FDXR variants cause adrenal insufficiency and atypical sexual development.

Genetic defects affecting steroid biosynthesis cause cortisol deficiency and differences of sex development; among them recessive mutations in the steroidogenic enzymes CYP11A1 and CYP11B, whose function is supported by reducing equivalents donated by ferredoxin reductase (FDXR) and ferredoxin. So far, mutations in the mitochondrial flavoprotein FDXR have been associated with a progressive neuropathic mitochondriopathy named FDXR-Related Mitochondriopathy (FRM), but cortisol insufficiency has not been documented. However, FRM patients often experience worsening or demise following stress associated with infections. We investigated two female FRM patients carrying the novel homozygous FDXR mutation p.G437R with ambiguous genitalia at birth and sudden death in the first year of life; they presented with cortisol deficiency and androgen excess compatible with 11-hydroxylase deficiency. In addition, steroidogenic FDXR-variant cell lines reprogrammed from three FRM patients' fibroblasts displayed deficient mineralocorticoid and glucocorticoid production. Finally, Fdxr-mutant mice allelic to the severe p.R386W human variant, showed reduced progesterone and corticosterone production. Therefore, our comprehensive studies show that human FDXR variants may cause compensated, but possibly life-threatening adrenocortical insufficiency in stress by affecting adrenal glucocorticoid and mineralocorticoid synthesis through direct enzyme inhibition, most likely in combination with disturbed mitochondrial redox balance.

TRAPPC6B biallelic variants cause a neurodevelopmental disorder with TRAPP II and trafficking disruptions.

Highly conserved transport protein particle (TRAPP) complexes regulate subcellular trafficking pathways. Accurate protein trafficking has been increasingly recognized to be critically important for normal development, particularly in the nervous system. Variants in most TRAPP complex subunits have been found to lead to neurodevelopmental disorders with diverse but overlapping phenotypes. We expand on limited prior reports on TRAPPC6B with detailed clinical and neuroradiologic assessments, and studies on mechanisms of disease, and new types of variants. We describe 29 additional patients from 18 independent families with biallelic variants in TRAPPC6B. We identified seven homozygous nonsense (n = 12 patients) and eight canonical splice-site variants (n = 17 patients). In addition, we identified one patient with compound heterozygous splice-site/missense variants with a milder phenotype and one patient with homozygous missense variants. Patients displayed non-progressive microcephaly, global developmental delay/intellectual disability, epilepsy and absent expressive language. Movement disorders including stereotypies, spasticity and dystonia were also observed. Brain imaging revealed reductions in cortex, cerebellum and corpus callosum size with frequent white matter hyperintensity. Volumetric measurements indicated globally diminished volume rather than specific regional losses. We identified a reduced rate of trafficking into the Golgi apparatus and Golgi fragmentation in patient-derived fibroblasts that was rescued by wild-type TRAPPC6B. Molecular studies revealed a weakened interaction between mutant TRAPPC6B (c.454C>T, p.Q152*) and its TRAPP binding partner TRAPPC3. Patient-derived fibroblasts from the TRAPPC6B (c.454C>T, p.Q152*) variant displayed reduced levels of TRAPPC6B as well as other TRAPP II complex-specific members (TRAPPC9 and TRAPPC10). Interestingly, the levels of the TRAPPC6B homologue TRAPPC6A were found to be elevated. Moreover, co-immunoprecipitation experiments showed that TRAPPC6A co-precipitates equally with TRAPP II and TRAPP III, while TRAPPC6B co-precipitates significantly more with TRAPP II, suggesting enrichment of the protein in the TRAPP II complex. This implies that variants in TRAPPC6B may preferentially affect TRAPP II functions compared to TRAPP III functions. Finally, we assessed phenotypes in a Drosophila TRAPPC6B-deficiency model. Neuronal TRAPPC6B knockdown impaired locomotion and led to wing posture defects, supporting a role for TRAPPC6B in neuromotor function. Our findings confirm the association of damaging biallelic TRAPPC6B variants with microcephaly, intellectual disability, language impairments, and epilepsy. A subset of patients also exhibited dystonia and/or spasticity with impaired ambulation. These features overlap with disorders arising from pathogenic variants in other TRAPP subunits, particularly components of the TRAPP II complex. These findings suggest that TRAPPC6B is essential for brain development and function, and TRAPP II complex activity may be particularly relevant for mediating this function.

