Cinzia Tiloca

Mutations in the Matrin 3 gene cause familial amyotrophic lateral sclerosis

MATR3 is an RNA- and DNA-binding protein that interacts with TDP-43, a disease protein linked to amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. Using exome sequencing, we identified mutations in MATR3 in ALS kindreds. We also observed MATR3 pathology in ALS-affected spinal cords with and without MATR3 mutations. Our data provide more evidence supporting the role of aberrant RNA processing in motor neuron degeneration.

Genetics of familial Amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a late onset, rapidly progressive and ultimately fatal neurodegenerative disease, caused by the loss of motor neurons in the brain and spinal cord. About 10% of all ALS cases are familial (FALS), and constitute a clinically and genetically heterogeneous entity. To date, FALS has been linked to mutations in 10 different genes and to four additional chromosomal loci. Research on FALS genetics, and in particular the discoveries of mutations in the SOD1, TARDBP, and FUS genes, has provided essential information toward the understanding of the pathogenesis of ALS in general. This review presents a tentative classification of all FALS-associated genes identified so far.

Novel optineurin mutations in patients with familial and sporadic amyotrophic lateral sclerosis

Background Optineurin (OPTN), a causative gene of hereditary primary open-angle glaucoma, has been recently associated with amyotrophic lateral sclerosis (ALS) with mainly autosomal recessive, but also dominant, traits. To further define the contribution of OPTN gene in ALS, we performed a mutational screening in a large cohort of Italian patients. Methods A group of 274 ALS patients, including 161 familial (FALS) and 113 sporadic (SALS) cases, were screened for OPTN mutations by direct sequencing of its coding sequence. All patients fulfilled the El Escorial criteria for probable or definite ALS and were negative for mutations in SOD1, ANG, TARDBP and FUS/TLS genes. Results The genetic analysis revealed six novel variants in both FALS and SALS patients, all occurring in an heterozygous state. We identified three missense (c.844A→C p.T282P, c.941A→T p.Q314L, c.1670A→C p.K557T), one nonsense (c.67G→T p.G23X) and two intronic mutations (c.552+1delG, c.1401+4A→G). The intronic c.552+1delG variant determined a splicing defect as demonstrated by mRNA analysis. All mutations were absent in 280 Italian controls and over 6800 worldwide glaucoma patients and controls screened so far. The clinical phenotype of OPTN-mutated patients was heterogeneous for both age of onset and disease duration but characterised by lower-limb onset and prevalence of upper motor neuron signs. Conclusion In this cohort, OPTN mutations were present both in FALS (2/161), accounting for 1.2% cases, and in SALS patients (4/113), thereby extending the spectrum of OPTN mutations associated with ALS. The study further supports the possible pathological role of optineurin protein in motor neuron disease.

C9ORF72 repeat expansion in a large Italian ALS cohort: evidence of a founder effect

A hexanucleotide repeat expansion (RE) in C9ORF72 gene was recently reported as the main cause of amyotrophic lateral sclerosis (ALS) and cases with frontotemporal dementia. We screened C9ORF72 in a large cohort of 259 familial ALS, 1275 sporadic ALS, and 862 control individuals of Italian descent. We found RE in 23.9% familial ALS, 5.1% sporadic ALS, and 0.2% controls. Two cases carried the RE together with mutations in other ALS-associated genes. The phenotype of RE carriers was characterized by bulbar-onset, shorter survival, and association with cognitive and behavioral impairment. Extrapyramidal and cerebellar signs were also observed in few patients. Genotype data revealed that 95% of RE carriers shared a restricted 10-single nucleotide polymorphism haplotype within the previously reported 20-single nucleotide polymorphism risk haplotype, detectable in only 27% of nonexpanded ALS cases and in 28% of controls, suggesting a common founder with cohorts of North European ancestry. Although C9ORF72 RE segregates with disease, the identification of RE both in controls and in patients carrying additional pathogenic mutations suggests that penetrance and phenotypic expression of C9ORF72 RE may depend on additional genetic risk factors.

A genome-wide association meta-analysis identifies a novel locus at 17q11.2 associated with sporadic amyotrophic lateral sclerosis

Identification of mutations at familial loci for amyotrophic lateral sclerosis (ALS) has provided novel insights into the aetiology of this rapidly progressing fatal neurodegenerative disease. However, genome-wide association studies (GWAS) of the more common (∼90%) sporadic form have been less successful with the exception of the replicated locus at 9p21.2. To identify new loci associated with disease susceptibility, we have established the largest association study in ALS to date and undertaken a GWAS meta-analytical study combining 3959 newly genotyped Italian individuals (1982 cases and 1977 controls) collected by SLAGEN (Italian Consortium for the Genetics of ALS) together with samples from Netherlands, USA, UK, Sweden, Belgium, France, Ireland and Italy collected by ALSGEN (the International Consortium on Amyotrophic Lateral Sclerosis Genetics). We analysed a total of 13 225 individuals, 6100 cases and 7125 controls for almost 7 million single-nucleotide polymorphisms (SNPs). We identified a novel locus with genome-wide significance at 17q11.2 (rs34517613 with P = 1.11 × 10(-8); OR 0.82) that was validated when combined with genotype data from a replication cohort (P = 8.62 × 10(-9); OR 0.833) of 4656 individuals. Furthermore, we confirmed the previously reported association at 9p21.2 (rs3849943 with P = 7.69 × 10(-9); OR 1.16). Finally, we estimated the contribution of common variation to heritability of sporadic ALS as ∼12% using a linear mixed model accounting for all SNPs. Our results provide an insight into the genetic structure of sporadic ALS, confirming that common variation contributes to risk and that sufficiently powered studies can identify novel susceptibility loci.

