FOXC2 Mutations in Familial and Sporadic Spinal Extradural Arachnoid Cyst

Spinal extradural arachnoid cyst (SEDAC) is a cyst in the spinal canal that protrudes into the epidural space from a defect in the dura mater. Most cases are sporadic; however, three familial SEDAC cases have been reported, suggesting genetic etiological factors. All familial cases are associated with lymphedema-distichiasis syndrome (LDS), whose causal gene is FOXC2. However, FOXC2 mutation analysis has been performed in only 1 family, and no mutation analysis has been performed on sporadic (non-familial) SEDACs. We recruited 17 SEDAC subjects consisting of 2 familial and 7 sporadic cases and examined FOXC2 mutations by Sanger sequencing and structural abnormalities by TaqMan copy number assay. We identified 2 novel FOXC2 mutations in 2 familial cases. Incomplete LDS penetrance was noted in both families. Four subjects presented with SEDACs only. Thus, SEDAC caused by the heterozygous FOXC2 loss-of-function mutation should be considered a feature of LDS, although it often manifests as the sole symptom. Seven sporadic SEDAC subjects had no FOXC2 mutations, no symptoms of LDS, and showed differing clinical characteristics from those who had FOXC2 mutations, suggesting that other gene(s) besides FOXC2 are likely to be involved in SEDAC.     

A new sporadic case of early-onset Loeys-Dietz syndrome due to the recurrent mutation p.R528C in theTGFBR2 gene substantiates interindividual clinical variability

We report on a 2-year-old Polish girl with typical manifestations of Loeys-Dietz syndrome (LDS), a rare genetic condition belonging to the group of Marfan-related disorders. The characteristic LDS symptoms observed in the girl included craniofacial dysmorphism (craniosynostosis, cleft palate, hypertelorism), arachnodactyly, camptodactyly, scoliosis, joint laxity, talipes equinovarus, translucent and hyperelastic skin, and umbilical hernia. Mild dilatation of the ascending aorta and tortuous course of the left internal carotid artery were recognized during her second year of life. Molecular genetic testing revealed a heterozygous missense mutation (c.1582C>T, p.R528C) in the transforming growth factor beta receptor II gene (TGFBR2). This mutation has been previously associated with LDS in 5 unrelated cases, and was never reported in patients with other Marfan-related disorders. Comparison of the phenotypes of our patient and these 5 individuals with c.1582C>T showed that only the hallmark triad of the syndrome — consisting of hypertelorism, aortic root dilatation/aneurysm, and cleft palate or bifid uvula — was present in all 6 cases. Interestingly, none of the 5 individuals who underwent psychological evaluation showed developmental delay. The pattern of all other LDS features showed interindividual variability. Our data support the recently reported observation that symptoms of LDS can develop at a very young age, making early diagnosis and management essential for these patients. This is the first report on a Polish infant with typical LDS symptoms caused by aTGFBR2 mutation.     

Genetic Characterization of the SufJ Frameshift Suppressor in SALMONELLA TYPHIMURIUM

A new suppressor of +1 frameshift mutations has been isolated in Salmonella typhimurium. This suppressor, sufJ, maps at minute 89 on the Salmonella genetic map between the argH and rpo(rif) loci, closely linked to the gene for the ochre suppressor tyrU(supM). The suppressor mutation is dominant to its wild-type allele, consistent with the suppressor phenotype being caused by an altered tRNA species. The sufJ map position coincides with that of a threonine tRNA(ACC/U) gene; the suppressor has been shown to read the related fourbase codons ACCU, ACCC, ACCA.--The ability of sufJ to correct one particular mutation depends on the presence of a hisT mutation which causes a defect in tRNA modification. This requirement is allele specific, since other frameshift mutations can be corrected by sufJ regardless of the state of the hisT locus.--Strains carrying both a sufJ and a hisT mutation are acutely sensitive to growth inhibition by uracil; the inhibition is reversed by arginine. This behavior is characteristic of strains with mutations affecting the arginine-uracil biosynthetic enzyme carbamyl phosphate synthetase. The combination of two mutations affecting tRNA structure may reduce expression of the structural gene for this enzyme (pyrA).     

Multiple correcting COL17A1 mutations in patients with revertant mosaicism of epidermolysis bullosa

Revertant mosaicism by somatic reversion of inherited mutations has been described for a number of genetic diseases. Several mechanisms can underlie this reversion process, such as gene conversion, crossing-over, true back mutation, and second-site mutation. Here, we report the occurrence of multiple corrections in two unrelated probands with revertant mosaicism of non-Herlitz junctional epidermolysis bullosa, an autosomal recessive genodermatosis due to mutations in the COL17A1 gene. Immunofluorescence microscopy and laser dissection microscopy, followed by DNA and RNA analysis, were performed on skin biopsy specimens. In patient 1, a true back mutation, 3781T-->C, was identified in the specimen from the arm, and a second-site mutation, 4463-1G-->A, which compensated for the frameshift caused by the inherited 4424-5insC mutation, was identified in the 3' splice site of exon 55 in a specimen from the middle finger. Patient 2 showed--besides two distinct gene conversion events in specimens from the arm and hand sites, both of which corrected the 1706delA mutation--a second-site mutation (3782G-->C) in an ankle specimen, which prevented the premature ending of the protein by the 3781C-->T nonsense mutation (R1226X). Thus, both inherited mutations, paternal as well as maternal, reverted at least once by different reversion events in distinct cell clusters in the described patients. The occurrence of multiple correcting mutations within the same patient indicates that in vivo reversion is less unusual than was generally thought. Furthermore, in the male patient, mosaic patterns of type XVII collagen-positive keratinocytes were present in clinically unaffected and affected skin. This latter observation makes it likely that reversion may be overlooked and may happen more often than expected.     