A transposase-derived gene required for human brain development.

DNA transposable elements and transposase-derived genes are present in most living organisms, including vertebrates, but their function is largely unknown. PiggyBac Transposable Element Derived 5 (PGBD5) is an evolutionarily conserved vertebrate DNA transposase-derived gene with retained nuclease activity in cells. Vertebrate brain development is known to be associated with prominent neuronal cell death and DNA breaks, but their causes and functions are not well understood. Here, we show that PGBD5 contributes to normal brain development in mice and humans, where its deficiency causes disorder of intellectual disability, movement and seizures. In mice, Pgbd5 is required for the developmental induction of post-mitotic DNA breaks and recurrent somatic genome rearrangements in neurons. Together, these studies nominate PGBD5 as the long-hypothesized neuronal DNA nuclease required for brain function in mammals.

ATP6V0C variants impair V-ATPase function causing a neurodevelopmental disorder often associated with epilepsy.

The vacuolar H+-ATPase is an enzymatic complex that functions in an ATP-dependent manner to pump protons across membranes and acidify organelles, thereby creating the proton/pH gradient required for membrane trafficking by several different types of transporters. We describe heterozygous point variants in ATP6V0C, encoding the c-subunit in the membrane bound integral domain of the vacuolar H+-ATPase, in 27 patients with neurodevelopmental abnormalities with or without epilepsy. Corpus callosum hypoplasia and cardiac abnormalities were also present in some patients. In silico modelling suggested that the patient variants interfere with the interactions between the ATP6V0C and ATP6V0A subunits during ATP hydrolysis. Consistent with decreased vacuolar H+-ATPase activity, functional analyses conducted in Saccharomyces cerevisiae revealed reduced LysoSensor fluorescence and reduced growth in media containing varying concentrations of CaCl2. Knockdown of ATP6V0C in Drosophila resulted in increased duration of seizure-like behaviour, and the expression of selected patient variants in Caenorhabditis elegans led to reduced growth, motor dysfunction and reduced lifespan. In summary, this study establishes ATP6V0C as an important disease gene, describes the clinical features of the associated neurodevelopmental disorder and provides insight into disease mechanisms.

A Novel Variant of ATP5MC3 Associated with Both Dystonia and Spastic Paraplegia.

BACKGROUND: In a large pedigree with an unusual phenotype of spastic paraplegia or dystonia and autosomal dominant inheritance, linkage analysis previously mapped the disease to chromosome 2q24-2q31. OBJECTIVE: The aim of this study is to identify the genetic cause and molecular basis of an unusual autosomal dominant spastic paraplegia and dystonia. METHODS: Whole exome sequencing following linkage analysis was used to identify the genetic cause in a large family. Cosegregation analysis was also performed. An additional 384 individuals with spastic paraplegia or dystonia were screened for pathogenic sequence variants in the adenosine triphosphate (ATP) synthase membrane subunit C locus 3 gene (ATP5MC3). The identified variant was submitted to the "GeneMatcher" program for recruitment of additional subjects. Mitochondrial functions were analyzed in patient-derived fibroblast cell lines. Transgenic Drosophila carrying mutants were studied for movement behavior and mitochondrial function. RESULTS: Exome analysis revealed a variant (c.318C > G; p.Asn106Lys) (NM_001689.4) in ATP5MC3 in a large family with autosomal dominant spastic paraplegia and dystonia that cosegregated with affected individuals. No variants were identified in an additional 384 individuals with spastic paraplegia or dystonia. GeneMatcher identified an individual with the same genetic change, acquired de novo, who manifested upper-limb dystonia. Patient fibroblast studies showed impaired complex V activity, ATP generation, and oxygen consumption. Drosophila carrying orthologous mutations also exhibited impaired mitochondrial function and displayed reduced mobility. CONCLUSION: A unique form of familial spastic paraplegia and dystonia is associated with a heterozygous ATP5MC3 variant that also reduces mitochondrial complex V activity.

Fetal brain small vessel disease 1 caused by a novel mutation in the COL4A1 gene.

