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Biomedical subjects

S Tomatsu

Publications and source records attributed to S Tomatsu.

At least 55 records · Page 3Linked to original sources
PubMed1997

Mucopolysaccharidosis IVA: a novel splice acceptor site mutation in intron 4 of the N-acetylgalactosamine-6-sulfate sulfatase gene in an Afghanistan girl with classical Morquio disease.

We report here a novel splice site mutation in intron 4 of the gene for N-acetylgalactosamine-6-sulfate sulfatase (GALNS) in an Afghanistan girl with severe mucopolysaccharidosis IVA (classical Morquio disease). Direct sequencing revealed a homozygous G to A transition in the conserved splice acceptor site in intron 4 (cagG-->caaG: designated IVS 4(-I) G-->A) which eliminates 144 nucleotides of exon 5 in her GALNS transcript and introduces an immediate premature termination codon (at Trp 141 of exon 4). The IVS 4(-1) G-->A has not been seen in other populations and this is the first report of the molecular basis of classical Morquio disease in an Afghanistan patient.

Afghanistan↗
PubMed1997

A novel common missense mutation G301C in the N-acetylgalactosamine-6-sulfate sulfatase gene in mucopolysaccharidosis IVA.

Mucopolysaccharidosis IVA (MPS IVA) is an autosomal recessive lysosomal storage disorder caused by a genetic defect in N-acetylgalactosamine-6-sulfate sulfatase (GALNS). In previous studies, we have found two common mutations in Caucasians and Japanese, respectively. To characterize the mutational spectrum in various ethnic groups, mutations in the GALNS gene in Colombian MPS IVA patients were investigated, and genetic backgrounds were extensively analyzed to identify racial origin, based on mitochondrial DNA (mtDNA) lineages. Three novel missense mutations never identified previously in other populations and found in 16 out of 19 Colombian MPS IVA unrelated alleles account for 84.2% of the alleles in this study. The G301C and S162F mutations account for 68.4% and 10.5% of mutations, respectively, whereas the remaining F69V is limited to a single allele. The skewed prevalence of G301C in only Colombian patients and haplotype analysis by restriction fragment length polymorphisms in the GALNS gene suggest that G301C originated from a common ancestor. Investigation of the genetic background by means of mtDNA lineages indicate that all our patients are probably of native American descent.

Asian People↗
PubMed1996

Heteroallelic missense mutations of the galactosamine-6-sulfate sulfatase (GALNS) gene in a mild form of Morquio disease (MPS IVA).

Morquio disease (MPS IVA) is an autosomal recessive disorder caused by a deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS) activity. Patients commonly present in early infancy with growth failure, spondyloepiphyseal dysplasia, corneal opacification, and keratan sulfaturia, but milder forms have been described. We report on a patient who grew normally until age 5 years. Her keratan sulfaturia was not detected until adolescence, and she now has changes restricted largely to the axial skeleton. She has experienced only mildly impaired vision. At age 22, thin-layer chromatography of purified glycosaminoglycans showed some keratan sulfaturia. GALNS activity in fibroblast homogenate supernatants was 20 +/- 5% of controls (as compared to 5 +/- 3% of controls in severe MPS IVA, P < .003). Kinetic analysis of residual fibroblast GALNS activity in patient and parents revealed decreased K(m) and increased Vmax in the mother and daughter, but not in the father, compatible with compound heterozygosity. GALNS exons were amplified from patient genomic DNA and screened by SSCP. Two missense mutations, a C to T transition at position 335 (predicting R94C) and a T to G transversion at position 344 (predicting F97V), were found on sequencing an abnormally migrating exon 3 amplicon. Digestion of the amplicon with FokI and AccI restriction enzymes (specific for the R94C and F97V mutations, respectively) confirmed heterozygosity. In fibroblast transfection experiments, heterozygous R94C and F97V mutants independently expressed as severe and mild GALNS deficiency, respectively. We interpret these findings to indicate that our patient bears heteroallelic GALNS missense mutations, leading to GALNS deficiency and mild MPS IVA. Our findings expand the clinical and biochemical phenotype of MPS IVA, but full delineation of the genotype-phenotype relationship requires further study of native and transfected mutant cell lines.