Linkage to D3S47 (C17) in one large autosomal dominant retinitis pigmentosa family and exclusion in another: confirmation of genetic heterogeneity.

Recently Dryja and his co-workers observed a mutation in the 23d codon of the rhodopsin gene in a proportion of autosomal dominant retinitis pigmentosa (ADRP) patients. Linkage analysis with a rhodopsin-linked probe C17 (D3S47) was carried out in two large British ADRP families, one with diffuse-type (D-type) RP and the other with regional-type (R-type) RP. Significantly positive lod scores (lod score maximum [Zmax] = +5.58 at recombination fraction [theta] = .0) were obtained between C17 and our D-type ADRP family showing complete penetrance. Sequence and oligonucleotide analysis has, however, shown that no point mutation at the 23d codon exists in affected individuals in our complete-penetrance pedigree, indicating that another rhodopsin mutation is probably responsible for ADRP in this family. Significantly negative lod scores (Z less than -2 at theta = .045) were, however, obtained between C17 and our R-type family which showed incomplete penetrance. Previous results presented by this laboratory also showed no linkage between C17 and another large British R-type ADRP family with incomplete penetrance. This confirms genetic heterogeneity. Some types of ADRP are being caused by different mutations in the rhodopsin locus (3q21-24) or another tightly linked gene in this region, while other types of ADRP are the result of mutations elsewhere in the genome.     

Detection of a new mutation (T1140C) in a patient with Hunter syndrome from Guangdong, China

This study identified mutations of the idurnate-2-sulfatase (IDS) gene in a patient with Hunter syndrome, and established a basis for the diagnosis of the prenatal gene of Hunter syndrome. Urine glyeosaminoglycan (GAG) assay was used to make the preliminary diagnosis of mucopolysaccharidosis type II. Polymerase chain reaction (PCR) from dried blood spots and DNA sequencing were applied to analyze hotspot mutations in exons 9,3 and 8 of the IDS gene in the proband and his parents. A new missense mutation (T1140C) in exon 8 of the IDS gene was found by using DNA sequencing. This mutation caused a substitution of codon 339 from CTA (leucine) to CCA (praline). The patient is a hemizygote, and his mother is a heterozygote. The new missense mutation results in a change in the primary and tertiary structure of the IDS protein. It is possible that this mutation severely impairs enzymatic activity and is the underlying basis for the pathology seen in this patient with Hunter syndrome. __________ Translated from Hereditas, 2006, 28(5): 521–524 [译自: 遗传]     

Involvement of the Escherichia coli folate-binding protein YgfZ in RNA modification and regulation of chromosomal replication initiation

The Escherichia coli hda gene codes for a DnaA-related protein that is essential for the regulatory inactivation of DnaA (RIDA), a system that controls the initiation of chromosomal replication. We have identified the ygfZ gene, which encodes a folate-binding protein, as a suppressor of hda mutations. The ygfZ null mutation suppresses an hda null mutation. The over-initiation and abortive elongation phenotypes conferred by the hda mutations are partially suppressed in an hda ygfZ background. The accumulation of the active form of DnaA, ATP-DnaA, in the hda mutant is suppressed by the disruption of the ygfZ gene, indicating that YgfZ is involved in regulating the level of ATP-DnaA. Although ygfZ is not an essential gene, the ygfZ disruptant grows slowly, especially at low temperature, demonstrating that this gene is important for cellular proliferation. We have identified mnmE (trmE) as a suppressor of the ygfZ disruption. This gene encodes a GTPase involved in tRNA modification. Examination of RNA modification in the ygfZ mutant reveals reduced levels of 2-methylthio N(6)-isopentenyladenosine [corrected] indicating that YgfZ participates in the methylthio-group formation of this modified nucleoside in some tRNAs. These results suggest that YgfZ is a key factor in regulatory networks that act via tRNA modification.     

Efficient gene correction of an aberrant splice site in β‐thalassaemia iPSCs by CRISPR/Cas9 and single‐strand oligodeoxynucleotides

β‐thalassaemia is a prevalent hereditary haematological disease caused by mutations in the human haemoglobin β (HBB) gene. Among them, the HBB IVS2‐654 (C > T) mutation, which is in the intron, creates an aberrant splicing site. Bone marrow transplantation for curing β‐thalassaemia is limited due to the lack of matched donors. The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9), as a widely used tool for gene editing, is able to target specific sequence and create double‐strand break (DSB), which can be combined with the single‐stranded oligodeoxynucleotide (ssODN) to correct mutations. In this study, according to two different strategies, the HBB IVS2‐654 mutation was seamlessly corrected in iPSCs by CRISPR/Cas9 system and ssODN. To reduce the occurrence of secondary cleavage, a more efficient strategy was adopted. The corrected iPSCs kept pluripotency and genome stability. Moreover, they could differentiate normally. Through CRISPR/Cas9 system and ssODN, our study provides improved strategies for gene correction of β‐Thalassaemia, and the expression of the HBB gene can be restored, which can be used for gene therapy in the future.     

中国广东Hunter综合征患者的T1140C新突变

应用尿黏多糖含量检测、干血滤纸片直接扩增、PCR产物直接测序法对患者及其父母等的IDS基因的突变热点exons9,3,8进行突变检测。发现患儿IDS基因的exon8发生一新的错义突变,突变部位在第339位密码子（CTA）内,即cDNA第1140bp的T突变为C,导致原339位的“亮氨酸CTA”突变为“脯氨酸CCA”。该患儿为这一突变的半合子,而其母为这一突变的杂合子。该错义突变改变了IDS酶的一级结构和三级空间结构,从而可能引起IDS酶活性大大降低,这可能是该Hunter综合征患者的真正致病原因。