A singleton fetus was referred to fetal magnetic resonance imaging (MRI) at 25 weeks due to mild ventriculomegaly and an abnormal fetal echocardiogram showing cardiomegaly, right ventricular hypertrophy and tricuspid insufficiency. Patchy areas of ischemic infarction, extensive subacute and chronic hemorrhage not respecting vascular territories, encephaloclastic cysts and closed lip schizencephaly were identified. Cataract was detected postnatally. The anomalies were caused by a pathogenic mutation (c.353 G>A; p.G118D) in the COL4A1 gene. The phenotype seen in this case, i.e. small vessel cerebral disease with or without ocular anomalies caused by COL4A1 mutations, is likely an underrecognized cause of perinatal stroke. The pattern of abnormalities reported herein should prompt strong consideration for diagnosis and molecular testing.

Central Diabetes Insipidus in a Patient With NFKB2 Mutation: Expanding the Endocrine Phenotype in DAVID Syndrome.

CONTEXT: Deficient anterior pituitary with variable immune deficiency (DAVID) syndrome is a recently described, rare disorder characterized by anterior pituitary hormone deficiencies and common variable immunodeficiency associated with NFKB2 mutations. Posterior pituitary hormone deficiencies have not been reported in patients with DAVID syndrome. CASE DESCRIPTION: We report a pediatric patient who initially presented with hypogammaglobulinemia and alopecia totalis, who was identified to have a de novo NFKB2 mutation at one year of age. He developed central diabetes insipidus and central adrenal insufficiency at three and four years of age, respectively. At seven years of age, he had not developed GH or TSH deficiencies. Whole exome sequencing ruled out known genetic causes of central diabetes insipidus, adrenal insufficiency, and hypopituitarism. CONCLUSION: This is a report of central diabetes insipidus in a patient with DAVID syndrome caused by an NFKB2 mutation. This case report expands the evolving endocrine phenotype associated with NFKB2 mutations beyond anterior pituitary deficiencies.

The Clinical Utility of a Single-Nucleotide Polymorphism Microarray in Patients With Epilepsy at a Tertiary Medical Center.

Microarray testing has revolutionized clinical cytogenetics, as it provides a significantly higher resolution and greater clinical yield than karyotype analysis. This study assessed the clinical utility of single-nucleotide polymorphism microarray in patients with epilepsy. Study subjects were patients between the ages of birth to 23 years who were diagnosed with epilepsy and had a microarray performed at Cincinnati Children's Hospital Medical Center. Statistical analysis explored the association of microarray results and brain magnetic resonance imaging (MRI), seizure type, and structural malformations. Approximately 17.7% (26/147) of participants had an abnormal microarray as defined by laboratory guidelines. There were no differences in frequency of abnormal brain MRI or seizure type between the abnormal and normal microarray groups. There was a higher prevalence of musculoskeletal malformations (P < .0035) and cardiovascular malformations (P < .0081) in subjects with abnormal microarrays. Clinicians should consider microarray analysis in individuals who have epilepsy, especially in combination with musculoskeletal malformation or cardiovascular malformation.

Activating PIK3CA alleles and lymphangiogenic phenotype of lymphatic endothelial cells isolated from lymphatic malformations.

Lymphatic malformations (LMs) are developmental anomalies of the lymphatic system associated with the dysmorphogenesis of vascular channels lined by lymphatic endothelial cells (LECs). Seeking to identify intrinsic defects in affected LECs, cells were isolated from malformation tissue or fluid on the basis of CD31 and podoplanin (PDPN) expression. LECs from five unrelated LM lesions were characterized, including cells derived from one patient previously diagnosed with CLOVES. CLOVES-related LECs carried a known, activating mutation in PIK3CA (p.H1047L), confirmed by direct sequencing. Activating PIK3CA mutations (p.E542K and p.E545A) were identified in lesion-derived cells from the other four patients, also by direct sequencing. The five LM-LEC cultures shared a lymphangiogenic phenotype distinguished by PI3K/AKT activation, enhanced sprouting efficiency, elevated VEGF-C expression and COX2 expression, shorter doubling times and reduced expression of angiopoietin 2 and CXCR4. Nine additional LM-LEC populations and 12 of 15 archived LM tissue samples were shown to bear common PIK3CA variants by allele-specific PCR. The activation of a central growth/survival pathway (PI3K/AKT) represents a feasible target for the non-invasive treatment of LMs bearing in mind that background genetics may individualize lesions and influence treatments.

