Tohru Sonoda

Loss of Dermatan-4-Sulfotransferase 1 Function Results in Adducted Thumb-Clubfoot Syndrome

Adducted thumb-clubfoot syndrome is an autosomal-recessive disorder characterized by typical facial appearance, wasted build, thin and translucent skin, congenital contractures of thumbs and feet, joint instability, facial clefting, and coagulopathy, as well as heart, kidney, or intestinal defects. We elucidated the molecular basis of the disease by using a SNP array-based genome-wide linkage approach that identified distinct homozygous nonsense and missense mutations in CHST14 in each of four consanguineous families with this disease. The CHST14 gene encodes N-acetylgalactosamine 4-O-sulfotransferase 1 (D4ST1), which catalyzes 4-O sulfation of N-acetylgalactosamine in the repeating iduronic acid-alpha1,3-N-acetylgalactosamine disaccharide sequence to form dermatan sulfate. Mass spectrometry of glycosaminoglycans from a patients fibroblasts revealed absence of dermatan sulfate and excess of chondroitin sulfate, showing that 4-O sulfation by CHST14 is essential for dermatan sulfate formation in vivo. Our results indicate that adducted thumb-clubfoot syndrome is a disorder resulting from a defect specific to dermatan sulfate biosynthesis and emphasize roles for dermatan sulfate in human development and extracellular-matrix maintenance.

Sotos syndrome and haploinsufficiency of NSD1: clinical features of intragenic mutations and submicroscopic deletions

Sotos syndrome (MIM 117550) is a congenital developmental disorder characterised by overgrowth and advanced bone age in infancy to early childhood, mental retardation, and various minor anomalies such as macrocephaly, prominent forehead, hypertelorism, downward slanting palpebral fissures, large ears, high and narrow palate, and large hands and feet.1,2 It is also frequently associated with brain, cardiovascular, and urinary anomalies3–6 and is occasionally accompanied by malignant lesions such as Wilms tumour and hepatocarcinoma.7,8 This condition has been classified as an autosomal dominant disorder, because several familial cases consistent with dominant inheritance have been described previously.9 Thus, sporadic cases accounting for most of the Sotos syndrome patients are assumed to be the result of de novo dominant mutations.nnWe have recently shown that Sotos syndrome is caused by haploinsufficiency of the gene for NSD1 (nuclear receptor binding Su-var, enhancer of zeste, and trithorax domain protein 1).10 NSD1 consists of 23 exons and encodes at least six functional domains possibly related to chromatin regulations (SET, PWWP-I, PWWP-II, PHD-I, PHD-II, and PHD-III), in addition to 10 putative nuclear localisation signals.11 It is expressed in several tissues including fetal/adult brain, kidney, skeletal muscle, spleen, and thymus11 and is likely to interact with nuclear receptors as a bifunctional transcriptional cofactor.12 In this paper, we report on clinical findings in Japanese patients with proven point mutations in NSD1 and those with submicroscopic deletions involving the entire NSD1 gene and discuss genotype-phenotype correlation.nnThis study consisted of five patients with heterozygous NSD1 point mutations and 21 patients with heterozygous submicroscopic deletions involving the entire NSD1 gene. The mutations were identified by direct sequencing of exons 2–23 and their flanking introns covering the whole coding region of NSD1 ,11 using genomic DNA extracted from peripheral leucocytes or …

Preferential Paternal Origin of Microdeletions Caused by Prezygotic Chromosome or Chromatid Rearrangements in Sotos Syndrome

Sotos syndrome (SoS) is characterized by pre- and postnatal overgrowth with advanced bone age; a dysmorphic face with macrocephaly and pointed chin; large hands and feet; mental retardation; and possible susceptibility to tumors. It has been shown that the major cause of SoS is haploinsufficiency of the NSD1 gene at 5q35, because the majority of patients had either a common microdeletion including NSD1 or a truncated type of point mutation in NSD1. In the present study, we traced the parental origin of the microdeletions in 26 patients with SoS by the use of 16 microsatellite markers at or flanking the commonly deleted region. Deletions in 18 of the 20 informative cases occurred in the paternally derived chromosome 5, whereas those in the maternally derived chromosome were found in only two cases. Haplotyping analysis of the marker loci revealed that the paternal deletion in five of seven informative cases and the maternal deletion in one case arose through an intrachromosomal rearrangement, and two other cases of the paternal deletion involved an interchromosomal event, suggesting that the common microdeletion observed in SoS did not occur through a uniform mechanism but preferentially arose prezygotically.

