Molecular interaction between fukutin and POMGnT1 in the glycosylation pathway of alpha-dystroglycan

The recent identification of mutations in genes encoding demonstrated or putative glycosyltransferases has revealed a novel mechanism for congenital muscular dystrophy. Hypoglycosylated alpha-dystroglycan (alpha-DG) is commonly seen in Fukuyama-type congenital muscular dystrophy (FCMD), muscle-eye-brain disease (MEB), Walker-Warburg syndrome (WWS), and Large(myd) mice. POMGnT1 and POMTs, the gene products responsible for MEB and WWS, respectively, synthesize unique O-mannose sugar chains on alpha-DG. The function of fukutin, the gene product responsible for FCMD, remains undetermined. Here we show that fukutin co-localizes with POMGnT1 in the Golgi apparatus. Direct interaction between fukutin and POMGnT1 was confirmed by co-immunoprecipitation and two-hybrid analyses. The transmembrane region of fukutin mediates its localization to the Golgi and participates in the interaction with POMGnT1. Y371C, a missense mutation found in FCMD, retains fukutin in the ER and also redirects POMGnT1 to the ER. Finally, we demonstrate reduced POMGnT1 enzymatic activity in transgenic knock-in mice carrying the retrotransposal insertion in the fukutin gene, the prevalent mutation in FCMD. From these findings, we propose that fukutin forms a complex with POMGnT1 and may modulate its enzymatic activity.     

Membrane topology of the human seipin protein

The Berardinelli-Seip congenital lipodystrophy type 2 (BSCL2) gene encodes an integral membrane protein, called seipin, of unknown function localized to the endoplasmic reticulum of eukaryotic cells. Seipin is associated with the heterogeneous genetic disease BSCL2, and mutations in an N-glycosylation motif links the protein to two other disorders, autosomal-dominant distal hereditary motor neuropathy type V and Silver syndrome. Here, we report a topological study of seipin using an in vitro topology mapping assay. Our results suggest that the predominant form of seipin is 462 residues long and has an N(cyt)-C(cyt) orientation with a long luminal loop between the two transmembrane helices.     

MLPA: A prenatal diagnostic tool for the study of congenital heart defects?

Congenital heart defects (CHD) represent the most common birth defects, so they are not a rare finding when performing routine ultrasound examinations during pregnancy. Once chromosome abnormalities have been excluded in a fetus with a CHD, chromosome 22q11.2 deletion is usually investigated by FISH, as it is the most frequent microdeletion syndrome and is generally associated with cardiac malformations. If 22q11.2 microdeletion is ruled out, the etiology of the CHD remains generally unexplained, making familial genetic counseling difficult. To evaluate the usefulness of Multiplex Ligation-dependent Probe Amplification (MLPA) kits designed for the study of 22q11.2 and other genomic regions previously associated with syndromic CHD, we performed MLPA in 55 pregnancies with fetuses presenting CHD, normal karyotype and negative FISH results for 22q11.2 microdeletion, which constitutes the largest prenatal series reported. Definitive MLPA results were obtained in 50 pregnancies, and in this setting such MLPA kits did not detect any imbalance. On the other hand, to compare FISH and MLPA techniques for the study of 22q11.2 microdeletions, we performed MLPA in 4 pregnancies known to have 22q11.2 deletions (by FISH). All four 22q11.2 microdeletions were also detected by MLPA, which corroborates that it is a reliable technique for the diagnosis and characterization of 22q11.2 deletions. Finally, we evaluated the possibility of replacing conventional FISH by MLPA for the prenatal diagnosis of CHD, comparing the diagnostic potential, results delivery times, repetition and failure rates and cost of both techniques, and concluded that FISH should still be the technique of choice for the prenatal diagnosis of fetuses with CHD.     

Steroid 21-hydroxylase gene mutational spectrum in 50 Tunisian patients: Characterization of three novel polymorphisms

Congenital adrenal hyperplasia (CAH) is an autosomal recessive disease of steroid biosynthesis in humans. More than 90% of all CAH cases are caused by mutations of the 21-hydroxylase gene (CYP21A2), and approximately 75% of the defective CYP21A2 genes are generated through an intergenic recombination with the neighboring CYP21A1P pseudogene. In this study, the CYP21A2 gene was genotyped in 50 patients in Tunisia with the clinical diagnosis of 21-hydroxylase deficiency. CYP21A2 mutations were identified in 87% of the alleles. The most common point mutation in our population was the pseudogene specific variant p.Q318X (26%). Three novel single nucleotide polymorphism (SNP) loci were identified in the CYP21A2 gene which seems to be specific for the Tunisian population. The overall concordance between genotype and phenotype was 98%. With this study the molecular basis of CAH has been characterized, providing useful results for clinicians in terms of prediction of disease severity, genetic and prenatal counseling.     

