Genetic and phenotypic characterization of NKX6-2-related spastic ataxia and hypomyelination.

BACKGROUND AND PURPOSE: Hypomyelinating leukodystrophies are a heterogeneous group of genetic disorders with a wide spectrum of phenotypes and a high rate of genetically unsolved cases. Bi-allelic mutations in NKX6-2 were recently linked to spastic ataxia 8 with hypomyelinating leukodystrophy. METHODS: Using a combination of homozygosity mapping, exome sequencing, and detailed clinical and neuroimaging assessment a series of new NKX6-2 mutations in a multicentre setting is described. Then, all reported NKX6-2 mutations and those identified in this study were combined and an in-depth analysis of NKX6-2-related disease spectrum was provided. RESULTS: Eleven new cases from eight families of different ethnic backgrounds carrying compound heterozygous and homozygous pathogenic variants in NKX6-2 were identified, evidencing a high NKX6-2 mutation burden in the hypomyelinating leukodystrophy disease spectrum. Our data reveal a phenotype spectrum with neonatal onset, global psychomotor delay and worse prognosis at the severe end and a childhood onset with mainly motor phenotype at the milder end. The phenotypic and neuroimaging expression in NKX6-2 is described and it is shown that phenotypes with epilepsy in the absence of overt hypomyelination and diffuse hypomyelination without seizures can occur. CONCLUSIONS: NKX6-2 mutations should be considered in patients with autosomal recessive, very early onset of nystagmus, cerebellar ataxia with hypotonia that rapidly progresses to spasticity, particularly when associated with neuroimaging signs of hypomyelination. Therefore, it is recommended that NXK6-2 should be included in hypomyelinating leukodystrophy and spastic ataxia diagnostic panels.

Mitochondrial complex I NUBPL mutations cause combined dystonia with bilateral striatal necrosis and cerebellar atrophy.

BACKGROUND AND PURPOSE: The recent advances in genetics have helped to unravel the cause of many dystonia syndromes. With the broadening spectrum of genetically defined dystonia syndromes, distinct clinico-radiological phenotypes are a welcome handle to guide the diagnostic work-up. METHODS: Exome sequencing was used to elucidate the genetic cause of a syndrome characterized by generalized dystonia, pyramidal and cerebellar involvement, with bilateral striatal necrosis (BSN) and cerebellar atrophy on magnetic resonance imaging. Homozygosity mapping and linkage analysis were used in a supportive role. Known genetic causes of BSN were excluded by use of exome data or Sanger sequencing. RESULTS: Compound heterozygous mutations were identified in the NUBPL gene in a small UK kindred. The gene lay in a region of positive linkage and segregated with disease in a family of six individuals. CONCLUSION: NUBPL mutations cause early onset, autosomal recessive generalized dystonia with cerebellar ataxia, pyramidal signs, preserved cognition and a distinct magnetic resonance imaging appearance with BSN and cerebellar atrophy.

Genetic overlap between Alzheimer's disease and Parkinson's disease at the MAPT locus.

We investigated the genetic overlap between Alzheimer's disease (AD) and Parkinson's disease (PD). Using summary statistics (P-values) from large recent genome-wide association studies (GWAS) (total n=89 904 individuals), we sought to identify single nucleotide polymorphisms (SNPs) associating with both AD and PD. We found and replicated association of both AD and PD with the A allele of rs393152 within the extended MAPT region on chromosome 17 (meta analysis P-value across five independent AD cohorts=1.65 × 10(-7)). In independent datasets, we found a dose-dependent effect of the A allele of rs393152 on intra-cerebral MAPT transcript levels and volume loss within the entorhinal cortex and hippocampus. Our findings identify the tau-associated MAPT locus as a site of genetic overlap between AD and PD, and extending prior work, we show that the MAPT region increases risk of Alzheimer's neurodegeneration.

Olfaction in Parkin single and compound heterozygotes in a cohort of young onset Parkinson's disease patients.

