The study of inherited retinal diseases has advanced our knowledge of the cellular and molecular mechanisms involved in sensory neural signaling. Dysfunction of two specific sensory modalities, vision and proprioception, characterizes the phenotype of the rare, autosomal-recessive disorder posterior column ataxia and retinitis pigmentosa (PCARP). Using targeted DNA capture and high-throughput sequencing, we analyzed the entire 4.2 Mb candidate sequence on chromosome 1q32 to find the gene mutated in PCARP in a single family. Employing comprehensive bioinformatic analysis and filtering, we identified a single-nucleotide coding variant in the feline leukemia virus subgroup C cellular receptor 1 (FLVCR1), a gene encoding a heme-transporter protein. Sanger sequencing confirmed the FLVCR1 mutation in this family and identified different homozygous missense mutations located within the protein’s transmembrane channel segment in two other unrelated families with PCARP. To determine whether the selective pathologic features of PCARP correlated with FLVCR1 expression, we examined wild-type mouse Flvcr1 mRNA levels in the posterior column of the spinal cord and the retina via quantitative real-time reverse-transcriptase PCR. The Flvcr1 mRNA levels were most abundant in the retina, followed by the posterior column of the spinal cord and other brain regions. These results suggest that aberrant FLVCR1 causes a selective degeneration of a subpopulation of neurons in the retina and the posterior columns of the spinal cord via dysregulation of heme or iron homeostasis. This finding broadens the molecular basis of sensory neural signaling to include common mechanisms that involve proprioception and vision.

Bardet-Biedl syndrome (BBS) is a multisystem genetically heterogeneous disorder, the clinical features of which are largely the consequence of ciliary dysfunction. BBS is typically inherited in an autosomal recessive fashion, and mutations in at least 14 genes have been identified. Here, we report the identification of a founder mutation in the BBS2 gene as the cause for the increased incidence of this developmental disorder in the Hutterite population. To ascertain the Hutterite BBS locus, we performed a genome-wide single nucleotide polymorphism (SNP) analysis on a single patient and his three unaffected siblings from a Hutterite family. The analysis identified two large SNP blocks that were homozygous in the patient but not in his unaffected siblings, one of these regions contained the BBS2 gene. Sequence analysis and subsequent RNA studies identified and confirmed a novel splice site mutation, c.472-2A>G, in BBS2. This mutation was also found in homozygous form in three subsequently studied Hutterite BBS patients from two different leuts, confirming that this is a founder mutation in the Hutterite population. Further studies are required to determine the frequency of this mutation and its role, if any, in the expression of other ciliopathies in this population.

The Hutterites are a genetically isolated Anabaptist group living on the North American prairies; their population numbers over 40,000, the majority of whom are descendants of 89 founders. An autosomal recessive developmental disorder was identified in four patients from two consanguineous Hutterite families. To our knowledge the clinical presentation is unique and undescribed. The patients have distinctive facial features, congenital malformations of the heart and genitourinary system, head circumference at the 2nd centile and developmental delay. The facial features include tall forehead with high anterior hairline, deep-set eyes with short, upslanted palpebral fissures, long nose with low-hanging columella, and thick vermilion of the upper and lower lip. Karyotype and baseline metabolic studies were normal. An identity-by-descent mapping approach was used to localize the gene for this disorder. The patients were genotyped using Affymetrix GeneChip Human Mapping 10 K (Xba 2.0) and 50 K (Xba 240) Arrays which identified a single 5.5 Mb homozygous region at chromosome 16p13.3. To confirm and refine the boundaries of this region, microsatellite markers were used to genotype the patients, their parents, and the available unaffected siblings. The disease locus was refined to a region of 5.1 Mb containing 173 known or predicted genes. No other recessive disorders with similar clinical features are currently mapped to this region. The coding regions of over fifteen genes, prioritized by microarray expression analysis and information available in public databases, have been sequenced, but no potential pathogenic mutations have been identified. The identification of the gene for this syndrome will provide new insights into development and learning.

