A Novel AMELX Mutation, Its Phenotypic Features, and Skewed X Inactivation.

Amelogenesis imperfecta (AI) is a group of genetic disorders of defective dental enamel. Mutation of AMELX encoding amelogenin on the X chromosome is a major cause of AI. Here we report a Chinese family with hypoplastic and hypomineralized AI. Whole exome analysis revealed a novel mutation c.185delC in exon 5 of AMELX causing the frame shift p.Pro62ArgfsTer47 (or p.Pro62Argfs*47). By sequencing of polymerase chain reaction products and T-vector clones, the mutation was confirmed as homozygous in the proband, hemizygous in her father, and heterozygous in her mother. The proband and her father had small and yellowish teeth with thin and rough enamel that was radiographically indistinguishable from the underlying dentin. Scanning electronic microscopy of 1 maternal tooth showed cracks and exposed loosely packed enamel prisms in affected areas. Consistent with a 25:75 skewing of X inactivation in the peripheral blood DNA as measured by androgen receptor allele methylation, the surface of the mother's tooth had alternating vertical ridges of transparent normal and white chalky enamel in a 34:66 ratio. In summary, this study provides one of the few phenotypic comparisons of hemizygous and homozygous AMELX mutations and suggests that the skewing of X inactivation in AI contributes to the phenotypic variations in heterozygous carriers of X-linked AI.

Dental abnormalities in Schimke immuno-osseous dysplasia.

Described for the first time in 1971, Schimke immuno-osseous dysplasia (SIOD) is an autosomal-recessive multisystem disorder that is caused by bi-allelic mutations of SMARCAL1, which encodes a DNA annealing helicase. To define better the dental anomalies of SIOD, we reviewed the records from SIOD patients with identified bi-allelic SMARCAL1 mutations, and we found that 66.0% had microdontia, hypodontia, or malformed deciduous and permanent molars. Immunohistochemical analyses showed expression of SMARCAL1 in all developing teeth, raising the possibility that the malformations are cell-autonomous consequences of SMARCAL1 deficiency. We also found that stimulation of cultured skin fibroblasts from SIOD patients with the tooth morphogens WNT3A, BMP4, and TGFß1 identified altered transcriptional responses, raising the hypothesis that the dental malformations arise in part from altered responses to developmental morphogens. To the best of our knowledge, this is the first systematic study of the dental anomalies associated with SIOD.

19p13.2 microduplication causes a Sotos syndrome-like phenotype and alters gene expression.

Up to 90% of individuals affected by Sotos syndrome have a pathogenic alteration of NSD1 (encodes nuclear receptor-binding Su-var, enhancer of zeste, and trithorax domain protein 1), a histone methyltransferase that functions as both a transcriptional activator and a repressor. Genomic copy number variations may also cause a Sotos-like phenotype. We evaluated a three-generation family segregating a Sotos-like disorder characterized by typical facial features, overgrowth, learning disabilities, and advanced bone age. Affected individuals did not have a detectable NSD1 mutation, but rather were found to have a 1.9 Mb microduplication of 19p13.2 with breakpoints in two highly homologous Alu elements. Because the duplication included the DNA methyltransferase gene (DNMT1), we assessed DNA methylation of peripheral blood and buccal cell DNA and detected no alterations. We also examined peripheral blood gene expression and found evidence for increased expression of genes within the duplicated region. We conclude that microduplication of 19p13.2 is a novel genomic disorder characterized by variable neurocognitive disability, overgrowth, and facial dysmorphism similar to Sotos syndrome. Failed compensation of gene duplication at the transcriptional level, as seen in peripheral blood, supports gene dosage as the cause of this disorder.

Schimke immuno-osseous dysplasia: SMARCAL1 loss-of-function and phenotypic correlation.