BACKGROUND: Parkin related Parkinson's disease (PD) is differentiated from idiopathic PD by absent or sparse Lewy bodies, and preserved olfaction. The significance of single Parkin mutations in the pathogenesis of PD is debated. OBJECTIVES: To assess olfaction results according to Parkin mutation status. To compare the prevalence of Parkin single heterozygous mutations in patients diagnosed with PD to the rate in healthy controls in order to establish whether these single mutations could be a risk factor for developing PD. METHODS: Parkin gene mutation testing was performed in young onset PD (diagnosed <50 years old) to identify three groups: Parkin homozygous or compound heterozygote mutation carriers, Parkin single heterozygote mutation carriers, and non-carriers of Parkin mutations. Olfaction was tested using the 40-item British version of the University of Pennsylvania smell identification test (UPSIT). RESULTS: Of 344 young onset PD cases tested, 8 (2.3%) were Parkin compound heterozygotes and 13 (3.8%) were Parkin single heterozygotes. Olfaction results were available in 282 cases (eight compound heterozygotes, nine single heterozygotes, and 265 non-carriers). In Parkin compound heterozygotes, the median UPSIT score was 33, interquartile range (IQR) 28.5-36.5, which was significantly better than in single Parkin heterozygotes (median 19, IQR 18-28) and non-carriers (median score 22, IQR 16-28) (ANOVA P < 0.001). These differences persisted after adjusting for age, disease duration, gender, and smoking (P < 0.001). There was no significant difference in UPSIT scores between single heterozygotes and non-carriers (P = 0.90). CONCLUSIONS: Patients with Parkin compound heterozygous mutations have relatively preserved olfaction compared to Parkin single heterozygotes and non-carriers. The prevalence of Parkin single heterozygosity is similar to the 3.7% rate reported in healthy controls.

A conserved sorting-associated protein is mutant in chorea-acanthocytosis.

Chorea-acanthocytosis (CHAC, MIM 200150) is an autosomal recessive neurodegenerative disorder characterized by the gradual onset of hyperkinetic movements and abnormal erythrocyte morphology (acanthocytosis). Neurological findings closely resemble those observed in Huntington disease. We identified a gene in the CHAC critical region and found 16 different mutations in individuals with chorea-acanthocytosis. CHAC encodes an evolutionarily conserved protein that is probably involved in protein sorting.

LRRK2 expression in idiopathic and G2019S positive Parkinson's disease subjects: a morphological and quantitative study.

AIMS: Mutations in the gene encoding leucine-rich repeat kinase-2 (LRRK2) have been established as a common genetic cause of Parkinson's disease (PD). The distribution of LRRK2 mRNA and protein in the human brain has previously been described, although it has not been reported in PD cases with the common LRRK2 G2019S mutation. METHODS: To further elucidate the role of LRRK2 in PD, we determined the localization of LRRK2 mRNA and protein in post-mortem brain tissue from control, idiopathic PD (IPD) and G2019S positive PD cases. RESULTS: Widespread neuronal expression of LRRK2 mRNA and protein was recorded and no difference was observed in the morphological localization of LRRK2 mRNA or protein between control, IPD and G2019S positive PD cases. Using quantitative real-time polymerase chain reaction, we demonstrated that there is no regional variation in LRRK2 mRNA in normal human brain, but we have identified differential expression of LRRK2 mRNA with significant reductions recorded in limbic and neocortical regions of IPD cases compared with controls. Semi-quantitative analysis of LRRK2 immunohistochemical staining demonstrated regional variation in staining intensity, with weak LRRK2 immunoreactivity consistently recorded in the striatum and substantia nigra. No clear differences were identified in LRRK2 immunoreactivity between control, IPD and G2019S positive PD cases. LRRK2 protein was identified in a small proportion of Lewy bodies. CONCLUSIONS: Our data suggest that widespread dysregulation of LRRK2 mRNA expression may contribute to the pathogenesis of IPD.