Adolescent↗
PubMed1996

Mucopolysaccharidosis type I: identification of common mutations that cause Hurler and Scheie syndromes in Japanese populations.

alpha-L-Iduronidase (IDUA) deficiency (mucopolysaccharidosis type I; MPS-I) is an inborn error of lysosomal degradation of glycosaminoglycans that results in storage of undegraded glycosaminoglycans in lysosomes. Previous studies in Caucasian populations showed that (1) homozygosity or compound heterozygosity for the W402X and Q70X mutations are the common causes of MPS-I with a severe form (Hurler syndrome), and (2) the presence of R89Q may lead to a milder phenotype. We studied mutations in the IDUA gene from 19 MPS-I patients, including two pairs of siblings, with various clinical phenotypes (Hurler, 6 cases; Hurler/Scheie, 7 cases; Scheie, 6 cases). We report the presence of two common mutations that account for 42% of the 38 alleles in these patients. One is a novel 5-bp insertion between the thymidine at nt 704 and a cytosine at nt 705 (704ins5), which is seen only in the Japanese population. The other is a missense mutation, R89Q, which is also seen in Caucasians, although uncommonly. In the 19 Japanese MPS-I patients, the 704ins5 mutation accounted for 7 of 38 alleles (18%), while the R89Q accounted for 9 of 38 (24%). No Japanese patient was found to carry the W402X or Q70X alleles, the two most common MPS-I mutations in Caucasians. Homozygosity for the 704ins5 mutation is associated with a severe phenotype, and for the R89Q mutation with a mild phenotype. Compound heterozygosity for these two mutations produced an intermediate phenotype. Haplotype analysis using polymorphisms linked to the IDUA locus demonstrated that each mutation occurs on a different specific haplotype, suggesting that individuals with each of these common mutations derive from common founders. These data continue to document the molecular heterogeneity and racial differences in mutations in MPS-I.

Adult↗
PubMed1996

Mucopolysaccharidosis IVA: submicroscopic deletion of 16q24.3 and a novel R386C mutation of N-acetylgalactosamine-6-sulfate sulfatase gene in a classical Morquio disease.

The N-acetylgalactosamine-6-sulfate sulfatase (GALNS) gene, which is responsible for autosomal recessive mucopolysaccharidosis IVA (MPSIVA), has been assigned to the long arm of chromosome 16, subregion 24.3, an area where the adenine phosophoribosyltransferase (APRT) gene and renal dipeptidase (DPEP I) gene are also localized. Molecular genetic studies on a severely affected patient with MPSIVA (Morquio disease), without karyotypic abnormality, revealed a partial submicroscopic deletion of 16q24.3 and a single point mutation on the other allele, with no functional GALNS activity. The patient, his mother, and siblings were hemizygous for GALNS and APRT loci, evidenced by informative RFLP and gene dosage analyses combined with a fluorescence in situ hybridization, utilizing a partial genomic clone of GALNS, but heterozygosity was retained at the DPEP I locus and proximal D16S7. Haplotyping of the family members revealed recombinational events between DPEP I locus and three other polymorphic loci on the paternal chromosome, localizing GALNS gene on the proximal side to DPEP I gene. As estimated from the genetic distance between two flanking markers of proximal D16S7 and distal DPEP I locus, size of the deletion was less than 3Mb. Mother of the boy and two older siblings were asymptomatic, despite this interstitial deletion of the Giemsa-light G band. The remaining paternal allele had no gene rearrangement but GALNS activity was not encoded as Arginine at 386 was replaced with Cysteine (R386C), suggesting this alteration accounts for the severe phenotype. Allelic loss of APRT is frequently observed in cancer tissues, thereby suggesting that the tumor suppressor gene locates near the APRT locus. No family member has evidence of any malignant disease. This study is apparently the first documentation of interstitial deletion of 16q24.3, involving GALNS and APRT genes.

Adenine Phosphoribosyltransferase↗
PubMed1996

Mucopolysaccharidosis IVA: a comparative study of polymorphic DNA haplotypes in the Caucasian and Japanese populations.

