Mutations in LRP2, which encodes the multiligand receptor megalin, cause Donnai-Barrow and facio-oculo-acoustico-renal syndromes

Donnai-Barrow syndrome is associated with agenesis of the corpus callosum, congenital diaphragmatic hernia, facial dysmorphology, ocular anomalies, sensorineural hearing loss and developmental delay. By studying multiplex families, we mapped this disorder to chromosome 2q23.3-31.1 and identified LRP2 mutations in six families with Donnai-Barrow syndrome and one family with facio-oculo-acoustico-renal syndrome. LRP2 encodes megalin, a multiligand uptake receptor that regulates levels of diverse circulating compounds. This work implicates a pathway with potential pharmacological therapeutic targets.     

The transmembrane protein meckelin (MKS3) is mutated in Meckel-Gruber syndrome and the wpk rat

Meckel-Gruber syndrome is a severe autosomal, recessively inherited disorder characterized by bilateral renal cystic dysplasia, developmental defects of the central nervous system (most commonly occipital encephalocele), hepatic ductal dysplasia and cysts and polydactyly. MKS is genetically heterogeneous, with three loci mapped: MKS1, 17q21-24 (ref. 4); MKS2, 11q13 (ref. 5) and MKS3 (ref. 6). We have refined MKS3 mapping to a 12.67-Mb interval (8q21.13-q22.1) that is syntenic to the Wpk locus in rat, which is a model with polycystic kidney disease, agenesis of the corpus callosum and hydrocephalus. Positional cloning of the Wpk gene suggested a MKS3 candidate gene, TMEM67, for which we identified pathogenic mutations for five MKS3-linked consanguineous families. MKS3 is a previously uncharacterized, evolutionarily conserved gene that is expressed at moderate levels in fetal brain, liver and kidney but has widespread, low levels of expression. It encodes a 995-amino acid seven-transmembrane receptor protein of unknown function that we have called meckelin.     

Multiple apical plasma membrane constituents are associated with susceptibility to meconium ileus in individuals with cystic fibrosis

Variants associated with meconium ileus in cystic fibrosis were identified in 3,763 affected individuals by genome-wide association study (GWAS). Five SNPs at two loci near SLC6A14 at Xq23-24 (minimum P = 1.28 × 10(-12) at rs3788766) and SLC26A9 at 1q32.1 (minimum P = 9.88 × 10(-9) at rs4077468) accounted for ~5% of phenotypic variability and were replicated in an independent sample of affected individuals (n = 2,372; P = 0.001 and 0.0001, respectively). By incorporating the knowledge that disease-causing mutations in CFTR alter electrolyte and fluid flux across surface epithelium into a hypothesis-driven GWAS (GWAS-HD), we identified associations with the same SNPs in SLC6A14 and SLC26A9 and established evidence for the involvement of SNPs in a third solute carrier gene, SLC9A3. In addition, GWAS-HD provided evidence of association between meconium ileus and multiple genes encoding constituents of the apical plasma membrane where CFTR resides (P = 0.0002; testing of 155 apical membrane genes jointly and in replication, P = 0.022). These findings suggest that modulating activities of apical membrane constituents could complement current therapeutic paradigms for cystic fibrosis.     

Mutations in SPG11, encoding spatacsin, are a major cause of spastic paraplegia with thin corpus callosum

Autosomal recessive hereditary spastic paraplegia (ARHSP) with thin corpus callosum (TCC) is a common and clinically distinct form of familial spastic paraplegia that is linked to the SPG11 locus on chromosome 15 in most affected families. We analyzed 12 ARHSP-TCC families, refined the SPG11 candidate interval and identified ten mutations in a previously unidentified gene expressed ubiquitously in the nervous system but most prominently in the cerebellum, cerebral cortex, hippocampus and pineal gland. The mutations were either nonsense or insertions and deletions leading to a frameshift, suggesting a loss-of-function mechanism. The identification of the function of the gene will provide insight into the mechanisms leading to the degeneration of the corticospinal tract and other brain structures in this frequent form of ARHSP.     

