Michael Volodarsky

A Deletion Mutation in TMEM38B Associated with Autosomal Recessive Osteogenesis Imperfecta

Autosomal recessive osteogenesis imperfecta (OI) was diagnosed in three unrelated Israeli Bedouin consanguineous families. Fractures were evident in all cases in infancy. Genome‐wide linkage analysis ruled out association with any of the known OI genes, and identified a single homozygosity locus of approximately 2 Mb on chromosome 9 common to all affected individuals (maximum multipoint lod score 6.5). Whole exome sequencing identified only a single mutation within this locus that was shared by all affected individuals: a homozygous deletion mutation of exon 4 of TMEM38B, leading to an early stop codon and a truncated protein, as well as low TMEM38B mRNA levels. TMEM38B encodes TRIC‐B, a ubiquitous component of TRIC, a monovalent cation‐specific channel involved in Ca2+ release from intracellular stores that has been shown to act in cell differentiation. Molecular mechanisms through which a TMEM38B mutation might lead to an OI phenotype are yet to be explored.

Autosomal recessive Adams–Oliver syndrome caused by homozygous mutation in EOGT, encoding an EGF domain-specific O-GlcNAc transferase

Autosomal recessive Adams–Oliver syndrome was diagnosed in three remotely related Bedouin consanguineous families. Genome-wide linkage analysis ruled out association with known Adams–Oliver syndrome genes, identifying a single-homozygosity ∼1.8-Mb novel locus common to affected individuals (LOD score 3.37). Whole-exome sequencing followed by Sanger sequencing identified only a single mutation within this locus, shared by all affected individuals and found in patients from five additional apparently unrelated Bedouin families: a 1-bp deletion mutation in a predicted alternative splice variant of EOGT, leading to a putative truncated protein. RT-PCR demonstrated that the EOGT-predicted alternative splice variant is ubiquitously expressed. EOGT encodes EGF-domain-specific O-linked N-acetylglucosamine transferase, responsible for extracellular O-GlcNAcylation of epidermal growth factor-like domain-containing proteins, and is essential for epithelial cell-matrix interactions. F-actin staining in diseased fibroblasts showed apparently intact cell cytoskeleton and morphology, suggesting the EOGT mutation acts not through perturbation of cytoskeleton but through other mechanisms yet to be elucidated.

UNC80 mutation causes a syndrome of hypotonia, severe intellectual disability, dyskinesia and dysmorphism, similar to that caused by mutations in its interacting cation channel NALCN

Background A syndrome of profound hypotonia, intellectual disability, intrauterine growth retardation with subsequent failure to thrive, dyskinesia and epilepsy was diagnosed in Bedouin Israeli families. Mild dysmorphism was evident: plagiocephaly, broad forehead with prominent nose, smooth philtrum and congenital esotropia. We set out to decipher the molecular basis of this syndrome. Methods Genome-wide linkage analysis and fine mapping were done. Whole exome sequencing data were filtered for candidate variants within locus. Validation and segregation of the mutation was assayed via Sanger sequencing. UNC80 expression pattern was analysed through reverse transcription PCR. Results Homozygosity mapping followed by fine mapping identified a 7.5 Mb disease-associated locus (logarithm of odds score 3.5) on chromosome 2. Whole exome and Sanger sequencing identified a single homozygous nonsense mutation within this locus, segregating within the families as expected for recessive heredity and not found in a homozygous state in 150 Bedouin controls: c.151C>T, p.(R51*) in UNC80. Conclusions The syndrome described is caused by a mutation in UNC80, truncating most of the 3258 amino acids highly conserved encoded protein, that has no known motifs. UNC80 bridges between UNC79 and the cation channel NALCN, enabling NALCNs role in basal Na+ leak conductance in neurons, essential for neuronal function. The phenotype caused by the UNC80 mutation resembles that previously described for homozygous NALCN mutations.