NEK1 variants confer susceptibility to amyotrophic lateral sclerosis

To identify genetic factors contributing to amyotrophic lateral sclerosis (ALS), we conducted whole-exome analyses of 1,022 index familial ALS (FALS) cases and 7,315 controls. In a new screening strategy, we performed gene-burden analyses trained with established ALS genes and identified a significant association between loss-of-function (LOF) NEK1 variants and FALS risk. Independently, autozygosity mapping for an isolated community in the Netherlands identified a NEK1 p.Arg261His variant as a candidate risk factor. Replication analyses of sporadic ALS (SALS) cases and independent control cohorts confirmed significant disease association for both p.Arg261His (10,589 samples analyzed) and NEK1 LOF variants (3,362 samples analyzed). In total, we observed NEK1 risk variants in nearly 3% of ALS cases. NEK1 has been linked to several cellular functions, including cilia formation, DNA-damage response, microtubule stability, neuronal morphology and axonal polarity. Our results provide new and important insights into ALS etiopathogenesis and genetic etiology.

Ubiquilin 2 mutations in Italian patients with amyotrophic lateral sclerosis and frontotemporal dementia

Objectives Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease mainly involving cortical and spinal motor neurones. Molecular studies have recently identified different mutations in the ubiquilin-2 (UBQLN2) gene as causative of a familial form of X-linked ALS, 90% penetrant in women. The aim of our study was to analyse the UBQLN2 gene in a large cohort of patients with familial (FALS) and sporadic (SALS) amyotrophic lateral sclerosis, with or without frontotemporal dementia (FTD), and in patients with FTD. Methods We analysed the UBQLN2 gene in 819 SALS cases, 226 FALS cases, 53 ALS–FTD patients, and 63 patients with a clinical record of FTD. Molecular analysis of the entire coding sequence was carried out in all FALS and ALS–FTD patients, while SALS and FTD patients were analysed specifically for the genomic region coding for the PXX repeat tract. Healthy controls were 845 anonymous blood donors and were screened for the PXX repeat region only. Results We found five different variants in the UBQLN2 gene in five unrelated ALS patients. Three variants, including two novel ones, involved a proline residue in the PXX repeat region and were found in three FALS cases. The other two were novel variants, identified in one FALS and one SALS patient. None of these variants was present in controls, while one control carried a new heterozygous variant. Conclusions Our data support the role of the UBQLN2 gene in the pathogenesis of FALS, being conversely a rare genetic cause in SALS even when complicated by FTD.

Screening of the PFN1 gene in sporadic amyotrophic lateral sclerosis and in frontotemporal dementia

Mutations in the profilin 1 (PFN1) gene, encoding a protein regulating filamentous actin growth through its binding to monomeric G-actin, have been recently identified in familial amyotrophic lateral sclerosis (ALS). Functional studies performed on ALS-associated PFN1 mutants demonstrated aggregation propensity, alterations in growth cone, and cytoskeletal dynamics. Previous screening of PFN1 gene in sporadic ALS (SALS) cases led to the identification of the p.E117G mutation, which is likely to represent a less pathogenic variant according to both frequency data in control subjects and cases, and functional experiments. To determine the effective contribution of PFN1 mutations in SALS, we analyzed a large cohort of 1168 Italian SALS patients and also included 203 frontotemporal dementia (FTD) cases because of the great overlap between these 2 neurodegenerative diseases. We detected the p.E117G variant in 1 SALS patient and the novel synonymous change p.G15G in another patient, but none in a panel of 1512 control subjects. Our results suggest that PFN1 mutations in sporadic ALS and in FTD are rare, at least in the Italian population.

RNA-binding proteins and RNA metabolism: a new scenario in the pathogenesis of Amyotrophic Lateral Sclerosis

Several RNA-processing genes have been implicated in the pathogenesis of Amyotrophic lateral sclerosis (ALS). In particular, causative mutations in the genes encoding for two DNA/RNA binding proteins, TAR DNA binding protein-43 (TDP-43) and fused in sarcoma/translocated in liposarcoma (FUS/TLS), were recently identified in ALS patients. These genetic findings and the presence of abnormal aggregates of these two RNA-binding proteins in ALS affected tissues suggest that molecular mechanisms regulating RNA metabolism are implicated in ALS pathogenesis through common pathways. In this review similarities and differences between TDP-43 and FUS/TLS proteins and their activities in physiological and pathological conditions will be discussed.

No association of DPP6 with amyotrophic lateral sclerosis in an Italian population.

We have attempted to replicate a recently reported association of polymorphism rs10260404, in the Dipeptidyl-peptidase 6 gene (DPP6), with susceptibility to amyotrophic lateral sclerosis (ALS) in a large independent Italian cohort of 904 cases and 1036 controls. Minor allele frequency was 0.38 in cases and 0.39 in controls and no evidence of association with ALS was observed (P=0.638). Our negative results agree with those recently reported in additional Polish and Italian cohorts.