Hartnup disorder is caused by mutations in the gene encoding the neutral amino acid transporter SLC6A19

Hartnup disorder (OMIM 234500) is an autosomal recessive abnormality of renal and gastrointestinal neutral amino acid transport noted for its clinical variability. We localized a gene causing Hartnup disorder to chromosome 5p15.33 and cloned a new gene, SLC6A19, in this region. SLC6A19 is a sodium-dependent and chloride-independent neutral amino acid transporter, expressed predominately in kidney and intestine, with properties of system B(0). We identified six mutations in SLC6A19 that cosegregated with disease in the predicted recessive manner, with most affected individuals being compound heterozygotes. The disease-causing mutations that we tested reduced neutral amino acid transport function in vitro. Population frequencies for the most common mutated SLC6A19 alleles are 0.007 for 517G --> A and 0.001 for 718C --> T. Our findings indicate that SLC6A19 is the long-sought gene that is mutated in Hartnup disorder; its identification provides the opportunity to examine the inconsistent multisystemic features of this disorder.     

Mutations in the gene encoding GlyT2 (SLC6A5) define a presynaptic component of human startle disease

Hyperekplexia is a human neurological disorder characterized by an excessive startle response and is typically caused by missense and nonsense mutations in the gene encoding the inhibitory glycine receptor (GlyR) alpha1 subunit (GLRA1). Genetic heterogeneity has been confirmed in rare sporadic cases, with mutations affecting other postsynaptic glycinergic proteins including the GlyR beta subunit (GLRB), gephyrin (GPHN) and RhoGEF collybistin (ARHGEF9). However, many individuals diagnosed with sporadic hyperekplexia do not carry mutations in these genes. Here we show that missense, nonsense and frameshift mutations in SLC6A5 (ref. 8), encoding the presynaptic glycine transporter 2 (GlyT2), also cause hyperekplexia. Individuals with mutations in SLC6A5 present with hypertonia, an exaggerated startle response to tactile or acoustic stimuli, and life-threatening neonatal apnea episodes. SLC6A5 mutations result in defective subcellular GlyT2 localization, decreased glycine uptake or both, with selected mutations affecting predicted glycine and Na+ binding sites.     

Senataxin, the ortholog of a yeast RNA helicase, is mutant in ataxia-ocular apraxia 2

Ataxia-ocular apraxia 2 (AOA2) was recently identified as a new autosomal recessive ataxia. We have now identified causative mutations in 15 families, which allows us to clinically define this entity by onset between 10 and 22 years, cerebellar atrophy, axonal sensorimotor neuropathy, oculomotor apraxia and elevated alpha-fetoprotein (AFP). Ten of the fifteen mutations cause premature termination of a large DEAxQ-box helicase, the human ortholog of yeast Sen1p, involved in RNA maturation and termination.     

Mutations in a new gene encoding a thiamine transporter cause thiamine-responsive megaloblastic anaemia syndrome.

Thiamine-responsive megaloblastic anaemia syndrome (TRMA; MIM 249270) is an autosomal recessive disorder with features that include megaloblastic anaemia, mild thrombocytopenia and leucopenia, sensorineural deafness and diabetes mellitus. Treatment with pharmacologic doses of thiamine ameliorates the megaloblastic anaemia and diabetes mellitus. A defect in the plasma membrane transport of thiamine has been demonstrated in erythrocytes and cultured skin fibroblasts from TRMA patients. The gene causing TRMA was assigned to 1q23.2-q23.3 by linkage analysis. Here we report the cloning of a new gene, SLC19A2, identified from high-through-put genomic sequences due to homology with SLC19A1, encoding reduced folate carrier 1 (refs 8-10). We cloned the entire coding region by screening a human fetal brain cDNA library. SLC19A2 encodes a protein (of 497 aa) predicted to have 12 transmembrane domains. We identified 2 frameshift mutations in exon 2. a 1-bp insertion and a 2-bp deletion, among four Iranian families with TRMA. The sequence homology and predicted structure of SLC19A2, as well as its role in TRMA, suggest that its gene product is a thiamine carrier, the first to be identified in complex eukaryotes.     

The gene encoding ganglioside-induced differentiation-associated protein 1 is mutated in axonal Charcot-Marie-Tooth type 4A disease.

We identified three distinct mutations and six mutant alleles in GDAP1 in three families with axonal Charcot-Marie-Tooth (CMT) neuropathy and vocal cord paresis, which were previously linked to the CMT4A locus on chromosome 8q21.1. These results establish the molecular etiology of CMT4A (MIM 214400) and suggest that it may be associated with both axonal and demyelinating phenotypes.     

