FUS/TLS gene mutations are the second most frequent cause of familial ALS in the Spanish population.

Our objective was to investigate the prevalence of FUS/TLS mutations in a Catalan familial ALS cohort undergoing a mutational study for SOD1 in 2006. We screened 25 probands from non-SOD1 families for FUS/TLS mutations. We identified two FALS probands with FUS/TLS mutations. One carried a C-to-T transition at nucleotide position 1561 (c.1561C>T) producing a p.R521C sequence change at protein level. The phenotype was characterized by a young age at onset (38.2 years old), proximal limb girdle weakness, predominant lower motor neuron signs and dropped head. Survival time ranged from 10 to 36 months. Obligate asymptomatic carriers were detected. Our second ALS6 pedigree carried a C-to-T transition at nucleotide position 1528 (c.1528G>A) producing a p.K510E sequence change at protein level. The phenotype was of an early onset (<40 years old), predominant lower motor neuron disease with short survival (nine months). In conclusion, these are the first two FUS/TLS mutations identified in Spain. The prevalence of this form of FALS (8%) is similar to the Dutch and British populations. FUS/TLS mutations are the second most common cause of FALS in our population.

I112M SOD1 mutation causes ALS with rapid progression and reduced penetrance in four Mediterranean families.

We evaluated a possible genotype-phenotype correlation and looked for a founder effect in four Mediterranean families carrying the I112M SOD1 mutation. The structural characteristics of the mutated protein were also analysed. Clinical data of FALS subjects from four families were evaluated. Mutational analysis of the SOD1 gene was carried out by direct sequencing. A haplotype study was carried out using 11 polymorphic markers flanking the SOD1 gene. Structural analysis was performed by means of homology modelling and molecular graphics methods. The clinical pattern of 17 FALS patients was characterized by prevalent spinal onset, mean age at onset of 47.1 years and mean duration of 20.7 months. Several obligate carriers were observed. These findings indicate that the I112M mutation is consistently associated with a uniform, fast-progressing phenotype with reduced penetrance of the disease. The haplotype analysis did not show a common haplotype among the Spanish families and the Italian family; however, a possible common founder could be hypothesized for Spanish families. From a structural viewpoint, mutation at codon 112 seems to confer a severe phenotype, probably related to altered protein functionality.

Profilin induces lamellipodia by growth factor-independent mechanism.

Profilin has been implicated in cell motility and in a variety of cellular processes, such as membrane extension, endocytosis, and formation of focal complexes. In vivo, profilin replenish the pool of ATP-actin monomers by increasing the rate of nucleotide exchange of ADP-actin for ATP-actin, promoting the incorporation of new actin monomers at the barbed end of actin filaments. For this report, we generated a membrane-permeable version of profilin I (PTD4-PfnI) for the alteration of intracellular profilin levels taking advantage of the protein transduction technique. We show that profilin I induces lamellipodia formation independently of growth factor presence in primary bovine trabecular meshwork (BTM) cells. The effects are time- and concentration-dependent and specific to the profilin I isoform. Profilin II, the neuronal isoform, failed to extend lamellipodia in the same degree as profilin I. H133S, a mutation in the polyproline binding domain, showed a reduced ability to induce lamellipodia. H199E, mutation in the actin binding domain failed to induce membrane spreading and inhibit fetal bovine serum (FBS) -induced lamellipodia extension. Incubation with a synthetic polyproline domain peptide (GP5)3, fused to a transduction domain, abolished lamellipodia induction by profilin or FBS. Time-lapse microscopy confirmed the effects of profilin on lamellipodia extension with a higher spreading velocity than FBS. PTD4-Pfn I was found in the inner lamellipodia domain, at the membrane leading edge where it colocalizes with endogenous profilin. While FBS-induced lamellipodia formation activates Rac1, PTD4-Pfn I stimulation did not induce Rac1 activation. We propose a role of profilin I favoring lamellipodia formation by a mechanism downstream of growth factor.

Haplotype Analysis of the First A4V-SOD1 Spanish Family: Two Separate Founders or a Single Common Founder?

