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Biomedical subjects

M Schalling

Publications and source records attributed to M Schalling.

At least 91 records · Page 5Linked to original sources
PubMed1996

Clinical implications of unstable DNA repeat sequences.

In this article we review the clinical and genetic features characteristic of a number of diseases recently explained by a novel genetic mechanism: unstable segments of the genome containing trinucleotide repeat sequences. Disorders identified to date are mostly progressive, and display unusual inheritance patterns such as anticipation. Anticipation is manifested as an earlier age at onset or a more severe phenotype in later generations of a family, and can be correlated to an increased repeat expansion size. Thus in later generations the disease onset can take place in childhood whereas affected individuals in earlier generations had only adult symptoms. Paediatric cases of typically adult disorders have been shown to be caused by exceptionally long repeat sequences. Anticipation has been observed in a number of disorders not yet identified at the molecular level. Such disorders could be caused by repeat expansions, and are presently subject to intense research efforts. If repeat sequence expansions are related to these disorders, the longest expansions should be seen in the childhood cases, making these the optimal cases to study. Various DNA-based methods have been developed for the detection of these mutations, making possible preclinical and prenatal diagnostics as well as detection of novel expansions.

Adult↗
PubMed1996

Inhibitory activity of the equine infectious anemia virus major 5' splice site in the absence of Rev.

The major 5' splice site of equine infectious anemia virus (EIAV) conforms to the consensus 5' splice site in eight consecutive positions and is located immediately upstream of the gag AUG. Our results show that the presence of this 5' splice site on the EIAV gag mRNA decreases Gag production 30- to 60-fold. This is caused by inefficient nuclear mRNA export and inefficient mRNA utilization. Inhibition could be overcome by providing human immunodeficiency virus type 1 Rev/Rev-responsive element, human T-cell leukemia virus type 1 Rex/Rex-responsive element, or simian retrovirus type 1 constitutive transport element. In addition, inhibition could be abolished by introducing single point mutations in the 5' splice site or by moving the 5' splice site away from its natural position immediately upstream of the gag AUG. This demonstrates that both maintenance of a perfect consensus 5' splice site and its proper location on the mRNA are important for inhibitory activity of the EIAV major 5' splice site.

Base Sequence↗
PubMed1996

Increased amounts of a high molecular weight insulin-like growth factor II (IGF-II) peptide and IGF-II messenger ribonucleic acid in pancreatic islets of diabetic Goto-Kakizaki rats.

Insulin-like growth factor II (IGF-II), a member of the insulin family, regulates cell growth and differentiation. The IGF-II gene is localized close to the insulin gene in man and rat. IGF-II peptide binds weakly to the insulin receptor and exerts insulin-like effects on the blood glucose level. We studied IGF-II in endocrine pancreas in an animal model of noninsulin-dependent diabetes mellitus, the Goto-Kakizaki (GK) rat. At the age of 2 months, these rats have structural islet changes, with fibrosis and irregular configuration, so-called starfish-shaped islets. Immunohistochemical investigation revealed IGF-II immunoreactivity in the beta-cells in both GK and control rats. Pancreatic extraction, followed by size separation using gel chromatography, disclosed a high mol wt form of IGF-II in all animals, and RIA measurements revealed a considerably larger amount of the IGF-II peptide in the 2-and 6-month-old GK rats than in the 1-month GK and control rats. In situ hybridization of 3-month-old GK rats showed increased IGF-II messenger RNA expression in the starfish-shaped islets of GK rats than in the islets with normal structure in both diabetic and control animals. The reason for the increased amount of IGF-II is unclear. As the animals are diabetic before the islet changes occur, it might be a compensatory effect in response to hyperglycemia, but could also be a cause of the islet fibrosis.

Animals↗
PubMed1996

Mutation detection in Machado-Joseph disease using repeat expansion detection.