Non-polarized targeting of AE1 causes autosomal dominant distal renal tubular acidosis

Autosomal dominant distal renal tubular acidosis (ddRTA) is caused by mutations in SLC4A1, which encodes the polytopic chloride-bicarbonate exchanger AE1 that is normally expressed at the basolateral surface of alpha-intercalated cells in the distal nephron. Here we report that, in contrast with many disorders in which mutant membrane proteins are retained intracellularly and degraded, ddRTA can result from aberrant targeting of AE1 to the apical surface.     

A new gene, encoding an anion transporter, is mutated in sialic acid storage diseases

Sialic acid storage diseases (SASD, MIM 269920) are autosomal recessive neurodegenerative disorders that may present as a severe infantile form (ISSD) or a slowly progressive adult form, which is prevalent in Finland (Salla disease). The main symptoms are hypotonia, cerebellar ataxia and mental retardation; visceromegaly and coarse features are also present in infantile cases. Progressive cerebellar atrophy and dysmyelination have been documented by magnetic resonance imaging (ref. 4). Enlarged lysosomes are seen on electron microscopic studies and patients excrete large amounts of free sialic acid in urine. A H+/anionic sugar symporter mechanism for sialic acid and glucuronic acid is impaired in lysosomal membranes from Salla and ISSD patients. The locus for Salla disease was assigned to a region of approximately 200 kb on chromosome 6q14-q15 in a linkage study using Finnish families. Salla disease and ISSD were further shown to be allelic disorders. A physical map with P1 and PAC clones was constructed to cover the 200-kb area flanked by the loci D6S280 and D6S1622, providing the basis for precise physical positioning of the gene. Here we describe a new gene, SLC17A5 (also known as AST), encoding a protein (sialin) with a predicted transport function that belongs to a family of anion/cation symporters (ACS). We found a homozygous SLC17A5 mutation (R39C) in five Finnish patients with Salla disease and six different SLC17A5 mutations in six ISSD patients of different ethnic origins. Our observations suggest that mutations in SLC17A5 are the primary cause of lysosomal sialic acid storage diseases.     

Calcium oxalate urolithiasis in mice lacking anion transporter Slc26a6

Urolithiasis is one of the most common urologic diseases in industrialized societies. Calcium oxalate is the predominant component in 70-80% of kidney stones, and small changes in urinary oxalate concentration affect the risk of stone formation. SLC26A6 is an anion exchanger expressed on the apical membrane in many epithelial tissues, including kidney and intestine. Among its transport activities, SLC26A6 mediates Cl(-)-oxalate exchange. Here we show that mutant mice lacking Slc26a6 develop a high incidence of calcium oxalate urolithiasis. Slc26a6-null mice have significant hyperoxaluria and elevation in plasma oxalate concentration that is greatly attenuated by dietary oxalate restriction. In vitro flux studies indicated that mice lacking Slc26a6 have a defect in intestinal oxalate secretion resulting in enhanced net absorption of oxalate. We conclude that the anion exchanger SLC26A6 has a major constitutive role in limiting net intestinal absorption of oxalate, thereby preventing hyperoxaluria and calcium oxalate urolithiasis.     

Hot-spot residue in small heat-shock protein 22 causes distal motor neuropathy

Distal hereditary motor neuropathies are pure motor disorders of the peripheral nervous system resulting in severe atrophy and wasting of distal limb muscles. In two pedigrees with distal hereditary motor neuropathy type II linked to chromosome 12q24.3, we identified the same mutation (K141N) in small heat-shock 22-kDa protein 8 (encoded by HSPB8; also called HSP22). We found a second mutation (K141E) in two smaller families. Both mutations target the same amino acid, which is essential to the structural and functional integrity of the small heat-shock protein alphaA-crystallin. This positively charged residue, when mutated in other small heat-shock proteins, results in various human disorders. Coimmunoprecipitation experiments showed greater binding of both HSPB8 mutants to the interacting partner HSPB1. Expression of mutant HSPB8 in cultured cells promoted formation of intracellular aggregates. Our findings provide further evidence that mutations in heat-shock proteins have an important role in neurodegenerative disorders.     