BACKGROUND: Schimke immuno-osseous dysplasia (SIOD) is an autosomal recessive pleiotropic disorder caused by mutations in SMARCAL1. SMARCAL1 encodes an enzyme with homology to the SNF2 chromatin remodelling proteins. METHODS: To assess the affect of SMARCAL1 mutations associated with SIOD on SMARCAL1 expression and function, we characterised the effects of various mutations on mRNA and protein expression in patient tissues and cell lines, and the ATPase activity, subcellular localisation, and chromatin binding of SMARCAL1 missense mutants. RESULTS: The SIOD associated SMARCAL1 mutations affected SMARCAL1 protein expression, stability, subcellular localisation, chromatin binding, and enzymatic activity. Further, expressing SMARCAL1 missense mutants in Drosophila melanogaster showed that disease severity was inversely proportionate to overall SMARCAL1 activity. CONCLUSION: Our results show for the first time that SMARCAL1 binds chromatin in vivo and that SIOD arises from impairment of diverse SMARCAL1 functions.

Vaso-occlusion in Schimke-immuno-osseous dysplasia: is the NO pathway involved?

Schimke-immuno-osseous dysplasia (SIOD) is an autosomal recessive disorder with the main clinical findings of spondyloepiphyseal dysplasia, nephrotic syndrome, and defective cellular immunity. Vaso-occlusive processes, especially generalized atherosclerosis, are a life-limiting complication in patients with severe SIOD. The nitric oxide synthase (NOS) oxidizes L-arginine to nitric oxide (NO). NO is a potent vasodilator with inhibitory effects on platelet aggregation and the development of atherosclerosis. We hypothesized that reduced NO production due to antagonism of NOS by asymmetric dimethylarginine (ADMA) would be a possible pathophysiological mechanism for vaso-occlusion in SIOD. We tested this hypothesis in 10 patients with SIOD and 10 age-matched healthy controls. Plasma and urine levels of nitrite and nitrate, the indicators of NO synthesis, and of ADMA, an endogenous NOS inhibitor, in children suffering from SIOD were not significantly different from those in the age-matched healthy controls. Our results suggest that the L-arginine/NO pathway is not altered in SIOD. Antagonism of NOS by ADMA does not seem to be the cause of premature general atherosclerosis in SIOD. The underlying pathology of vaso-occlusion in SIOD still remains unclear.

A novel 8.5 MB dup(1)(p34.1p34.3) characterized by FISH in a child presenting with congenital heart defect and dysmorphic features.

Chromosome duplications involving 1p are rarely reported but are apparently associated with short survival as well as congenital malformations and impaired development. Several of these have had congenital heart defects, although too few patients have been reported with similar breakpoints to characterize a syndrome. We present a girl with a novel interstitial duplication in the short arm of chromosome 1 [46,XX,dup(1)(p34.1p34.3)]. She presented with congenital heart defects at 1 month and by 1 year of age manifested delayed acquisition of motor milestones and subsequently of language milestones. By breakpoint-mapping using FISH analysis, we determined that her 1p duplication spans 8.5 megabases. Her 1p duplication is the smallest reported to date to contain 1p34 in patients with congenital heart defect due to abnormalities of heart looping during development. Thus, her 8.5 MB duplication provides a target region to search for a potentially dosage-sensitive gene(s) causing abnormal heart looping when duplicated. Two patients have been reported with duplication including 1p34 but without congenital heart defect, and their duplications span all but the distal approximately 2 MB segment duplicated in our patient. Thus, within our patient's 8.5 MB target region for a dosage sensitive gene leading to looping abnormalities (and thereby congenital heart defect), the distal 2 MB region might well be the region to begin the search.

SET binding factor 2 (SBF2) mutation causes CMT4B with juvenile onset glaucoma.

The authors report a Japanese family segregating autosomal recessive Charcot-Marie-Tooth disease (CMT) with focally folded myelin, juvenile-onset glaucoma, and a nonsense mutation of SET binding factor 2 (SBF2). The consistent phenotypic features associated with SBF2 mutations are early-onset demyelinating neuropathy, myelin folding, and markedly decreased motor nerve conduction velocities; glaucoma associates with SBF2 nonsense mutations.