BACKGROUND: Several neurological disorders have recently been explained through the discovery of expanded DNA repeat sequences. Among these is Machado-Joseph disease, one of the most common spinocerebellar ataxias (MJD/SCA3), caused by a CAG repeat expansion on chromosome 14. A useful way of detecting repeat sequence mutations is offered by the repeat expansion detection method (RED), in which a thermostable ligase is used to detect repeat expansions directly from genomic DNA. We have used RED to detect CAG expansions in families with either MJD/SCA3 or with previously uncharacterized spinocerebellar ataxia (SCA). MATERIALS AND METHODS: Five MJD/SCA3 families and one SCA family where linkage to SCA1-5 had been excluded were analyzed by RED and polymerase chain reaction (PCR). RESULTS: An expansion represented by RED products of 180-270 bp segregated with MJD/SCA3 (p < 0.00001) in five families (n = 60) and PCR products corresponding to 66-80 repeat copies were observed in all affected individuals. We also detected a 210-bp RED product segregating with disease (p < 0.01) in a non-SCA1-5 family (n = 16), suggesting involvement of a CAG expansion in the pathophysiology. PCR analysis subsequently revealed an elongated MJD/SCA3 allele in all affected family members. CONCLUSIONS: RED products detected in Machado-Joseph disease families correlated with elongated PCR products at the MJD/SCA3 locus. We demonstrate the added usefulness of RED in detecting repeat expansions in disorders where linkage is complicated by phenotyping problems in gradually developing adult-onset disorders, as in the non-SCA1-5 family examined. The RED method is informative without any knowledge of flanking sequences. This is particularly useful when studying diseases where the mutated gene is unknown. We conclude that RED is a reliable method for analyzing expanded repeat sequences in the genome.

Alleles↗
PubMed1996

Developmental expression of splicing variants of fibroblast growth factor receptor 3 (FGFR3) in mouse.

A characteristic feature of the fibroblast growth factor receptor (FGFR) family is the structural diversity generated by alternative splicing. The FGFR3 gene encodes two splice variants because of the mutually exclusive use of the exons IIIb and IIIc. In the present study we examined the expression of the two different splice forms IIIb and IIIc of FGFR3 in developing mouse embryos (12 days p.c., 14 days p.c., 20 days p.c.). The overall level of the IIIc exon splice product surpassed that of the IIIb exon form. The IIIc mRNA was detected in the developing brain and in the spinal cord. Outside the nervous system very strong expression was observed in the vertebra and in all other bony structures. In contrast, the IIIb splice form was restricted to epithelial structures with no expression detected in the central nervous system and bone.

Alternative Splicing↗
PubMed1995

A role for a new herpes virus (KSHV) in different forms of Kaposi's sarcoma.

Kaposi's sarcoma (KS) is a previously rare, tumour-like lesion of controversial biological nature. KS has since the early 1980s become frequent in patients with AIDS, particularly in homosexuals. KS is also endemic in Central Africa predominantly in otherwise healthy men but also in women and children. Recently, evidence for the presence of novel, herpes virus DNA sequences in more than 90% of AIDS Kaposi lesions (AKS) was presented. This DNA was identified using representational difference analysis (RDA) generating short, unique sequences with variable homology to several herpes virus, but no intact virus was recovered. If these DNA-sequences are also present in other, non-HIV-associated forms of Kaposi's sarcoma this would strongly suggest a specific, aetiopathological involvement of this putative new herpes virus in the pathogenesis of Kaposi's sarcoma, rather than a contamination of yet another opportunistic virus in immunosuppressed AIDS patients.

Acquired Immunodeficiency Syndrome↗
PubMed1995

Overexpression of the obese (ob) gene in adipose tissue of human obese subjects.

Obesity is accompanied by complications such as hypertension, non-insulin-dependent diabetes mellitus and atherosclerosis, which in turn cause ischaemic heart disease, stroke and premature death. The underlying mechanisms behind imbalance in energy intake and energy expenditure that lead to obesity are still controversial. In most populations, obesity is more common among women than men and is a multifactorial phenotype, which may result from a complex network of genetic and nongenetic factors. The relative importance of genetic factors for obesity is under debate. Genome searches using polymorphic markers in inbred mice with phenotypes that result in extreme obesity and studies of human candidate genes are being performed in an attempt to identify genes that contribute to obesity. There is evidence that body weight is physiologically regulated and it has been postulated that the storage of fat may provide signals to the brain that the body is obese, which in turn may make the subject eat less and burn more fuel. One of the molecules that may be involved in such signalling is the obese (ob) gene product. Mutations in ob result in profound obesity and type II diabetes in mice. The mouse ob gene and its human homologue have been cloned and sequenced. The gene is expressed in adipose tissue and the product has features of a secreted protein. We have investigated human ob expression in subcutaneous and omental adipose tissue obtained from non-obese and massively obese subjects using in situ hybridization histochemistry and report on overexpression in obese people.

Adipose Tissue↗
PubMed1995

Foci of trinucleotide repeat transcripts in nuclei of myotonic dystrophy cells and tissues.