Midtrimester triple test levels in women with severe preeclampsia and HELLP syndrome

Background.  The levels of midtrimester triple test constituents are known to be altered in hypertensive disorders of pregnancy.

SLC30A9 mutation affecting intracellular zinc homeostasis causes a novel cerebro-renal syndrome

Few cerebro-renal syndromes have been described to date. Perez et al. identify a novel autosomal recessive cerebro-renal syndrome in a consanguineous Bedouin kindred, caused by a mutation in SLC30A9. The mutation disrupts the role of SLC30A9 in Zn2+ transport, leading to impaired regulation of cytosolic zinc homeostasis.

ALFY-Controlled DVL3 Autophagy Regulates Wnt Signaling, Determining Human Brain Size

Primary microcephaly is a congenital neurodevelopmental disorder of reduced head circumference and brain volume, with fewer neurons in the cortex of the developing brain due to premature transition between symmetrical and asymmetrical cellular division of the neuronal stem cell layer during neurogenesis. We now show through linkage analysis and whole exome sequencing, that a dominant mutation in ALFY, encoding an autophagy scaffold protein, causes human primary microcephaly. We demonstrate the dominant effect of the mutation in drosophila: transgenic flies harboring the human mutant allele display small brain volume, recapitulating the disease phenotype. Moreover, eye-specific expression of human mutant ALFY causes rough eye phenotype. In molecular terms, we demonstrate that normally ALFY attenuates the canonical Wnt signaling pathway via autophagy-dependent removal specifically of aggregates of DVL3 and not of Dvl1 or Dvl2. Thus, autophagic attenuation of Wnt signaling through removal of Dvl3 aggregates by ALFY acts in determining human brain size.

Novel FAM20A mutation causes autosomal recessive amelogenesis imperfecta

OBJECTIVE To relate the peculiar phenotype of amelogenesis imperfecta in a large Bedouin family to the genotype determined by whole genome linkage analysis. DESIGN Amelogenesis imperfecta (AI) is a broad group of inherited pathologies affecting enamel formation, characterized by variability in phenotypes, causing mutations and modes of inheritance. Autosomal recessive or compound heterozygous mutations in FAM20A, encoding sequence similarity 20, member A, have been shown to cause several AI phenotypes. Five members from a large consanguineous Bedouin family presented with hypoplastic amelogenesis imperfecta with unerupted and resorbed permanent molars. Following Soroka Medical Center IRB approval and informed consent, blood samples were obtained from six affected offspring, five obligatory carriers and two unaffected siblings. Whole genome linkage analysis was performed followed by Sanger sequencing of FAM20A. RESULTS The sequencing unravelled a novel homozygous deletion mutation in exon 11 (c.1523delC), predicted to insert a premature stop codon (p.Thr508Lysfs*6). CONCLUSIONS We provide an interesting case of novel mutation in this rare disorder, in which the affected kindred is unique in the large number of family members sharing a similar phenotype.

Controlled ovarian hyperstimulation using multi-dose gonadotropin-releasing hormone (GnRH) antagonist results in less systemic inflammation than the GnRH-agonist long protocol.

Objective. The aim of the study was to investigate whether controlled ovarian hyperstimulation (COH) using multi-dose gonadotropin-releasing hormone (GnRH) antagonist results in a lesser degree of systemic inflammation than the GnRH-agonist long protocol. Design. Prospective, observational study. Patients and methods. Blood was drawn three times during the COH cycle from patients undergoing the long GnRH-agonist protocol (agonist group) (n = 12) or the multi-dose GnRH-antagonist protocol (antagonist group) (n = 15): the day on which adequate suppression was obtained (agonist group), or day 2 or 3 of the menstrual cycle and before gonadotropin treatment (antagonist group) (Day-0); the day of or prior to administration of human chorionic gonadotropin (Day-hCG); and the day of ovum pick-up (Day-OPU). Levels of sex steroids and serum C-reactive protein (CRP) were compared between the two study groups among the three time points. Results. While no between-group differences were observed in patient age or ovarian stimulation characteristics, a significantly higher number of oocytes were retrieved in the antagonist compared with the agonist group. In both groups, serum CRP levels were significantly higher on Day-OPU than on Day-hCG and Day-0. While serum CRP levels were higher on Day-hCG than Day-0, the difference was statistically significant only for the agonist group (p < 0.05). Moreover, Day-OPU serum CRP levels were significantly higher in the agonist than in the antagonist subgroup. Conclusion. COH using the multi-dose GnRH-antagonist protocol yields a lesser degree of systemic inflammation, as reflected by CRP levels, than the GnRH-agonist long protocol.

