analysis indicated a common origin for R117H in our population, whereas another frequent CAVD mutation, viz. the "5T allele" was a recurrent mutation on different intragenic haplotypes and multiple ethnic backgrounds. We identified a total of 46 different mutations and variants, of which 15 mutations have not previously been reported. Thirteen novel missense mutations and one unique amino-acid insertion may be confined to the CAVD phenotype. A few splice or missense variants, such as F508C or 1716 G-->A, are proposed here as possible candidate CAVD mutations with an apparently reduced penetrance. Clinical examination of patients with CFTR mutations on both chromosomes revealed elevated sweat chloride concentrations and discrete symptoms of respiratory disease in a subset of patients. Thus, our collaborative study shows that CAVD without renal malformation is a primary genital form of cystic fibrosis in the vast majority of German patients and links the particular expression of clinical symptoms in CAVD with a distinct subset of CFTR mutation genotypes.

Adult↗
PubMed1997

Conserved Y-chromosomal location of TSPY in Bovidae.

We determined the chromosomal location of TSPY, the testis-specific protein, Y-encoded, by fluorescence in situ hybridization (FISH) to chromosome spreads of cattle, goat and sheep. Using a cloned polymerase chain reaction (PCR) product of one bovine TSPY family member, we were able to show a conserved Y chromosomal localization for TSPY in all three species. In contrast to a limited regional distribution of TSPY FISH signals on the chromosome of man, other primates, great apes, goat and sheep, in cattle TSPY-related sequences appear to be spread over most of the Y chromosome. The painting effect observed in this species reflects the higher complexity of the bovine TSPY gene family, being composed not only of a tandemly repeated cluster, but harbouring a large number of different family members dispersed all over the Y chromosome.

Animals↗
PubMed1997

Detection of 100% of the CFTR mutations in 63 CF families from Tyrol.

We identified 100% of the CFTR gene mutations, including three novel mutations, in 126 unrelated cystic fibrosis chromosomes from Tyrol, Austria. The frequency of the major mutation deltaF508 (74.6%) was not significantly different in Tyrolian CF-patients than in patients from Bavaria (71.0%) and Middle- and Northern Germany (71.9%), but was significantly higher than in patients from Styria (58.1%) or Northern Italy (47.6%). Interestingly, the distribution of the next most frequent mutations, R1162X (8.7%) 2183AA-->G, 2789+5G-->A and G542X (2.4% each), was more similar to the distribution of these mutations among CF-patients from Northern Italy than to those from Styria, Bavaria or Middle- and Northern Germany. Nine further mutations occurred once or twice. One of these, the missense mutation M1101K, is rare worldwide but very frequent in the Hutterite brethren, a small founder population which came from Southern Austria to Northern America. Three other different mutations (deltaL453, 1874insT and 4108delT) were present in single Tyrolian families and have not been described before. The identification of 100% of CFTR gene mutations in a particular CF population demonstrates the power of genetic analysis for the diagnosis and counselling of CF families in this restricted geographical area of Austria. Our study provides evidence for a closer genetic relation between CF patients from Tyrol and those from Bavaria or Middle- and Northern Germany as well as Northern Italy, than between CF patients from the two Austrian states Tyrol and Styria.

Adolescent↗
PubMed1997

Novel and characteristic CFTR mutations in Saudi Arab children with severe cystic fibrosis.

More than 600 different CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations have been identified so far that are considered to cause the fatal genetic disorder cystic fibrosis (CF). We have investigated 15 Arab children from 12 families, who were diagnosed as having CF, for mutations in the coding region and in the flanking intron sequences of the CFTR gene. Six different CFTR mutations were identified including two novel mutations, 1548delG in exon 10 and 406-2A-->G in intron 3. Prominent mutations were the splice mutation 3120 + 1G-->A (intron 16) followed by N1303K (exon 21) and 1548delG (exon 10). Most CF children were homozygotes who presented with a severe form of the disease including failure to thrive, recurrent chest infections, particularly with Pseudomonas aeruginosa, and frequent hospital admissions. Identification of the CFTR mutations facilitates molecular investigation of the disease and better understanding of its pathophysiology in Arab children, among whom CF is probably an underdiagnosed disease.

Child↗
PubMed1996

Geographic distribution and origin of CFTR mutations in Germany.

The geographic distribution and origin of CFTR mutations in Germany was evaluated in 658 three-generation families with cystic fibrosis (CF). Fifty different mutations were detected on 1305 parental CF chromosomes from 22 European countries and overseas. The major mutation. delta F508 was identified on 71.5% of all CF chromosomes, followed by R553X (1.8%), N1303K (1.3%), G542X (1.1%), G551D (0.8%) and R347P (0.8%). According to the grandparents' birthplace, 74% of CF chromosomes had their origin in Germany; the delta F508 percentage was 77%, 75%, 70% and 62% in northern, southern, western and eastern Germany, respectively. Ten or more mutant alleles in the investigated CF gene pool originated from Austria, the Czech Republic, Poland, Russia, Turkey and the Ukraine. This widespread geographic origin of CFTR mutations in today's Germany reflects the many demographic changes and migrations in Central Europe during the 20th century.

Cystic Fibrosis↗
PubMed1996

Testis-specific protein, Y-encoded (TSPY) expression in testicular tissues.