Familial 9q22.3 microduplication spanning PTCH1 causes short stature syndrome with mild intellectual disability and dysmorphic features.

Partial trisomy 9q involving the duplication of band 9q22 is manifested by a constellation of symptoms including short stature, intellectual disability, microcephaly, pyloric stenosis, facial dysmorphism, and various defects of the heart, distal extremities, eyes, thyroid, and esophagus. In three family members with growth retardation, mild intellectual disability, and mild facial dysmorphism, array-based comparative genomic hybridization analyses showed a familial microduplication at 9q22.3. On the basis of the described functions of the duplicated genes, PTCH1 represents a candidate gene that may be responsible for the phenotypic findings, although the 14 other genes in this duplicated segment may also contribute to the phenotype. The current report provides evidence to support a specific phenotype associated with a 9q22.3 microduplication and confirm localization of a subset of the trisomy 9q phenotype to this chromosomal region.

Acute necrotizing encephalopathy in 3 brothers.

Acute necrotizing encephalopathy (ANE) is a devastating and rapidly progressive neurologic disorder that occurs in healthy children after common viral infections. Typically, ANE is sporadic and does not recur. However, familial (ANE1) and recurrent cases have been reported and were recently linked to mutations in RANBP2 (RAN-binding protein 2). We report here a multiply affected kindred with recurrent familial ANE. These affected male siblings (a set of twins and their older brother) all presented with prodromal fever and upper respiratory tract infection that progressed within 72 hours to seizures, coma, and ultimately death, a course that is typical of ANE. It should be noted that 1 brother was treated with early aggressive management, including corticosteroids, and he survived for an additional 5 years. This represents the second reported case of familial ANE in the United States and the only case of male siblings with consanguineous parents. We hope that early recognition and growing awareness can lead to more effective treatment and better outcomes in the future.

The interplay of infection and genetics in acute necrotizing encephalopathy.

PURPOSE OF REVIEW: Acute necrotizing encephalopathy (ANE) presents with fulminant encephalopathy and characteristic brain lesions following viral infection. The rarity and unpredictability of the disorder have significantly impaired its study. Growing recognition of ANE and the discovery of causative missense mutations in the nuclear pore gene RANBP2 give promising steps toward unraveling this disease. This review summarizes recent advances of clinical and scientific understanding of ANE. RECENT FINDINGS: Inflammatory factors participate in the pathogenesis of ANE, but the lack of difference between influenza and noninfluenza ANE focuses attention on the abnormal host response as causative. Early treatment with steroids provides the best outcome for patients who do not have brainstem lesions. Missense mutations in RANBP2 cause the majority of familial and recurrent ANE cases, but other single-gene causes of ANE are possible for familial, recurrent, and sporadic cases. SUMMARY: Early recognition and systematic evaluation of ANE are necessary. Modeling ANE as a genetic disorder may provide the most immediate gains in the understanding and treatment of ANE and related disorders.

Recurrent acute necrotizing encephalopathy following influenza A in a genetically predisposed family.

Acute necrotizing encephalopathy (ANE) typically affects young, healthy children who develop rapid-onset severe encephalopathy triggered by viral infections. This disease is more commonly reported in Japan but occurs worldwide, although it remains under-recognized in Western countries. An autosomal dominant form, ANE1, was recently identified. We report the details of a 9-year-old Caucasian female who experienced recurrent ANE episodes at the ages of 9 months and 9 years. Brain magnetic resonance imaging findings were characteristic of ANE during both episodes, although more extensive in the recent episode, which resulted in severe neurological sequelae; influenza A was identified on bronchoalveolar lavage during this episode. Interestingly, there was evidence of peripheral polyneuropathy during the recent episode, which has not previously been described in sporadic ANE. Both the patient and her mother, who had also had postviral polyneuritis in the past, harbour a mutation in Ran-binding protein 2 (RANBP2); this occurred de novo in the mother and confers genetic susceptibility to ANE. Our case suggests that recurrent disease and/or an expanded clinical phenotype raises the possibility of ANE1; positive family history, although supportive, is not necessary as the mutation can occur de novo. Increased awareness may lead to earlier recognition and better treatment options.

Infection-triggered familial or recurrent cases of acute necrotizing encephalopathy caused by mutations in a component of the nuclear pore, RANBP2.