We have analyzed the intracellular localization of transcripts from the myotonin protein kinase (Mt-PK) gene in fibroblasts and muscle biopsies from myotonic dystrophy patients and normal controls. In affected individuals, a trinucleotide expansion in the gene results in the phenotype, the severity of which is proportional to the repeat length. A fluorochrome-conjugated probe (10 repeats of CAG) hybridized specifically to this expanded repeat. Mt-PK transcripts containing CTG repeat expansions were detected in the nucleus as bright foci in DM patient fibroblasts and muscle biopsies, but not from normal individuals. These foci represented transcripts from the Mt-PK gene since they simultaneously hybridized to fluorochrome-conjugated probes to the 5'-end of the Mt-PK mRNA. A single oligonucleotide probe to the repeat and the sense strand each conjugated to different fluorochromes revealed the gene and the transcripts simultaneously, and indicated that these focal concentrations (up to 13 per nucleus) represented predominately posttranscriptional RNA since only a single focus contained both the DNA and the RNA. This concentration of nuclear transcripts was diagnostic of the affected state, and may represent aberrant processing of the RNA.

Adult↗
PubMed1995

No evidence for association of familial Parkinson's disease with CAG repeat expansion.

In some kindreds, familial Parkinson's disease (PD) exhibits genetic anticipation. Thus, we postulated that familial PD in certain kindreds may be associated with a CAG repeat expansion. However, using the repeat expansion detection method, we found no significant increase in the frequency of CAG repeat expansion among 46 unrelated PD probands compared with controls. Nor did we find evidence for CAG repeat expansion between generations in 11 different PD families that exhibit anticipation in age at onset.

Adult↗
PubMed1995

Familial periodic cerebellar ataxia without myokymia maps to a 19-cM region on 19p13.

Familial periodic cerebellar ataxia (FPCA) is a heterogeneous group of rare autosomal dominant disorders characterized by episodic cerebellar disturbance. A potassium-channel gene (KCNA1) has been found to be responsible for one of its subgroups, familial periodic cerebellar ataxia with myokymia (FPCA/+M; MIM 160120). A different subgroup that is not associated with myokymia (FPCA/-M; MIM 108500) was recently mapped to chromosome 19p. Here we have performed linkage analysis in two large families with FPCA/-M that also demonstrated neurodegenerative pathology of the cerebellum. Three markers in 19p13 gave significant lod scores (> 3.0), while linkage to KCNA1 and three known loci for spinocerebellar ataxia (SCA1, SCA2, and SCA3) was excluded. The highest lod score was obtained with the marker D19S413 (4.4 at recombination fraction 0), and identification of meiotic recombinants in affected individuals placed the locus between the flanking markers D19S406 and D19S226, narrowing the interval to 19 cM. A CAG trinucleotide-repeat expansion was detected in one family but did not cosegregate with the disease.

Cerebellar Ataxia↗
PubMed1994

Rapid and efficient construction of yeast artificial chromosome contigs in the mouse genome with interspersed repetitive sequence PCR (IRS-PCR): generation of a 5-cM, > 5 megabase contig on mouse chromosome 1.

We have developed a new technique for the generation of YAC contigs in the mouse genome that is based on the ability to detect overlapping clones by hybridization of shared IRS-PCR products. As a demonstration of the technique, a 5-cM, > 5 megabase contig was developed on the distal half of mouse Chromosome (Chr) 1, spanning the region from Lamb2 to At3. The contig covers roughly 5% of the genetic distance of the chromosome and is comprised of more than 80 clones; 71 probes were assigned physical order to the chromosome, of which 59 were new markers generated in this study. Eight of the new probes were shown to be polymorphic between C3H/HeJ-gld and M. spretus. Three probes were mapped on a [(C3H/HeJ-gld x M. spretus) x C3H/HeJ-gld] interspecific backcross to integrate the physical map with a high-resolution genetic map of the region.

Animals↗
PubMed1993

Gtx: a novel murine homeobox-containing gene, expressed specifically in glial cells of the brain and germ cells of testis, has a transcriptional repressor activity in vitro for a serum-inducible promoter.

Although it is likely that a highly complex network of transcription factors acts in concert during mammalian brain development, relatively few such genes have been characterized to date. We describe here a novel murine homeobox gene, denoted Gtx, which in adult animals is specifically expressed within glial cells of the central nervous system, including the forebrain, and in germ cells of the testis. Gtx resides on chromosome 7 and does not cosegregate with any previously mapped homeobox gene. The amino acid sequence of the predicted protein encoded by Gtx is highly divergent from that of any other known homeobox genes. The Gtx homeodomain contains unique residues at positions predicted to contact DNA bases. It did not bind to known target sites for other homeobox genes in vitro but bound with high affinity to the MEF-2 motif, a binding site for the serum response factor-related proteins. GTX efficiently competed with RSRF to bind the MEF-2 element in vitro. Co-transfection of Gtx prevented the serum-induced activation of the MEF-2-containing reporter genes. Although the true biological role of Gtx is not known, these results suggest that Gtx is a novel cell-type specific homeobox gene that has the potential to act as a transcriptional repressor for a subset of serum-inducible genes.