CDC174, a novel component of the exon junction complex whose mutation underlies a syndrome of hypotonia and psychomotor developmental delay

Siblings of non-consanguineous Jewish-Ethiopian ancestry presented with congenital axial hypotonia, weakness of the abducens nerve, psychomotor developmental delay with brain ventriculomegaly, variable thinning of corpus callosum and cardiac septal defects. Homozygosity mapping identified a single disease-associated locus of 3.5 Mb on chromosome 3. Studies of a Bedouin consanguineous kindred affected with a similar recessive phenotype identified a single disease-associated 18 Mb homozygosity locus encompassing the entire 3.5 Mb locus. Whole exome sequencing demonstrated only two homozygous mutations within a shared identical haplotype of 0.6 Mb, common to both Bedouin and Ethiopian affected individuals, suggesting an ancient common founder. Only one of the mutations segregated as expected in both kindreds and was not found in Bedouin and Jewish-Ethiopian controls: c.1404A>G, p.[*468Trpext*6] in CCDC174. We showed that CCDC174 is ubiquitous, restricted to the cell nucleus and co-localized with EIF4A3. In fact, yeast-two-hybrid assay demonstrated interaction of CCDC174 with EIF4A3, a component of exon junction complex. Knockdown of the CCDC174 ortholog in Xenopus laevis embryos resulted in poor neural fold closure at the neurula stage with later embryonic lethality. Knockdown embryos exhibited a sharp reduction in expression of n-tubulin, a marker for differentiating primary neurons, and of hindbrain markers krox20 and hoxb3. The Xenopus phenotype could be rescued by the human normal, yet not the mutant CCDC174 transcripts. Moreover, overexpression of mutant but not normal CCDC174 in neuroblastoma cells caused rapid apoptosis. In line with the hypotonia phenotype, the CCDC174 mutation caused depletion of RYR1 and marked myopathic changes in skeletal muscle of affected individuals.

Are Ultrasonographic Findings Suggestive of Ovarian Stromal Edema Associated with Ischemic Adnexal Torsion

Objective: To study whether sonographic findings suggestive of ovarian stromal edema are associated with tissue ischemia in patients with adnexal torsion. Methods: A study of 79 patients with adnexal torsion was performed. Patients were divided into an ischemic group, in which the twisted adnexa were seen as blue or black, and a non-ischemic group, in which the adnexa retained normal color and appeared viable. Clinical and ultrasonographic findings, specifically the presence of ultrasonographic signs suggestive of ovarian stromal edema, were compared between the two groups. Results: Of the 79 patients with torsion, in 44 (55.7%) the adnexa appeared ischemic at surgery. The presence of ischemia was not associated with age, pregnancy, duration of pain, vomiting or findings at physical examination. There was no significant difference between the ischemic and the non-ischemic group in the proportion of patients with signs of ovarian stromal edema (59 vs. 40%, p = 0.11), in the proportion of patients with absent/diminished stromal Doppler flow (36 vs. 28%, p = 0.12%) or in the proportion of patients with both signs of stroma edema and absent/diminished stromal Doppler flow (20 vs. 12%, p = 0.36). Conclusion: Ultrasonographic signs of ovarian stromal edema do not assist in differentiating between ischemic and non-ischemic adnexal torsion.