Disrupted function and axonal distribution of mutant tyrosyl-tRNA synthetase in dominant intermediate Charcot-Marie-Tooth neuropathy

Charcot-Marie-Tooth (CMT) neuropathies are common disorders of the peripheral nervous system caused by demyelination or axonal degeneration, or a combination of both features. We previously assigned the locus for autosomal dominant intermediate CMT neuropathy type C (DI-CMTC) to chromosome 1p34-p35. Here we identify two heterozygous missense mutations (G41R and E196K) and one de novo deletion (153-156delVKQV) in tyrosyl-tRNA synthetase (YARS) in three unrelated families affected with DI-CMTC. Biochemical experiments and genetic complementation in yeast show partial loss of aminoacylation activity of the mutant proteins, and mutations in YARS, or in its yeast ortholog TYS1, reduce yeast growth. YARS localizes to axonal termini in differentiating primary motor neuron and neuroblastoma cultures. This specific distribution is significantly reduced in cells expressing mutant YARS proteins. YARS is the second aminoacyl-tRNA synthetase found to be involved in CMT, thereby linking protein-synthesizing complexes with neurodegeneration.     

Mutations in SLC6A19, encoding B0AT1, cause Hartnup disorder

Hartnup disorder, an autosomal recessive defect named after an English family described in 1956 (ref. 1), results from impaired transport of neutral amino acids across epithelial cells in renal proximal tubules and intestinal mucosa. Symptoms include transient manifestations of pellagra (rashes), cerebellar ataxia and psychosis. Using homozygosity mapping in the original family in whom Hartnup disorder was discovered, we confirmed that the critical region for one causative gene was located on chromosome 5p15 (ref. 3). This region is homologous to the area of mouse chromosome 13 that encodes the sodium-dependent amino acid transporter B(0)AT1 (ref. 4). We isolated the human homolog of B(0)AT1, called SLC6A19, and determined its size and molecular organization. We then identified mutations in SLC6A19 in members of the original family in whom Hartnup disorder was discovered and of three Japanese families. The protein product of SLC6A19, the Hartnup transporter, is expressed primarily in intestine and renal proximal tubule and functions as a neutral amino acid transporter.     

Rare independent mutations in renal salt handling genes contribute to blood pressure variation

The effects of alleles in many genes are believed to contribute to common complex diseases such as hypertension. Whether risk alleles comprise a small number of common variants or many rare independent mutations at trait loci is largely unknown. We screened members of the Framingham Heart Study (FHS) for variation in three genes-SLC12A3 (NCCT), SLC12A1 (NKCC2) and KCNJ1 (ROMK)-causing rare recessive diseases featuring large reductions in blood pressure. Using comparative genomics, genetics and biochemistry, we identified subjects with mutations proven or inferred to be functional. These mutations, all heterozygous and rare, produce clinically significant blood pressure reduction and protect from development of hypertension. Our findings implicate many rare alleles that alter renal salt handling in blood pressure variation in the general population, and identify alleles with health benefit that are nonetheless under purifying selection. These findings have implications for the genetic architecture of hypertension and other common complex traits.     

The gene encoding gigaxonin, a new member of the cytoskeletal BTB/kelch repeat family, is mutated in giant axonal neuropathy

Disorganization of the neurofilament network is a prominent feature of several neurodegenerative disorders including amyotrophic lateral sclerosis (ALS), infantile spinal muscular atrophy and axonal Charcot-Marie-Tooth disease. Giant axonal neuropathy (GAN, MIM 256850), a severe, autosomal recessive sensorimotor neuropathy affecting both the peripheral nerves and the central nervous system, is characterized by neurofilament accumulation, leading to segmental distension of the axons. GAN corresponds to a generalized disorganization of the cytoskeletal intermediate filaments (IFs), to which neurofilaments belong, as abnormal aggregation of multiple tissue-specific IFs has been reported: vimentin in endothelial cells, Schwann cells and cultured skin fibroblasts, and glial fibrillary acidic protein (GFAP) in astrocytes. Keratin IFs also seem to be alterated, as most patients present characteristic curly or kinky hairs. We report here identification of the gene GAN, which encodes a novel, ubiquitously expressed protein we have named gigaxonin. We found one frameshift, four nonsense and nine missense mutations in GAN of GAN patients. Gigaxonin is composed of an amino-terminal BTB (for Broad-Complex, Tramtrack and Bric a brac) domain followed by a six kelch repeats, which are predicted to adopt a beta-propeller shape. Distantly related proteins sharing a similar domain organization have various functions associated with the cytoskeleton, predicting that gigaxonin is a novel and distinct cytoskeletal protein that may represent a general pathological target for other neurodegenerative disorders with alterations in the neurofilament network.