Branched-chain ketoacid dehydrogenase deficiency results in complex and volatile metabolic derangements that threaten brain development. Treatment for classical maple syrup urine disease (MSUD) should address this underlying physiology while also protecting children from nutrient deficiencies. Based on a 20-year experience managing 79 patients, we designed a study formula to (1) optimize transport of seven amino acids (Tyr, Trp, His, Met, Thr, Gln, Phe) that compete with branched-chain amino acids (BCAAs) for entry into the brain via a common transporter (LAT1), (2) compensate for episodic depletions of glutamine, glutamate, and alanine caused by reverse transamination, and (3) correct deficiencies of omega-3 essential fatty acids, zinc, and selenium widespread among MSUD patients. The formula was enriched with LAT1 amino acid substrates, glutamine, alanine, zinc, selenium, and alpha-linolenic acid (18:3n-3). Fifteen Old Order Mennonite children were started on study formula between birth and 34 months of age and seen at least monthly in the office. Amino acid levels were checked once weekly and more often during illnesses. All children grew and developed normally over a period of 14-33 months. Energy demand, leucine tolerance, and protein accretion were tightly linked during periods of normal growth. Rapid shifts to net protein degradation occurred during illnesses. At baseline, most LAT1 substrates varied inversely with plasma leucine, and their calculated rates of brain uptake were 20-68% below normal. Treatment with study formula increased plasma concentrations of LAT1 substrates and normalized their calculated uptakes into the nervous system. Red cell membrane omega-3 polyunsaturated fatty acids and serum zinc and selenium levels increased on study formula. However, selenium and docosahexaenoic acid (22:6n-3) levels remained below normal. During the study period, hospitalizations decreased from 0.35 to 0.14 per patient per year. There were 28 hospitalizations managed with MSUD hyperalimentation solution; 86% were precipitated by common infections, especially vomiting and gastroenteritis. The large majority of catabolic illnesses were managed successfully at home using ‘sick-day’ formula and frequent amino acid monitoring. We conclude that the study formula is safe and effective for the treatment of classical MSUD. In principle, dietary enrichment protects the brain against deficiency of amino acids used for protein accretion, neurotransmitter synthesis, and methyl group transfer. Although the pathophysiology of MSUD can be addressed through rational formula design, this does not replace the need for vigilant clinical monitoring, frequent measurement of the complete amino acid profile, and ongoing dietary adjustments that match nutritional intake to the metabolic demands of growth and illness.

Ubiquitin ligases play an important role in the regulation of the immune system. Absence of Itch E3 ubiquitin ligase in mice has been shown to cause severe autoimmune disease. Using autozygosity mapping in a large Amish kindred, we identified a linkage region on chromosome 20 and selected candidate genes for screening. We describe, in ten patients, identification of a mutation resulting in truncation of ITCH. These patients represent the first reported human phenotype associated with ITCH deficiency. These patients not only have multisystem autoimmune disease but also display morphologic and developmental abnormalities. This disorder underscores the importance of ITCH ubiquitin ligase in many cellular processes.

We identified an autosomal recessive condition in 11 individuals in the Old Order Amish of northeastern Ohio. The syndrome was characterized by distinctive craniofacial dysmorphism, skeletal anomalies, and mental retardation. The typical craniofacial dysmorphism included brachycephaly, highly arched bushy eyebrows, synophrys, long eyelashes, low-set ears, microdontism of primary teeth, and generalized gingival hyperplasia, whereas Sprengel deformity of scapula, fusion of spine, rib abnormities, pectus excavatum, and pes planus represented skeletal anomalies. The genome-wide homozygosity mapping using six affected individuals localized the disease gene to a 3.3-Mb region on chromosome 1q23.3-q24.1. Candidate gene sequencing identified a homozygous frameshift mutation, c.139_140delAG, in the transmembrane and coiled-coil domains 1 (TMCO1) gene, as the pathogenic change in all affected members of the extended pedigree. This mutation is predicted to result in a severely truncated protein (p.Ser47Ter) of only one-fourth the original length. The TMCO1 gene product is a member of DUF841 superfamily of several eukaryotic proteins with unknown function. The gene has highly conserved amino acid sequence and is universally expressed in all human tissues examined. The high degree of conservation and the ubiquitous expression pattern in human adult and fetal tissues suggest a critical role for TMCO1. This report shows a TMCO1 sequence variant being associated with a genetic disorder in human. We propose “TMCO1 defect syndrome” as the name of this condition.