Amino Acid Sequence↗
PubMed1993

Tissue specific expression of FMR-1 provides evidence for a functional role in fragile X syndrome.

We have performed mRNA in situ hybridization studies and northern blot analysis in the mouse and human, respectively, to determine the normal gene expression patterns of FMR-1. Expression in the adult mouse was localized to several regions of the brain and the tubules of the testes, which are two of the major organs affected in fragile X syndrome. Universal and very strong expression was observed in early mouse embryos, with differentially decreasing expression during subsequent stages of embryonic development. The early embryonic onset and tissue specificity of FMR-1 gene expression is consistent with involvement in the fragile X phenotype, and also suggests additional organ systems in which clinical manifestations of reduced FMR-1 gene expression may occur.

Adult↗
PubMed1993

Direct detection of novel expanded trinucleotide repeats in the human genome.

Expansion of trinucleotide repeats can give rise to genetic disease. We have developed a technique, repeat expansion detection (RED), that can identify potentially pathological repeat expansion without prior knowledge of chromosomal location. Human genomic DNA is used as a template for a two-step cycling process that generates oligonucleotide multimers when expanded trinucleotide sequences are present at the level found in myotonic dystrophy and fragile-X patients. We have identified at least one new locus exhibiting trinucleotide expansion. Analysis of three families transmitting a long CTG repeat shows that the allele in these families corresponds to a locus on chromosome 18. RED constitutes a powerful tool to identify other diseases caused by this mechanism, particularly diseases associated with anticipation.

Chromosomes, Human, Pair 18↗
PubMed1993

The Wilms tumour gene WT1 is expressed in murine mesoderm-derived tissues and mutated in a human mesothelioma.

The tumour suppressor gene WT1 encodes a transcription factor expressed in tissues of the genito-urinary system. Inactivation of this gene is associated with the development of Wilms tumour a pediatric kidney cancer. We show that WT1 is also expressed at high levels in many supportive structures of mesodermal origin in the mouse. We also describe a case of adult human mesothelioma, a tumour derived from the peritoneal lining, that contains a homozygous point mutation within WT1. This mutation, within the putative transactivation domain, converts the protein from a transcriptional repressor of its target sequence to a transcriptional activator. The role of WT1 in normal development thus extends to diverse structures derived from embryonic mesoderm and disruption of WT1 function contributes to the onset of adult, as well as pediatric, tumours.

Adult↗
PubMed1993

Mouse beta-globin DNA-binding protein B1 is identical to a proto-oncogene, the transcription factor Spi-1/PU.1, and is restricted in expression to hematopoietic cells and the testis.

The hematopoietic-specific DNA-binding protein B1 binds to the DNA consensus sequence AAAGRGGAARYG located twice in intervening sequence 2 of both of the mouse beta-globin genes (D. L. Galson and D.E. Housman, Mol. Cell. Biol. 8:381-392, 1988). B1 was cloned by expression of a murine erythroleukemia (MEL) cell cDNA library in transfected COS cells and screening by electrophoretic mobility shift analysis. B1 is identical to the proto-oncogene Spi-1/PU.1 (Spi-1), an ets family member. Protein-DNA contacts are shown to resemble those of the helix-turn-helix homeodomain proteins. By Northern (RNA) analysis, we found that Spi-1 mRNA is present at low levels during murine CFU-E maturation and is at least 20-fold higher in uninduced MEL, a transformed proerythroblast-like cell line which contains an activating/transforming insertion of spleen focus-forming virus at the Spi-1 locus. Dimethyl sulfoxide-induced MEL cell differentiation decreases Spi-1 mRNA to approximately 20% of the uninduced level before commitment occurs. In addition to erythroid cells, Spi-1 mRNA is present in B cells, myelomonocytes, and mast cells but not in T cells and nonhematopoietic cell types. In situ hybridization demonstrated Spi-1 mRNA expression in bone marrow, spleen, interstitial nonhepatocytes of the liver, and interstitial nontubular cells of the testis. The Spi-1 locus was mapped on human chromosome 11 to the same interval as ACP2 (lysosomal acid phosphatase), between the anonymous DNA markers D11S33 and D11S14. This region has not yet been found to be associated with a human malignancy.

Amino Acid Sequence↗