Acute necrotizing encephalopathy (ANE) is a rapidly progressive encephalopathy that can occur in otherwise healthy children after common viral infections such as influenza and parainfluenza. Most ANE is sporadic and nonrecurrent (isolated ANE). However, we identified a 7 Mb interval containing a susceptibility locus (ANE1) in a family segregating recurrent ANE as an incompletely penetrant, autosomal-dominant trait. We now report that all affected individuals and obligate carriers in this family are heterozygous for a missense mutation (c.1880C-->T, p.Thr585Met) in the gene encoding the nuclear pore protein Ran Binding Protein 2 (RANBP2). To determine whether this mutation is the susceptibility allele, we screened controls and other patients with ANE who are unrelated to the index family. Patients from 9 of 15 additional kindreds with familial or recurrent ANE had the identical mutation. It arose de novo in two families and independently in several other families. Two other patients with familial ANE had different RANBP2 missense mutations that altered conserved residues. None of the three RANBP2 missense mutations were found in 19 patients with isolated ANE or in unaffected controls. We conclude that missense mutations in RANBP2 are susceptibility alleles for familial and recurrent cases of ANE.

Cohen syndrome in the Ohio Amish.

We describe eight members from two large Amish kindreds who share a phenotype characterized by early-onset pigmentary retinopathy and myopia, global developmental delay and mental retardation, microcephaly, short stature, hypotonia, joint hyperextensibility, small hands and feet, common facial appearance, and friendly disposition. Several of the children had intermittent granulocytopenia. The phenotypic occurrence in three siblings coupled with the increased coefficient of inbreeding in the Amish suggested that this disorder is autosomal recessive and due to a single founder allele. Despite similarity to the clinical features of Cohen syndrome, experienced dysmorphologists attending the 23rd David W. Smith Workshop suggested the facial gestalt of the Amish children was inconsistent with this diagnosis. We mapped the locus responsible for these individuals' phenotype to chromosome 8q22-q23, which contains the recently discovered Cohen syndrome gene, COH1. Complete sequencing of the COH1 gene identified a likely disease-causing frameshift mutation and a missense mutation in the Amish patients. A comparison of features among different Cohen syndrome populations with shared linkage to the COH1 locus or known COH1 gene mutations may allow for the determination of improved clinical criteria on which to suspect the diagnosis of Cohen syndrome. We conclude that facial gestalt seems to be an unreliable indicator of Cohen syndrome between ethnic populations, although it is quite consistent among affected individuals within a particular ethnic group. Other features common to almost all individuals with proven COH1 mutations, such as retinal dystrophy, myopia, microcephaly, mental retardation, global developmental delay, hypotonia, and joint hyperextensibility appear to be better clinical indicators of this disorder.

Autosomal dominant acute necrotizing encephalopathy maps to 2q12.1-2q13.

In autosomal dominant acute necrotizing encephalopathy (ADANE), apparently healthy children develop necrotizing lesions in their thalami and brainstems in the course of febrile illnesses. We used DNA from affected subjects and obligate carriers to map ADANE to a 6.5Mb region on chromosome 2. Sequencing of four candidate genes in the interval (BCL2L11, ST6GalII, CHT1, and FLJ20019), involved in apoptosis, viral recognition, choline transport, and electron transport, showed no disease causing mutations.

Mixed clefting type in Rapp-Hodgkin syndrome.

Mixed clefting type (MCT) is the rare occurrence of cleft lip, with or without cleft palate, and cleft palate alone in the same pedigree. Here we present a family with Rapp-Hodgkin syndrome (RHS) that manifests MCT, and use this rare finding to suggest that RHS may be related not only to phenotypically similar syndromes, but seemingly dissimilar ones as well. RHS has obvious phenotypic overlap with other ectodermal dysplasia-clefting syndromes (EDCS), such as ectrodactyly-ectodermal dysplasia-clefting syndrome (EEC) and ankyloblepharon-ectodermal dysplasia-clefting syndrome (AEC), all of which show MCT. MCT is also found in the allelic disorders van der Woude syndrome (VDW) and popliteal-pterygium syndrome (PPS). Therefore, while VDW and PPS have little clinical overlap with the EDCS, the common finding of MCT may indicate closer relationships at the developmental or genetic level.