Mutations in the neurofilament light chain gene (NEFL) cause early onset severe Charcot-Marie-Tooth disease.

Neurofilament light chain polypeptide (NEFL) is one of the most abundant cytoskeletal components of the neuron. Mutations in the NEFL gene were recently reported as a cause for autosomal dominant Charcot-Marie-Tooth type 2E (CMT2E) linked to chromosome 8p21. In order to investigate the frequency and phenotypic consequences of NEFL mutations, we screened 323 patients with CMT or related peripheral neuropathies. We detected six disease associated missense mutations and one 3-bp in-frame deletion clustered in functionally defined domains of the NEFL protein. Patients have an early onset and often a severe clinical phenotype. Electrophysiological examination shows moderately to severely slowed nerve conduction velocities. We report the first nerve biopsy of a CMT patient with a de novo missense mutation in NEFL, and found an axonal pathology with axonal regeneration clusters and onion bulb formations. Our findings provide further evidence that the clinical variation observed in CMT depends on the gene mutated and the specific type of mutation, and we also suggest that NEFL mutations need to be considered in the molecular evaluation of patients with sporadic or dominantly inherited CMT.

Screening for mutations in a genetically heterogeneous disorder: DHPLC versus DNA sequence for mutation detection in multiple genes causing Charcot-Marie-Tooth neuropathy.

PURPOSE: To determine the efficacy of denaturing high-performance liquid chromatography (DHPLC) for mutation detection in genetically heterogeneous diseases using Charcot-Marie-Tooth neuropathy as a model. METHODS: (1) Identification of the optimal conditions for mutation scanning by DHPLC using 50 known variants of PMP22, MPZ, GJB1 and EGR2. (2) Comparison of DHPLC with DNA sequencing for mutation detection in 168 patient DNA samples. RESULTS: We established the optimal conditions for screening PMP22, MPZ, GJB1, and EGR2 for mutations. Under optimized conditions, DHPLC was as sensitive as DNA sequencing and detected two mutations that were not identified by automated DNA sequence. CONCLUSIONS: DHPLC increases the efficiency and sensitivity of mutation screening in genetically heterogeneous diseases.

EGR2 mutation R359W causes a spectrum of Dejerine-Sottas neuropathy.

Heterozygous mutations in the early growth response gene 2 (EGR2), which encodes a zinc-finger transcription factor that regulates the late stages of myelination, cause myelinopathies including congenital hypomyelinating neuropathy, Dejerine-Sottas neuropathy (DSN), and Charcot-Marie-Tooth disease type 1. We screened 170 unrelated neuropathy patients without mutations involving the peripheral myelin protein 22 gene (PMP22), the myelin protein zero gene (MPZ), or the gap junction protein beta1 gene (GJB1) and identified two DSN patients with the heterozygous mutation R359W in the alpha-helix domain of the first zinc-finger of EGR2. We now report that this mutation is a recurrent cause of DSN, and that expressivity ranges from that typical for DSN to a more rapidly progressive neuropathy that can cause death by age 6 years. Furthermore, in contrast to patients with typical DSN, patients with the EGR2 R359W mutation have more respiratory compromise and cranial nerve involvement.

Denaturing high-performance liquid chromatography of the myotubularin-related 2 gene (MTMR2) in unrelated patients with Charcot-Marie-Tooth disease suggests a low frequency of mutation in inherited neuropathy.

Charcot-Marie-Tooth type 4B (CMT4B), an autosomal recessive demyelinating neuropathy characterized by focally folded myelin sheaths in the peripheral nerve, has been associated with mutations in the gene encoding myotubularin-related protein 2, MTMR2, on chromosome 11q22. To investigate whether mutations in MTMR2 may also cause different forms of CMT, we screened 183 unrelated patients with a broad spectrum of CMT and related neuropathies using denaturing high-performance liquid chromatography. We identified four frequent and three rare exonic variants; two of the rare variants were identified in two unrelated patients with congenital hypomyelinating neuropathy and not in the normal controls. Our results suggest that loss-of-function mutations in MTMR2 are preferentially associated with the CMT4B phenotype.