Four novel NIPBL mutations in Japanese patients with Cornelia de Lange syndrome

Cornelia de Lange syndrome (CdLS, OMIM #122470) is a multiple congenital anomaly syndrome characterized by dysmorphic facial features, hirsutism, severe growth and developmental delay, and malformed upper limbs [Ireland et al., 1993; Jackson et al., 1993]. The prevalence is estimated to be 1/10,000 [Opitz, 1985]. Recently, two independent groups proved that CdLS is caused by NIPBL mutations [Krantz et al., 2004; Tonkin et al., 2004]. NIPBL consists of 47 exons and encodes delangin, a 2,804 amino-acid protein, from exon 2 to 47. We analyzed 15 Japanese sporadic patients (CdL 1–15) with typical CdLS features (Table I) and their parents after obtaining written informed consent. All protocols in this study were approved by the Committee for the Ethical Issues on Human Genome and Gene analysis, Nagasaki University. Clinical geneticists diagnosed these patients based on mental and growth retardation, and characteristic facial features. Genomic DNA was extracted using a standard protocol. Fourty-six coding exons (from exon 2 to 47) of NIPBL were amplified by PCR as described previously [Krantz et al., 2004] except for exons 4, 33, 37, and 41, of which primers were originally designed (available on request). Sequence analysis was performed as described previously [Kurotaki et al., 2003]. We identified three novel nonsense mutations and one missense mutation in NIPBL among the 15 Japanese patients examined: 1885C>T (R629X) (CdL 4) and 1921G>T (E641X) (CdL 2) in exon 10, 3346G>T (E1116X) (CdL 15) in exon 12, and 5483G>A (R1828Q) (CdL 10) in exon 29. All the four mutations were not found in any of 97 normal Japanese controls or in the JSNP database (http://snp.ims.u-tokyo.ac.jp/). The altered amino acid (R1828Q) was de novo and located in the evolutionally conserved sequences at least in the human, rat, mouse, and fly homologs, thus the change is likely to be pathological. The C-terminal half 1500 amino acids of delangin is well conserved among homologs of flies, worms, plants, and fungi, and is expected to be biologically important [Tonkin et al., 2004], though it was not found to contain any obvious functional domains by analysis using PROSITE (http://kr. expasy.org/cgi-bin/prosite/PSScan.cgi). Three protein truncation mutations at amino acid positions 629, 641, and 1116 and a missense mutation at amino acid position 1828 could lose or impair the C-terminal half function. The Drosophila homolog of NIPBL, Nipped-B, is involved in activating the Ubx and Cut homeobox genes. Ubx suppresses the limb formation by repressing Dll that requires for the distal limb development, and Cut mutations cause leg and wing abnormalities [Tonkin et al., 2004]. Thus, it is plausible that reduced expression of human NIPBL may lead to limb anomalies in CdLS. Interestingly, limb abnormalities (oligodactyly and ulner deficiency) were observed in three of our four patients with a mutation, but only one of seven patients without any mutation whose clinical information was available did show some limb abnormality (oligodactyly), though Gillis et al. [2004] reported that severity of limb defects was not statistically different between mutation-positive and mutation-negative patients. Additionally, three single nucleotide polymorphisms (SNPs), 1151A>G (N384S) in exon 9, 2021A>G (N674S) in exon 10 and 5874T>C (S1958S) in exon 33, were identified, as they were found among normal controls and the second substitution (2021A>G) was previously reported as a SNP [Gillis et al., 2004]. Allele frequencies of the three SNPs in normal Japanese controls are 3.2% (6/186), 13.0% (25/192), and 64.5% (129/200), respectively. To exclude a submicroscopic deletion around NIPBL and its franking regions, fluorescence in situ hybridization (FISH) analysis was performed in 10 of 15 cases on their metaphase chromosomes using two BAC clones covering the NIPBL gene (Table I), RP11-14I21, and RP11-7M4, selected from the UCSC genome browser, 2003 July version (http://genome.ucsc.edu/ cgi-bin/hgGateway). FISH and subsequent photomicroscopy were performed as described previously [Miyake et al., 2004]. However, none of them showed any deletion. We also investigated core promoter regions in 11 affected individuals not having detectable point mutations in the coding regions. Two core promoter regions were identified, ranging 800 to 500 bp (CPR-A) and 400 to þ 200 bp (CPRB) from the beginning of NIPBL cDNA (NM_015384.3) using four different promoter prediction programs: neural network promoter prediction program (http://www.fruitfly.org/seq_ tools/promoter.html), human core-promoter finder (http:// rulai.cshl.org/tools/genefinder/CPROMOTER/human.htm), promoter 2.0 prediction server (http://www.cbs.dtu.dk/services/ promoter/), bioinformatics & molecular analysis section (http:// bimas.dcrt.nih.gov/molbio/proscan/). No nucleotide changes were detected among the 11 patients in the two core promoter regions except for a part of CPR-B sequence ( 60 þ 60), which was hardly determined due to high GC ratio (75.83%), suggesting that promoter mutations in NIBPL is less likely. In conclusion, we identified four novelNIPBLmutations and three SNPs. It is important to describe a full spectrum of phenotype in more patients with positive mutations and establish comprehensive diagnostic criteria. *Correspondence to: Dr. Naomichi Matsumoto, Department of Human Genetics, Yokohama City University Graduate School of Medicine, Fukuura 3-9, Kanazawa-ku, Yokohama 236-0004, Japan. E-mail: naomat@yokohama-cu.ac.jp