Mucopolysaccharidosis IVA (MPS IVA) is an autosomal recessive disorder caused by deficiency of the lysosomal enzyme N-acetylgalactosamine-6-sulphate sulphatase (GALNS). The genetic heterogeneity at the GALNS locus was studied in 62 mutant alleles and 376 normal alleles in the Caucasian population and also in 40 mutant and 100 normal alleles in the Japanese population. For this study, six different restriction fragment length polymorphisms (RFLPs) at the GALNS locus were analysed to search for the frequency of each RFLP produced by StyI, SphI, RsaI, HaeIII, StuI and HapII restriction endonucleases. We detected a total of 27 haplotypes in the Caucasian and Japanese population. Of these 27 haplotypes, 18 haplotypes were present in the Caucasian population and the most common of these was haplotype 1 (ABHcde) in both mutant and normal alleles. In contrast, in the Japanese population we found 20 of the 27 haplotypes and the most common in mutant and normal alleles was haplotype 2 (abhcDE). Within these two populations a parent in the MPS IVA family has an average probability of greater than 77% (in the Caucasian population 77.27% and in the Japanese population 78.26%) of being heterozygous, and hence informative for linkage, at one or more GALNS RFLP sites. Our results delineate the molecular heterogeneity of MPS IVA haplotypes, as well as their significant interpopulation variation, and make prenatal diagnosis and carrier detection possible in the majority of families with one affected child.

Acetylgalactosamine↗
PubMed1996

Carrier identification of X-linked adrenoleukodystrophy by measurement of very long chain fatty acids and lignoceric acid oxidation.

Because identification of carriers of X-linked adrenoleukodystrophy (ALD) results in 5-15% false negatives with very long chain fatty acids (VLCFA) assay in plasma, and mutation analysis of plasma VLCFA combined with of the ALD gene is not always practical, we studied whether the analysis of plasma VLCFA combined with lignoceric acid oxidation study in fibroblasts could improve the rate of carrier detection. Lignoceric acid oxidation was abnormal in 19 out of 19 patients (ALD or adrenomyeloneuropathy) and in three out of three obligate heterozygous women. Among ten women at risk of being a carrier, three women who had normal plasma VLCFA had abnormal lignoceric acid oxidation in fibroblasts. These data suggest that this combined biochemical procedure may help to improve carrier detection in families when the ALD gene mutation has not been identified.

Adrenoleukodystrophy↗
PubMed1996

Correction by gene expression of biochemical abnormalities in fibroblasts from Zellweger patients.

Zellweger syndrome is a prototype of peroxisomal biogenesis disorders and a fatal autosomal recessive disease with no effective therapy. We identified nine genetic complementation groups of these disorders, and mutations in peroxisome assembly factor-1 (PAF-1) and the 70-kD peroxisomal membrane protein (PMP70) genes have been detected by our group F and Roscher's group 1, respectively. We now describe permanent recovery from generalized peroxisomal abnormalities in fibroblasts of a Zellweger patient from group F, such as biochemical defects of peroxisomal beta-oxidation, plasmalogen biosynthesis, and morphologic absence of peroxisomes, by stable transfection of human cDNA encoding PAF-1. In the light of these observations, we designed a gene expression system using fibroblasts from patients with peroxisomal biogenesis disorders. In Zellweger fibroblasts obtained from Roscher's group 1 and transfected with human cDNA encoding PMP70, peroxisomes were not morphologically identifiable, and peroxisomal function did not normalize.

ATP-Binding Cassette Transporters↗
PubMed1996

Mucopolysaccharidosis IVA: four new exonic mutations in patients with N-acetylgalactosamine-6-sulfate sulfatase deficiency.

We report four new mutations in Japanese patients with mucopolysaccharidosis IVA (MPSIVA) who were heterozygous for a common double gene deletion. A nonsense mutation of CAG to TAG at codon 148 in exon 4 was identified, resulting in a change of Q to a stop codon and three missense mutations. V (GTC) to A (GCC) at codon 138 in exon 4, P (CCC) to S (TCC) at codon 151 in exon 5, and P (CCC) to L (CTC) at codon 151 in exon 5. Introduction of these mutations into the normal GALNS cDNA and transient expression in cultured fibroblasts resulted in a significant decrease in the enzyme activity. V138A and Q148X mutations result in changes of restriction site, which were analyzed by restriction-enzyme assay. P151S and P151L mutations that did not alter the restriction site were detected by direct sequencing or allele specific oligohybridization. Detection of the double gene deletion was initially done using Southern blots and was confirmed by PCR. Haplotypes were determined using seven polymorphisms to the GALNS locus in families with the double gene deletion. Haplotype analysis showed that the common double gene deletion occurred on a single haplotype, except for some variation in a VNTR-like polymorphism. This finding is consistent with a common founder for all individuals with this mutation.

Adolescent↗
PubMed1996

Human peroxisome assembly factor-2 (PAF-2): a gene responsible for group C peroxisome biogenesis disorder in humans.