In glutaric aciduria type 1, glutaryl-coenzyme A and its derivatives are produced from intracerebral lysine and entrapped at high concentrations within the brain, where they interfere with energy metabolism. Biochemical toxicity is thought to trigger stroke-like striatal degeneration in susceptible children under 2 years of age. Here, we explore vascular derangements that might also contribute to brain damage. We studied injured and non-injured Amish glutaric aciduria type 1 patients using magnetic resonance imaging (n = 26), transcranial Doppler ultrasound (n = 35) and perfusion computed tomography (n = 6). All glutaric aciduria type 1 patients had wide middle cerebral, internal carotid and basilar arteries. In non-injured patients, middle cerebral artery velocities were 18-26% below control values throughout late infancy and early childhood, whereas brain-injured children had an early velocity peak (18 months) and low values thereafter. Perfusion scans from six patients showed that tissue blood flow did not undergo a normal developmental surge. We observed four different perfusion patterns. (i) Three children (two non-injured) had low cerebral blood flow, prolonged mean transit time, elevated cerebral blood volume and high mean transit time/cerebral blood flow and cerebral blood volume/cerebral blood flow ratios. This pattern optimizes substrate extraction at any given flow rate but indicates low perfusion pressure and limited autoregulatory reserve. (ii) Ten hours after the onset of striatal necrosis in an 8-month-old infant, mean transit time and cerebral blood volume were low relative to cerebral blood flow, which varied markedly from region to region. This pattern indicates disturbed autoregulation, regional perfusion pressure gradients, or redistribution of flow from functional capillaries to non-exchanging vessels. (iii) In an infant with atrophic putaminal lesions, striatal flow was normal but mean transit time and cerebral blood volume were low, consistent with perfusion in excess of metabolic demand. (iv) Finally, a brain-injured adult with glutaric aciduria type 1 had regional perfusion values within the normal range, but the putamina, which normally have the highest regional perfusion, had cerebral blood flow values 24% below cortical grey matter. Although metabolic toxicity appears central to the pathophysiology of striatal necrosis, cerebrovascular changes probably also contribute to the process. These changes may be the primary cause of expanded cerebrospinal fluid volume in newborns, intracranial and retinal haemorrhages in infants and interstitial white matter oedema in children and adults. This pilot study suggests important new areas for clinical investigation.

The structure of the human glutaryl coenzyme A dehydrogenase (GCD) gene was determined to contain 11 exons and to span approximately 7 kb. Fibroblast DNA from 64 unrelated glutaric acidemia type I (GA1) patients was screened for mutations by PCR amplification and analysis of SSCP. Fragments with altered electrophoretic mobility were subcloned and sequenced to detect mutations that caused GA1. This report describes the structure of the GCD gene, as well as point mutations and polymorphisms found in 7 of its 11 exons. Several mutations were found in more than one patient, but no one prevalent mutation was detected in the general population. As expected from pedigree analysis, a single mutant allele causes GA1 in the Old Order Amish of Lancaster County, Pennsylvania. Several mutations have been expressed in Escherichia coli, and all produce diminished enzyme activity. Reduced activity in GCD encoded by the A421V mutation in the Amish may be due to impaired association of enzyme subunits.