Autosomal-dominant GTPCH1-deficient DRD: clinical characteristics and long-term outcome of 34 patients.

BACKGROUND: An autosomal dominantly inherited defect in the GCH1 gene that encodes guanosine triphosphate cyclohydrolase 1 (GTPCH1) is the most common cause of dopa-responsive dystonia (DRD). A classic phenotype of young-onset lower-limb dystonia, diurnal fluctuations and excellent response to levodopa has been well recognised in association with GCH1 mutations, and rare atypical presentations have been reported. However, a number of clinical issues remain unresolved including phenotypic variability, long-term response to levodopa and associated non-motor symptoms, and there are limited data on long-term follow-up of genetically proven cases. METHODS: A detailed clinical evaluation of 34 patients (19 women, 15 men), with confirmed mutations in the GCH1 gene, is presented. RESULTS AND CONCLUSIONS: The classic phenotype was most frequent (n = 23), with female predominance (F:M = 16:7), and early onset (mean 4.5 years) with involvement of legs. However, a surprisingly large number of patients developed craniocervical dystonia, with spasmodic dysphonia being the predominant symptom in two subjects. A subset of patients, mainly men, presented with either a young-onset (mean 6.8 years) mild DRD variant not requiring treatment (n = 4), or with an adult-onset (mean 37 years) Parkinson disease-like phenotype (n = 4). Two siblings were severely affected with early hypotonia and delay in motor development, associated with compound heterozygous GCH1 gene mutations. The study also describes a number of supplementary features including restless-legs-like symptoms, influence of female sex hormones, predominance of tremor or parkinsonism in adult-onset cases, initial reverse reaction to levodopa, recurrent episodes of depressive disorder and specific levodopa-resistant symptoms (writer's cramp, dysphonia, truncal dystonia). Levodopa was used effectively and safely in 20 pregnancies, and did not cause any fetal abnormalities.

Expression of BRI2 mRNA and protein in normal human brain and familial British dementia: its relevance to the pathogenesis of disease.

INTRODUCTION: Two different disease-specific mutations in the BRI2 gene, situated on chromosome 13, have been identified as giving rise to familial British dementia (FBD) and familial Danish dementia (FDD). Each mutation results in extension of the open reading frame generating the disease-specific precursor proteins which are cleaved by furin-like proteolysis releasing the amyloidogenic C-terminal peptides ABri and ADan in FBD and FDD, respectively. MATERIAL AND METHODS: To understand the mechanism of the formation of amyloid lesions in FBD, we studied the origin of the precursor proteins and furin in the human brain. We used control brains, cases of sporadic Alzheimer's disease (AD), variant AD with cotton wool plaques and FBD to study BRI2 mRNA expression using in situ hybridization. Furin and BRI2 protein expression was investigated using Western blotting and immunohistochemistry. RESULTS: BRI2 mRNA and BRI2 protein are widely expressed primarily by neurones and glia and are deposited in the amyloid lesions in FBD. They were, however, not expressed by cerebrovascular components. Furin expression showed a similar pattern except that it was also present in cerebrovascular smooth muscle cells. CONCLUSIONS: These findings suggest that neurones and glia and are a major source of BRI2 protein and that in FBD, the mutated precursor protein may undergo furin cleavage within neurones to produce the amyloid peptide ABri. The failure to demonstrate BRI2 in blood vessels under the conditions tested suggests that vascular amyloid peptide production does not contribute significantly to cerebral amyloid angiopathy (CAA) in FBD and FDD, lending indirect support to the drainage hypothesis of CAA.

The genetics of Parkinson's disease.