Peroxisome-biogenesis disorders (PBD) are genetically heterogeneous and can be classified into at least ten complementation groups. We recently isolated the cDNA for rat peroxisome assembly factor-2 (PAF-2) by functional complementation using the peroxisome-deficient Chinese-hamster-ovary cell mutant, ZP92. To clarify the novel pathogenic gene of PBD, we cloned the full-length human PAF-2 cDNA that morphologically and biochemically restores peroxisomes of group C Zellweger fibroblasts (the same as group 4 in the Kennedy-Krieger Institute) and identified two pathogenic mutations in the PAF-2 gene in two patients with group C Zellweger syndrome. The 2,940-bp open reading frame of the human PAF-2 cDNA encodes a 980-amino-acid protein that shows 87.1% identity with rat PAF-2 and also restored the peroxisome assembly after gene transfer to fibroblasts of group C patients. Direct sequencing of the PAF-2 gene revealed a homozygous 1-bp insertion at nucleotide 511 (511 insT) in one patient with group C Zellweger syndrome (ZS), which introduces a premature termination codon in the PAF-2 gene, and, in the second patient, revealed a splice-site mutation in intron 3 (IVS3+1G-->A), which skipped exon 3, an event that leads to peroxisome deficiency. Chromosome mapping utilizing FISH indicates that PAF-2 is located on chromosome 6p21.1. These results confirm that human PAF-2 cDNA restores peroxisome of group C cells and that defects in the PAF-2 produce peroxisome deficiency of group C PBD.

ATPases Associated with Diverse Cellular Activitie↗
PubMed1995

Mucopolysaccharidosis type IVA: common double deletion in the N-acetylgalactosamine-6-sulfatase gene (GALNS).

Mucopolysaccharidosis IVA (MPS IVA) is an autosomal recessive disorder caused by a deficiency in N-acetylgalactosamine-6-sulfatase (GALNS). We found two separate deletions of nearly 8.0 and 6.0 kb in the GALNS gene, including some exons. There are Alu repetitive elements near the breakpoints of the 8.0-kb deletion, and this deletion resulted from an Alu-Alu recombination. The other 6.0-kb deletion involved illegitimate recombinational events between incomplete short direct repeats of 8 bp at deletion breakpoints. The same rearrangement has been observed in a heteroallelic state in four unrelated patients. This is the first documentation of a common double deletion a gene that is not a member of a gene cluster.

Base Sequence↗
PubMed1995

Mucopolysaccharidosis type II (Hunter disease): identification and characterization of eight point mutations in the iduronate-2-sulfatase gene in Japanese patients.

Mucopolysaccharidosis type II (Hunter disease) is a lysosomal storage disorder caused by a deficiency of the enzyme iduronate-2-sulfatase. Varied clinical phenotypes of this disease have been described. To identify mutations in individual patients and to examine possible correlations between mutations and clinical phenotypes, we analyzed the iduronate-2-sulfatase gene in Japanese patients with different clinical phenotypes. Five missense mutations, S333L (severe), R468Q (severe), R468L (severe), W337R (intermediate), R48P (mild), and three nonsense mutations, W345X (severe), R443X (intermediate), Q531X (mild), were identified by the RT-PCR method. Transient expression in the enzyme-deficient fibroblasts revealed that all five missense mutant enzymes were synthesized as the normal-size precursor (73 kD), and the nonsense mutant enzymes were synthesized as truncated ones (W345X:54 kD, R443X:59 kD, and Q531X:69 kD), although stable mature enzymes (45-56 kD) were not detected by Western blot analysis. Furthermore, expression of the eight mutant cDNAs resulted in severe reductions of iduronate-2-sulfatase enzyme activity in comparison with a normal cDNA.

Adolescent↗
PubMed1995

Mucopolysaccharidosis IVA: structural gene alterations identified by Southern blot analysis and identification of racial differences.