Periaxin mutations cause recessive Dejerine-Sottas neuropathy.

The periaxin gene (PRX) encodes two PDZ-domain proteins, L- and S-periaxin, that are required for maintenance of peripheral nerve myelin. Prx(-/-) mice develop a severe demyelinating peripheral neuropathy, despite apparently normal initial formation of myelin sheaths. We hypothesized that mutations in PRX could cause human peripheral myelinopathies. In accordance with this, we identified three unrelated Dejerine-Sottas neuropathy patients with recessive PRX mutations-two with compound heterozygous nonsense and frameshift mutations, and one with a homozygous frameshift mutation. We mapped PRX to 19q13.13-13.2, a region recently associated with a severe autosomal recessive demyelinating neuropathy in a Lebanese family (Delague et al. 2000) and syntenic to the location of Prx on murine chromosome 7 (Gillespie et al. 1997).

Manifestations and treatment of Schimke immuno-osseous dysplasia: 14 new cases and a review of the literature.

UNLABELLED: Schimke immuno-osseous dysplasia (SIOD) is a rare autosomal recessive spondylo-epiphyseal dysplasia. The characteristic features of SIOD include 1) short stature with hyperpigmented macules and an unusual facies, 2) proteinuria with progressive renal failure, 3) lymphopenia with recurrent infections, and 4) cerebral ischaemia. Although 25 patients have been reported with this disorder, the clinical course and phenotype of SIOD are not well characterized. This report summarizes the clinical findings, course and treatment of reported patients and includes 14 additional patients with SIOD. We emphasize the high incidence of cerebral ischaemia and ocular abnormalities, define the high incidence of thyroid dysfunction and blood cytopenia, and confirm the absence of effective and durable medical therapies. CONCLUSION: Schimke immuno-osseous dysplasia is a multi-system autosomal recessive disorder with variable expression that affects the skeletal, renal, immune, vascular, and haematopoietic systems. Medical therapy is limited especially for more severely affected individuals.

Alström syndrome: further evidence for linkage to human chromosome 2p13.

Alström syndrome is a rare autosomal recessive disorder characterized by retinal degeneration, sensorineural hearing loss, early-onset obesity, and non-insulin-dependent diabetes mellitus. The gene for Alström syndrome (ALMS1) has been previously localized to human chromosome 2p13 by homozygosity mapping in two distinct isolated populations - French Acadian and North African. Pair-wise analyses resulted in maximum lod (logarithm of the odds ratio) scores of 3.84 and 2.9, respectively. To confirm these findings, a large linkage study was performed in twelve additional families segregating for Alström syndrome. A maximum two-point lod score of 7.13 (theta = 0.00) for marker D2S2110 and a maximum cumulative multipoint lod score of 9.16 for marker D2S2110 were observed, further supporting linkage to chromosome 2p13. No evidence of genetic heterogeneity was observed in these families. Meiotic recombination events have localized the critical region containing ALMS1 to a 6.1-cM interval flanked by markers D2S327 and D2S286. A fine resolution radiation hybrid map of 31 genes and markers has been constructed.

Molecular mechanisms for CMT1A duplication and HNPP deletion.

As the best characterized human genomic disorders, CMT1A and HNPP illustrate several common mechanistic features of genomic rearrangements. These features include the following: (1) Recombination occurs between homologous sequences flanking the duplicated/deleted genomic segment. (2) The evolution of the mammalian genome may result in an architecture consisting of region-specific low-copy repeats that predispose to rearrangement secondary to providing homologous regions as substrate for recombination. (3) Strand exchange occurs preferentially in a region of perfect sequence identity within the flanking repeat sequences. (4) Double-strand breaks likely initiate recombination between the flanking repeats. (5) The mechanism and rate of homologous recombination resulting in DNA rearrangement may differ for male and female gametogenesis. (6) Homologous recombination resulting in DNA rearrangement occurs with high frequency in the human genome. (7) Genomic disorders result from structural features of the human genome and not population specific alleles or founder effects; therefore, genomic disorders appear to occur with equal frequencies in different world populations.