The effort to map the entire human genome has led recently to the important milestone publication in late 1999 of the complete sequence of chromosome 22. This has been facilitated by increasingly sophisticated tools for genetic analysis and the ensuing wealth of detailed genetic information. The quest for genetic factors contributing to Parkinson's disease and parkinsonian disorders has revealed a progressively complex picture implicating gene mutations in the rarer, autosomally inherited forms of Parkinson's disease and the interplay of genetic and/or environmental factors in the common sporadic forms of the disorder. These findings not only reiterate the complex genetic heterogeneity of Parkinson's disease but could also point towards common pathogenic mechanisms in Parkinson's disease and related neurodegenerative disorders.

Multiple mitochondrial DNA deletions in monozygotic twins with OPMD.

BACKGROUND: Oculopharyngeal muscular dystrophy (OPMD) is caused by expansions of the poly (A) binding protein 2 (PABP2) gene. Previous histological analyses have revealed mitochondrial abnormalities in the muscles of OPMD patients but their significance remains uncertain. OBJECTIVE: We had the rare opportunity to study monozygotic twins with identical expansions of the PABP2 gene but with markedly different severities of OPMD. Both had histological features of mitochondrial myopathy. We determined whether mitochondrial DNA abnormalities underlay these changes. METHODS: Clinical information was obtained by history and examination. Muscle biopsies were obtained from each subject and genetic analysis was performed using long-range PCR and Southern blotting. RESULTS: We demonstrate, for the first time, the presence of mitochondrial DNA (mtDNA) deletions by Southern blotting in individuals with OPMD. This correlates with the presence of mitochondrial myopathy in both twins. Moreover, both twins had different mtDNA deletions, which might explain their phenotypic differences. CONCLUSION: We hypothesise that mitochondrial dysfunction may occur as a consequence of PABP2 gene mutations, and that this dysfunction may affect the phenotypic manifestations of OPMD.

Clinical heterogeneity and genotype-phenotype correlations in hereditary spastic paraplegia because of Spatacsin mutations (SPG11).

BACKGROUND: Autosomal recessive hereditary spastic paraplegia (ARHSP) with thin corpus callosum is a distinct and usually severe form of complex hereditary spastic paraplegia classified as SPG11. Recently mutations on SPG11 gene (KIAA1840), which is localized to chromosome 15q13-q15, were shown to cause the majority of SPG11 cases. METHODS: We analysed the 40 coding exons of this gene in the probands from eight families with complex ARHSP, four of these families had a thin corpus callosum and two has mild thinning. RESULTS: Three families were identified with novel mutations in the SPG11 gene. One family was of Asian origin with a homozygous nonsense mutation and had a very severe phenotype but only very mild thinning of the corpus callosum. In the other two English families the parents were unrelated and the mutations were compound heterozygotes. In these two families the phenotype was mild and both probands had a thin corpus callosum. CONCLUSION: Given the probable mechanism of action of the mutations in the Spatacsin gene, we discuss the probable genotype phenotype correlations in these families. This study confirms the frequent occurrence of Spatacsin mutations in complex ARHSP with genotype phenotype effects and exposes the spectrum of clinical heterogeneity in SPG11.

Test for LRRK2 mutations in patients with Parkinson's disease.

LRRK2-associated Parkinson's disease is common enough to raise clinical questions such as which patients should be tested and what advice should be given. We discuss practical issues in the light of our experiences with real life Parkinson's disease patients. Neurologists should consider testing LRRK2 in Parkinson's disease patients with affected first degree relatives where the onset is over the age of 40 years. A common G2019S mutation makes genetic testing straightforward and cost-effective. Age-related or reduced genetic penetrance means that LRRK2 mutations are also found in sporadic Parkinson's disease patients; however, at present, there is little to support the widespread testing of these patients except in high-risk ethnic groups such as North African Arabs and Ashkenazi Jews. Incomplete penetrance also complicates presymptomatic testing, which is best undertaken in the context of appropriate genetic counselling.