TSPY, the 'testis-specific protein, Y-encoded', is the product of a tandem gene cluster on human proximal Yp. In order to gain insight into the function of this locus, we have analysed (I) the diversity of RNAs transcribed from the cluster, (II) the sequence homology of the deduced TSPY to other proteins, and (iii) its protein properties both in tissue extracts and in tissue sections, using a TSPY-specific antiserum. We have identified a set of distinct TSPY transcripts with diverse exon compositions. We show that TSPY has homology with other human and non-human proteins, including SET and NAP, factors that are suggested to play a role in DNA replication. Protein analysis revealed TSPY to occur mainly in a modified, putatively phosphorylated form. By immunostaining it was detected in distinct subsets of spermatogonia. TSPY was also strongly immunostained in early testicular carcinoma in situ (CIS), while seminomatous tumour cells stained less intensely. The spermatogonial cells of two XY-TFM-females gave a strong immune response. The data presented here point to a phosphorylation-dependent TSPY-function in early spermatogenesis, immediately prior to the spermatogonia-to-spermatocyte transition, and in early testicular tumorigenesis.

Amino Acid Sequence↗
PubMed1995

Somatic spectrum of cancer-associated single basepair substitutions in the TP53 gene is determined mainly by endogenous mechanisms of mutation and by selection.

The spectrum of somatic TP53 single basepair substitutions detected in 955 cancers was compared with that of 2,224 different germline mutations in 279 different human genes (other than TP53), reported as the cause of inherited disease. This comparison reveals that, disregarding a relatively small subset (12%) of TP53 mutations that probably result from the action of exogenous mutagens, both the relative rates and the nearest-neighbor spectra of single basepair substitutions are similar in the two datasets. This spectral resemblance suggests that a substantial proportion of cancer-associated somatic TP53 mutations result from endogenous cellular mechanisms. The likelihood of clinical observation of a particular mutation type differs, however, between tumors and genetic diseases, when the chemical properties of the resulting amino acid substitutions are considered. Together with a sixfold higher observation likelihood for mutations at evolutionarily conserved residues, this finding argues that selection is a critical factor in determining which TP53 mutations are found to be associated with human cancer.

Base Sequence↗
PubMed1995

Transcript analysis of CFTR nonsense mutations in lymphocytes and nasal epithelial cells from cystic fibrosis patients.

The mutational effects at the mRNA level were investigated by RT-PCR analysis of nine different nonsense mutations (Q39X, E60X, R75X, G542X, L719X, Y1092X, R1162X, S1196X, W1282X) and one frameshift mutation (1078delT) within the CFTR gene. With the exception of mutation R1162X, reduced mRNA levels ranging from 30% to less than 5% of the wild type have been observed. In case of the R75X and E60X mutations, the mRNA reduction was accompanied by the appearance of atypical CFTR isoforms. Single exon 3 skipping, as well as joint exon 2 and 3 skipping, was observed in lymphocyte and nasal epithelial mRNA derived from R75X alleles. The analysis of mRNA transcribed from E60X alleles revealed skipping of exon 3 (lymphocytes and nasal epithelial cells) or skipping of exons 3 and 4 (nasal epithelial cells). With the exception of the E60X mutation, no obvious tissue-specific differences in the splicing pattern and ratios of mutation to wild-type transcripts were detected between lymphocytes and nasal epithelial cells. In addition to aberrant splicing, the reduction of transcripts is the most common effect of nonsense and frameshift mutations within the CFTR gene.

Alleles↗
PubMed1995

Direct molecular analysis of the fragile X syndrome in a sample of Egyptian and German patients using non-radioactive PCR and Southern blot followed by chemiluminescent detection.

Molecular genetic analysis of individuals from 6 Egyptian and 33 German families with fragile X syndrome and 240 further patients with mental retardation was performed applying a completely non-radioactive system. The aim of our study was the development of a non-radioactive detection method and its implementation in molecular diagnosis of the fragile X syndrome. Furthermore, we wanted to assess differences in the mutation sizes between Egyptian and German patients and between Egyptian and German carriers of a premutation. Using non-radioactive polymerase chain reaction (PCR), agarose gel electrophoresis and blotting of the PCR products, followed by hybridisation with a digoxigenin-labelled oligonucleotide probe (CGG)5 and chemiluminescent detection, we identified the fragile X full mutation (amplification of a CGG repeat in the FMR-1 gene ranging from several hundred to several thousand repeat units) in all patients. We observed no differences in the length of the CGG repeat between the Egyptian and German patients and carriers, respectively. However, in one prenatal diagnosis, we detected only one normal sized allele in a female fetus using the PCR-agarose assay, whereas Southern blot analysis with the digoxigenin labelled probe StB 12.3 revealed presence of a full mutation. Our newly established nonradioactive genomic blotting method is based on the conventional radioactive Southern blot analysis. Labelling of the probe StB 12.3 with digoxigenin via PCR allowed the detection of normal, premutated and fully mutated alleles. For exact sizing of small premutated or large normal alleles, we separated digoxigenin labelled PCR products through denaturing polyacrylamide gelelectrophoresis (PAGE) and transfered them to a nylon membrane using a gel dryer. The blotted PCR-fragments can easily be detected with alkaline phosphate-labelled anti-digoxigenin antibody. The number of trinucleotide repeat units can be determined by scoring the detected bands against a digoxigenated M13 sequencing ladder. Our newly developed digoxigenin/chemiluminescence approach using PCR and Southern blot analysis provides reliable results for routine detection of full fragile X mutations and premutations.

Base Sequence↗