Unique properties of cluster of differentiation 93 in the umbilical cord blood of neonates.

It has previously been reported by these authors that cluster of differentiation (CD) 93 is co‐expressed on naive T‐lymphocytes (CD4+CD45RA+ cells) in neonatal umbilical cord blood cells (UCBCs) but not on normal adult peripheral blood cells (PBCs). In this study, expression of CD93 on other lymphocyte subsets and the concentration of soluble formed CD93 (sCD93) in serum or culture supernatants from neonatal umbilical cord blood (UCB) was examined. It was found that CD93 is also co‐expressed on CD2+, CD16+, CD56+ or CD25+ cells in the lymphocyte population of neonatal UCBCs, but not on normal adult PBCs. The concentrations of sCD93 in serum and culture supernatants from neonatal UCB were significantly greater than those from normal adult peripheral blood. The concentrations of sCD93 in culture supernatants from neonatal UCBCs and normal adult PBCs treated with phorbol 12‐myristate 13‐acetate (PMA) were significantly enhanced compared with those without PMA treatment. The degree of enhancement of sCD93 by PMA in culture supernatants from neonatal UCBCs was significantly greater than that of normal adult PBCs and enhancement of sCD93 by PMA in the culture supernatants from neonatal UCBCs and normal adult PBCs was significantly suppressed by PKC inhibitor. Interestingly, the high concentration of serum sCD93 in neonates was significantly decreased in sera from infants at 1 month after birth. Expression of CD93 on the lymphocyte population of PBCs from infants at 1 month after birth was also significantly decreased, compared with that for neonatal UCBCs. These findings indicate that CD93 in neonatal UCB has unique properties as an immunological biomarker.

Duplication (22)(q11.22-q11.23) without coloboma and cleft lip or palate

reported.1 The manifestations of these trisomies partially overlap. In general the severity of the phenotype correlates with the size of the duplication.1 There are two well characterized syndromes associated with partial trisomy of chromosome 22q: (i) proximal 22q trisomy syndrome (the cat-eye syndrome (CES)); and (ii) distal 22q trisomy syndrome.2 Partial trisomies that are derived from a parental translocation involving chromosome 22q exhibit variable phenotypes because such trisomies are derivatives involving other chromosomes. Alternatively, the phenotypic difference may be mainly due to the extent of the segment involved or to different breakpoints. In the present paper we describe a patient with a ‘pure’ duplication of 22q, from 22q11.22 to q11.23, without involvement of any other chromosome segment. The patient does not have coloboma, which commonly occurs in CES, or cleft lip and palate, which commonly occurs in distal trisomy 22q. This case appears to support previous suggestions that coloboma is associated with the duplication of the 22q pericentromere,3 and that the critical region of the 22q distal trisomy is associated with 22q13.1-qter.4