Understanding the molecular causes of Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disease that is both common and incurable. The majority of cases are sporadic and of unknown origin but several genes have been identified that, when mutated, give rise to rare, familial forms of the disease. The principal genes that have been shown to cause PD are alpha-synuclein (SNCA), parkin, leucine-rich repeat kinase 2 (LRRK2), PTEN-induced putative kinase 1 (PINK1) and DJ-1. Here, we discuss what has been learnt from the study of these genes and what has been elucidated of the molecular pathways that lead to cell degeneration. Of importance is what these molecular events and pathways tell scientists of the common sporadic form of PD. Although complete knowledge of these genes' functions remains elusive, recent work implicates abnormal protein accumulation, protein phosphorylation, mitochondrial dysfunction and oxidative stress as common pathways to PD pathogenesis.

PINK1 protein in normal human brain and Parkinson's disease.

Parkinson's disease is a common incurable neurodegenerative disease whose molecular aetiology remains unclear. The identification of Mendelian genes causing rare familial forms of Parkinson's disease has revealed novel proteins and pathways that are likely to be relevant in the pathogenesis of sporadic Parkinson's disease. Recently, mutations in a novel gene, PINK1, encoding a 581 amino acid protein with both mitochondrial targeting and serine/threonine kinase domains, were identified as a cause of autosomal recessive parkinsonism. This provided important evidence for the role of the mitochondrial dysfunction and kinase pathways in neurodegeneration. In this study, we report the first characterization of the PINK1 protein in normal human and sporadic Parkinson's brains, in addition to Parkinson's cases with heterozygous PINK1 mutations. The possible role of the PINK1 protein was also assessed in a number of neurodegenerative diseases characterized by proteinaceous inclusions. For these studies, rabbit polyclonal antibodies were raised against two peptide sequences within the N-terminal hydrophilic loops of PINK1 protein. Using immunohistochemistry and western blotting we were able to demonstrate that PINK1 is a ubiquitous protein expressed throughout the human brain and it is found in all cell types showing a punctate cytoplasmic staining pattern consistent with mitochondrial localization. Fractionation studies of human and rat brain confirm that PINK1 is localized to the mitochondrial membranes. In addition, we show that PINK1 is detected in a proportion of Lewy bodies in cases of sporadic Parkinson's disease and Parkinson's disease associated with heterozygous mutations in the PINK1 gene, which are clinically and pathologically indistinguishable from the sporadic cases. PINK1 was absent in cortical Lewy bodies, in neurofibrillary tangles in Alzheimer's disease, progressive supranuclear palsy and corticobasal degeneration, and in the glial and neuronal alpha-synuclein positive inclusions in multiple system atrophy. These studies provide for the first time in vivo morphological and biochemical evidence to support a mitochondrial localization of PINK1 and underpin the significance of mitochondrial dysfunction in the pathogenesis of nigral cell degeneration in Parkinson's disease.

The sepiapterin reductase gene region reveals association in the PARK3 locus: analysis of familial and sporadic Parkinson's disease in European populations.

BACKGROUND: Parkinson's disease is a genetically complex disease with mixed mode of inheritance. Recently, a haplotype across the sepiapterin reductase (SPR) gene, which is located in the PARK3 linkage region, was shown to modulate age of onset of Parkinson's disease in sibships from North America. OBJECTIVE: To make a thorough assessment of the SPR gene region in sporadic Parkinson's disease. METHODS: A linkage study in 122 European sibship families with five microsatellite and 17 single nucleotide polymorphism (SNP) markers in and around the SPR gene region, and an association analysis in 340 sporadic cases of Parkinson's disease and 680 control subjects from Germany with 40 SNPs. Linkage was evaluated by non-parametric linkage scores and genotypic or haplotype association was tested by regression analysis, assuming different risk effect models. RESULTS: Significant LOD scores between 2 and 3 were obtained at the two SPR-flanking markers D2S2110 and D2S1394 and seven SNP markers around the SPR gene. We found the previously reported promoter SNP rs1876487 also significantly associated with age of onset in our sib pair families (p-value 0.02). One strong linkage disequilibrium (LD) block of 45 kb including the entire SPR gene was observed. Within this LD block all 14 inter-correlated SNPs were significantly associated with Parkinson's disease affection status (p-value 0.004). CONCLUSIONS: DNA polymorphisms in a highly intercorrelated LD block, which includes the SPR gene, appear to be associated with both sporadic and familial Parkinson's disease. This confirms a previous study showing that SPR potentially modulates the onset of or risk for Parkinson's disease.