Despite the genetic heterogeneity reported in familial amyotrophic lateral sclerosis (ALS) (fALS), Cu/Zn superoxide-dismutase (SOD1) gene mutations are the second most common cause of the disease, accounting for around 20% of all families (ALS1) and isolated sporadic cases (sALS). At least 186 different mutations in the SOD1 gene have been reported to date. The possibility of a single founder and separate founders have been investigated for D90A (p.D91A) and A4V (p.A5V), the most common mutations worldwide. High-throughput single nucleotide polymorphism genotyping studies have suggested two founders for A4V (one for the Amerindian population and another for the European population) although the possibility that the two populations are descended from a single ancient founder cannot be ruled out. We used 15 genetic variants spanning the human chromosome 21 from the SOD1 gene to the SCAF4 gene, comparing them with the population reference panels, to demonstrate that the first A4V Spanish pedigree shared the genetic background reported in the European population.

PFN1 mutations are also rare in the Catalan population with amyotrophic lateral sclerosis.

Evidence of genetic heterogeneity in ALS has been found, with at least 31 genes being identified to date as causing ALS, and other genes being suggested as risk factors for susceptibility to the disease and for phenotype modifications. In recent years, new molecular genetic methodologies, especially GWAS and exome sequencing, have contributed to the identification of new ALS genes. Some of these genes (SOD1, TARDBP, FUS, and C9orf72) have homogenous frequencies in different populations. However, a few genes are rare in populations other than those in which they were first identified. Here we investigate the frequency of the PFN1 gene in a Catalan ALS population. A mutational analysis of the PFN1 gene was carried out on a Catalan cohort of 42 ALS families (FALS) and 423 sporadic ALS patients (SALS). The screening included 600 healthy controls. No PFN1 mutations were identified in either the FALS or SALS group. We also found no mutations in the control group. Our results are consistent with those described in other populations with very low frequencies, suggesting that PFN1 is a very rare cause of ALS worldwide. Together with the absence of a distinctive phenotype associated with ALS18, these results mean that this gene should be a second or third line for inclusion in screening in patients requesting genetic counseling.

The p.E22G mutation in the Cu/Zn superoxide-dismutase gene predicts a long survival time: clinical and genetic characterization of a seven-generation ALS1 Spanish pedigree.

BACKGROUND: Despite the genetic heterogeneity reported in familial ALS (FALS), Cu/Zn superoxide-dismutase (SOD1) gene mutations are the most frequent cause of FALS, accounting for around 20% of familial cases (ALS1) and isolated sporadic cases. Some mutations are associated with a long survival time, while others are linked to a very rapid progression. Clinical-genetic characterization of ALS1 families is therefore important as it can provide information on the phenotype associated with a given mutation, the distribution of SOD1 mutations in different ethnic groups, and can clarify the genotype-phenotype correlation in patients with SOD1 gene mutations. OBJECTIVES: To describe the phenotype linked to this previously reported SOD1 gene mutation, p.E22G (E21G in the old nomenclature), in a large ALS1 Spanish kindred. This mutation was previously reported in a Canadian family but no clinical information was available. METHODS: Clinical characterization including gender, age at onset, site of onset and survival time was available from 15 affected members belonging to a seven-generation pedigree. The possibility of gender predominance or anticipation was also analyzed. DNA samples were available from three of the living symptomatic members. Informed consent for blood samples was obtained. We used direct sequencing to screen for SOD1 gene mutations. RESULTS: An A-to-G transition at nucleotide position 65 (c.65A>G) leading to a p.E22G sequence change at protein level was identified in the three affected ALS patients. The phenotype was similar in all affected members in our p.E22G family. Initial symptoms occurred in the distal limb muscles, predominantly in the legs, and there was a mean survival time of 13.2+/-8.6 years. The mean age at onset was 51.8+/-10.1. The prevalence in males and females was similar, with no difference in phenotype as regards gender. The age range for onset of symptoms was between 38 and 71 years of age, although 60% of the members presented symptoms before their fiftieth birthday. The information available for five affected parent/affected offspring pairs suggested no apparent anticipation. CONCLUSIONS: p.E22G is the ninth SOD1 gene mutation reported in Spain, and the third of these to be associated with long survival (the other two being p.G38R--previously G37R, and p.D77V--previously D76V). Our results emphasize the importance of genetic and clinical characterization of ALS1 families around the world for understanding the genotype-phenotype relationships of each SOD1 gene mutant and their relative frequency in different ethnic groups worldwide.