Frequent monoallelic loss of D13S319 in multiple myeloma patients shown by interphase fluorescence in situ hybridization.

Deletions or monosomy of chromosome 13 are frequent in multiple myeloma (MM). A candidate tumor suppressor gene might reside telomeric of the retinoblastoma gene (RBl) at band 13q14 and to play a role in B cell neoplasm. The D13S319 locus, between RB1 and D13S25 loci at 13q14 is the most commonly deleted marker in chronic lymphocytic leukemia (CLL) and non-Hodgkin's lymphoma (NHL). We evaluated the D13S319 locus in 24 MM cases by fluorescence in situ hybridization (FISH). We observed monosomy for D13S319 in 6/20 (30%) MM patients with an apparently normal karyotype. As expected, in four karyotypically abnormal MM cases with partial or complete monosomy for chromosome 13, all of them had monoallelic loss of D13S319. Our results indicated that the loss of D13S319 is commonly found in MM, even at diagnosis, and is more frequent than predicted based on conventional cytogenetic analysis of metaphase spreads. This finding implicates a candidate tumor suppressor gene at 13q14 in the pathogenesis of MM.

Molecular Mechanisms for CMT1A Duplication and HNPP Deletion.

As the best characterized human genomic disorders, 118 CMT1A and HNPP illustrate several common mechanistic features of genomic rearrangements. These features include the following: (1) Recombination occurs between homologous sequences flanking the duplicated/deleted genomic segment. (2) The evolution of the mammalian genome may result in an architecture consisting of region-specific low-copy repeats that predispose to rearrangement secondary to providing homologous regions as substrate for recombination. (3) Strand exchange occurs preferentially in a region of perfect sequence identity within the flanking repeat sequences. (4) Double-strand breaks likely initiate recombination between the flanking repeats. (5) The mechanism and rate of homologous recombination resulting in DNA rearrangement may differ for male and female gametogenesis. (6) Homologous recombination resulting in DNA rearrangement occurs with high frequency in the human genome. (7) Genomic disorders result from structural features of the human genome and not population specific alleles or founder effects; therefore, genomic disorders appear to occur with equal frequencies in different world populations.

Schimke immunoosseous dysplasia complicated by moyamoya phenomenon.

Schimke immunoosseous dysplasia (SID) is an autosomal recessive spondyloepiphyseal dysplasia that was first described by Schimke et al. [1971: Lancet 2:1088-1089]. It is associated with premature arteriosclerosis and cerebral ischemia; however, the cerebral vascular abnormalities causing ischemia have not been described [Spranger et al., 1991: J Pediatr 119:64-72; Ehrich et al., 1995: Clin Nephrol 43:89-95]. Based on magnetic resonance angiography (MRA) and magnetic resonance venography (MRV), we now report on 2 girls with SID who have cerebral ischemia associated with moyamoya phenomenon. In addition, one patient also has an absent or occluded left transverse sinus and diffuse aortic narrowing. This is the first characterization of the cerebral vascular abnormality found in SID and raises the possibility that cerebral moyamoya may represent another major manifestation of the underlying genetic defect in SID.

Lethal neonatal Menkes' disease with severe vasculopathy and fractures.

A male neonate presented with an acute onset of severe intra-abdominal bleeding, haemorrhagic shock and multiple fractures leading to death on d 27. Menkes' disease was diagnosed at autopsy and confirmed by copper accumulation studies on cultured fibroblasts. Such an early onset of fatal complications in this condition has not been previously reported. New insights into the pathogenesis of Menkes' disease provided by DNA mutation analysis and difficulties in neonatal diagnosis are discussed. Menkes' disease should be considered in male infants with pathological fractures and other signs of connective tissue disease, even in the neonatal period.