Genetic association studies of complex neurological diseases.

Genetic association studies offer a powerful approach to identify the multiple variants of small effect that modulate susceptibility to common, complex disease. They, however, have a poor reputation, mainly because of the consistent lack of replication of all but a few. Thousands of genetic studies have been carried out on multifactorial diseases in the past 30 years, yielding only about 50 variants that can be considered to be true positives. Although the positive studies show proof of principle, the multitude of negative studies indicate fundamental problems in the design and execution of association studies. Here, we discuss some of the more pertinent study design and data analysis issues which can affect the outcome of genetic association studies.

The fragile X tremor ataxia syndrome in the differential diagnosis of multiple system atrophy: data from the EMSA Study Group.

The recent identification of fragile X-associated tremor ataxia syndrome (FXTAS) associated with premutations in the FMR1 gene and the possibility of clinical overlap with multiple system atrophy (MSA) has raised important questions, such as whether genetic testing for FXTAS should be performed routinely in MSA and whether positive cases might affect the specificity of current MSA diagnostic criteria. We genotyped 507 patients with clinically diagnosed or pathologically proven MSA for FMR1 repeat length. Among the 426 clinically diagnosed cases, we identified four patients carrying FMR1 premutations (0.94%). Within the subgroup of patients with probable MSA-C, three of 76 patients (3.95%) carried premutations. We identified no premutation carriers among 81 patients with pathologically proven MSA and only one carrier among 622 controls (0.16%). Our results suggest that, with proper application of current diagnostic criteria, FXTAS is very unlikely to be confused with MSA. However, slowly progressive disease or predominant tremor are useful red flags and should prompt the consideration of FXTAS. On the basis of our data, the EMSA Study Group does not recommend routine FMR1 genotyping in typical MSA patients.

Linkage disequilibrium fine mapping and haplotype association analysis of the tau gene in progressive supranuclear palsy and corticobasal degeneration.

BACKGROUND: The haplotype H1 of the tau gene, MAPT, is highly associated with progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD). OBJECTIVE: To investigate the pathogenic basis of this association. METHODS: Detailed linkage disequilibrium and common haplotype structure of MAPT were examined in 27 CEPH trios using validated HapMap genotype data for 24 single nucleotide polymorphisms (SNPs) spanning MAPT. RESULTS: Multiple variants of the H1 haplotype were resolved, reflecting a far greater diversity of MAPT than can be explained by the H1 and H2 clades alone. Based on this, six haplotype tagging SNPs (htSNPs) that capture 95% of the common haplotype diversity were used to genotype well characterised PSP and CBD case-control cohorts. In addition to strong association with PSP and CBD of individual SNPs, two common haplotypes derived from these htSNPs were identified that are highly associated with PSP: the sole H2 derived haplotype was underrepresented and one of the common H1 derived haplotypes was highly associated, with a similar trend observed in CBD. There were powerful and highly significant associations with PSP and CBD of haplotypes formed by three H1 specific SNPs. This made it possible to define a candidate region of at least approximately 56 kb, spanning sequences from upstream of MAPT exon 1 to intron 9. On the H1 haplotype background, these could harbour the pathogenic variants. CONCLUSIONS: The findings support the pathological evidence that underlying variations in MAPT could contribute to disease pathogenesis by subtle effects on gene expression and/or splicing. They also form the basis for the investigation of the possible genetic role of MAPT in Parkinson's disease and other tauopathies, including Alzheimer's disease.