Congenital heart defect and mental retardation in a patient with a 13q33.1-34 deletion

13q deletion syndrome is a rare genetic disorder caused by deletions of the long arm of chromosome 13. Patients with 13q deletion display a variety of phenotypic features. We describe a one-year-old female patient with congenital heart defects (CHD), facial anomalies, development and mental retardation. We identified a 12.75Mb deletion in chromosome region 13q33.1-34 with high resolution SNP Array (Human660W-Quad, Illumina, USA). This chromosome region contains about 55 genes, including EFNB2, ERCC5, VGCNL1, F7, and F10. Comparing our findings with previously reported 13q deletion patients with congenital heart defects, we propose that the 13q33.1-34 deletion region might contain key gene(s) associated with cardiac development. Our study also identified a subclinical deficiency of Factors VII and X in our patient with Group 3 of 13q deletion syndrome.     

Identification of congenital muscular dystonia 2 associated with an inherited GlyT2 defect in Belgian Blue cattle from the United Kingdom

Two newborn Belgian Blue calves from a farm in the United Kingdom exhibited lateral recumbency, low head carriage and transient muscle spasms following tactile or auditory stimulation. DNA sequence analysis indicated that both calves were homozygous for the recessive congenital muscular dystonia type 2 (CMD2) mutation (c.809T>C, p.Leu270Pro) in SLC6A5, encoding the neuronal glycine transporter GlyT2. Further testing of animals from the index farm and a sample of Belgian Blue sires revealed an unexpectedly high frequency of CMD2 carriers. This implies that linked quantitative trait loci may be influencing the prevalence of CMD2 in the estimated 55,000 Belgian Blue cattle in the United Kingdom. We have therefore developed new inexpensive tests for the CMD2 allele that can be used to confirm diagnosis, identify carriers and guide future breeding strategy, thus avoiding animal distress/premature death and minimizing the future economic impact of this disorder.     

A spectrum of novel NPHS1 and NPHS2 gene mutations in pediatric nephrotic syndrome patients from Pakistan

Background

Mutations in the NPHS1 and NPHS2 genes are among the main causes of early-onset and familial steroid resistant nephrotic syndrome respectively. This study was carried out to assess the frequencies of mutations in these two genes in a cohort of Pakistani pediatric NS patients.

Methods

Mutation analysis was carried out by direct sequencing of the NPHS1 and NPHS2 genes in 145 nephrotic syndrome (NS) patients. This cohort included 36 samples of congenital or infantile onset NS cases and 39 samples of familial cases obtained from 30 families.

Results

A total of 7 homozygous (6 novel) mutations were found in the NPHS1 gene and 4 homozygous mutations in the NPHS2 gene. All mutations in the NPHS1 gene were found in the early onset cases. Of these, one patient has a family history of NS. Homozygous p.R229Q mutation in the NPHS2 gene was found in two children with childhood-onset NS.

Conclusions

Our results show a low prevalence of disease causing mutations in the NPHS1 (22% early onset, 5.5% overall) and NPHS2 (3.3% early onset and 3.4% overall) genes in the Pakistani NS children as compared to the European populations. In contrast to the high frequency of the NPHS2 gene mutations reported for familial SRNS in Europe, no mutation was found in the familial Pakistani cases. To our knowledge, this is the first comprehensive screening of the NPHS1 and NPHS2 gene mutations in sporadic and familial NS cases from South Asia.     

Agnathia and associated malformations in a male rhesus monkey

BACKGROUND: Agnathia is a rare malformation characterized by the absence of the mandible. METHODS: A male rhesus monkey with malformations was found dead and studied by internal examination, radiographs and histopathology. RESULTS: A case of a rare first branchial arch anomaly with agenesis of the mandible and tongue is presented. The animal also had visceral deformities. However, ears were normal in shape and only slightly low in position. The craniofacial malformations may reflect incomplete separation of the first branchial arch into its maxillary and mandibular processes. CONCLUSIONS: The association between the craniofacial and other corporal anomalies is unclear.