Ninety-six alleles (36 alleles of Japanese and 60 of Caucasian origin) from forty-eight patients with mucopolysaccharidosis IVA were investigated for structural gene alterations using Southern blot analysis. All patients had a previously demonstrated deficiency of N-acetyl-galactosamine-6-sulfate-sulfatase and exhibited a wide spectrum of clinical severity. Initially, using the full-length cDNA as a probe, five of 36 chromosomes from the Japanese patients revealed similar rearrangements with respect to DNA digested with BamHI, SacI, and XhoI. Subsequent analysis using seven genomic fragments, covering the entire gene, enhanced the detection of aberrant fragments produced by the above restriction enzymes. Conversely, the 60 chromosomes of Caucasian origin revealed no evidence of large structural rearrangements when analyzed by these methods. There was a statistically significant difference between the two populations (P < 0.01). A severely affected Japanese patient showed structural rearrangements on both chromosomes by means of BamHI blots. An 8.0-kb fragment and a highly polymorphic 7.0-kb to 11.0-kb fragment present in normal individuals disappeared and two aberrant fragments of 11.5 kb and 12.0 kb were observed. Three other Japanese patients also showed these two aberrant fragments, in addition to the normal fragment pattern, and were thus heterozygous for this rearrangement. Interpretation of Southern blots was difficult because of the complexity of polymorphic bands resulting from variable number of tandem repeat elements. However, by utilizing these aberrant fragments or polymorphic bands, carrier detection was effective, even in families with poorly characterized mutations. Hybridization with probe MG-A (5'-end genomic probe in intron 1) showed a 8.4-kb fragment in BamHI blots of one Japanese and one Caucasian patient; XhoI, SacI, and EcoRI blots were normal. Since this BamHI alteration was also observed in one normal control, it appears to be a rare nonpathological polymorphism.

Alleles↗
PubMed1995

Mucopolysaccharidosis IVA: polymorphic haplotypes and informative RFLPs in the Japanese population.

Seven different restriction fragment length polymorphisms (RFLPs) at the N-acetylgalactosamine-6-sulfate sulfatase (GALNS) locus were analyzed using Southern blotting and polymerase chain reaction based techniques to search for the frequency of each RFLP produced by StyI, SphI, HaeIII, StuI, HapII, XhoI, and BamHI restriction endonucleases, respectively, in 36 mutant alleles, including two sibling cases and 100 normal alleles. Calculation of heterozygosity indexes showed that these RFLPs were polymorphic, ranging from 0.31 to 0.69 in mucopolysaccharidosis IVA (MPS IVA) patients compared with 0.21 to 0.65 in normal individuals. There was some significant difference in several RFLPs and in the combination with four kinds of RFLPs (SphI, StuI, HapII, XhoI polymorphisms). The normal alleles were composed of 13 different RFLPs haplotypes; the most common among the Japanese population carrying normal alleles was haplotype 8 (bDEF1) (31.3%), the others being dispersed. The same haplotype 8 was the most frequent in the mutant alleles (44.4%), with seven further haplotypes. These findings revealed the striking variety of polymorphic haplotypes in the MPS IVA gene. By using these five kinds of RFLPs, we examined the theoretical informativity of haplotype analysis in heterozygote detection in nine unrelated MPS IVA families and ten unrelated normal families. All the members of the MPS IVA families studied were diagnosed as a patient, carrier, or non-carrier. We propose that prenatal diagnosis or family analysis in cases in which mutations have not been characterized is now feasible.

Base Sequence↗
PubMed1995

Mucopolysaccharidosis IVA: screening and identification of mutations of the N-acetylgalactosamine-6-sulfate sulfatase gene.

Mutations causing mucopolysaccharidosis IVA in 15 Japanese and one Caucasian patient were characterized. To screen these mutations, we used a combination of single strand conformation polymorphism analysis and heteroduplex analysis for PCR products of targeted cDNA or genomic DNA. Various small mutations were identified in 23 of 26 alleles, while the other six alleles had large rearrangements. Cycle sequencing of PCR products revealed 15 different mutations, including 12 missense, one nonsense, one frame shift (2 bp deletion) and one splice site mutation, in accord with the broad range of clinical phenotypes. Two alleles have different mutations in the same nucleotide position of exon 3 (R94C, CGC-->TGC; R94G, CGC-->GGC), diagnosed by sequencing and by allelic-specific oligohybridization (ASO). One allele had two amino acid changes, E450V in exon 12 and V488M in exon 13, thereby indicating a double point mutation. All 16 mutations reported were confirmed by restriction enzyme assay or by allelic-specific oligohybridization. Transfection of mutagenized cDNAs into patients' fibroblasts showed that all mutations caused completely deficient or markedly decreased N-acetylgalactosamine-6-sulfate sulfatase (GALNS) activity, thereby indicating that these mutations were responsible for the enzyme deficiency.

Alleles↗