J Neurol Neurosurg Psychiatry 80:354-355 doi:10.1136/jnnp.2008.157677
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TDP-43 M311V mutation in familial amyotrophic lateral sclerosis

  1. R Lemmens1,2,3,
  2. V Race4,
  3. N Hersmus1,2,
  4. G Matthijs4,
  5. L Van Den Bosch1,2,
  6. P Van Damme1,2,3,
  7. B Dubois1,3,
  8. S Boonen5,
  9. A Goris1,
  10. W Robberecht1,2,3
  1. 1
    Experimental Neurology, University of Leuven, Leuven, Belgium
  2. 2
    Vesalius Research Centre, Flanders Institute for Biotechnology (VIB), Leuven, Belgium
  3. 3
    Department of Neurology, University Hospital Leuven, University of Leuven, Leuven, Belgium
  4. 4
    Molecular Diagnostic Laboratory, Centre for Human Genetics, University of Leuven, Leuven, Belgium
  5. 5
    Gerontology and Geriatrics Section, Leuven University Department of Experimental Medicine, University of Leuven, Leuven, Belgium
  1. Dr R Lemmens, Department of Neurology, Herestraat 49, B-3000 Leuven, Belgium; robin.lemmens{at}
  • Received 7 July 2008
  • Revised 20 August 2008
  • Accepted 23 August 2008

Recently, mutations in TARDBP have been identified in sporadic amyotrophic lateral sclerosis (SALS), an adult onset motor neurodegenerative disorder, as well as in familial forms (FALS) of ALS.

Here we report an A to G (931 A>G) mutation in exon 6 of TARDBP, resulting in the substitution of methionine by valine at codon 311, in one patient with autosomal dominant FALS. This mutation was absent in 601 healthy controls (1202 chromosomes). Our data suggest a novel pathogenic missense mutation in exon 6 of TARDBP (M311V) causing ALS, confirming the aetiological role of the TAR DNA binding protein 43 (TDP-43) in the pathogenesis of motor neurodegeneration.

ALS is a motor neurodegenerative disease which is pathologically characterised by the selective loss of motor neurons in the motor cortex, brainstem and spinal cord.1 Furthermore, ubiquinated inclusions have been shown to be present in these motor neurons. TDP-43 was found to be a major component of these inclusions in ALS as well as in frontotemporal dementia (FTD). The exact pathogenic mechanism of TDP-43 has not yet been established but it is hypothesised that under pathological conditions TDP-43 is eliminated from the nuclei, resulting in decreased TDP-43 nuclear function.2

Approximately 5–10% of ALS cases are familial (FALS) of which 20% are caused by mutations in superoxide dismutase 1 (SOD1). The pathological findings lead to genetic screening of TARDBP in FTD and ALS patients. We and others failed to identify mutations in FTD and in patients with SALS.3 In contrast, others recently reported mutations in TARDBP in both SALS and FALS.4 5 All but one of these mutations are localised in exon 6 of the gene, which encodes the hnRNP (heterogeneous nuclear ribonucleoparticles) binding domain of the gene. We have now sequenced the TARDBP gene in FALS patients and identified one novel mutation.

A total of 72 patients with FALS from 31 families were recruited from the University Hospital Leuven. Patients were diagnosed according to the El Escorial criteria. Control individuals from the same population were recruited among spouses of patients suffering from neurological diseases (ALS, ischaemic stroke or multiple sclerosis) and from the Leuven University Gerontology Database in which mean age was 62.5 (13.5) years and 52.2% were men. The study was approved by the local ethics committee and patients provided informed consent.

DNA was extracted from venous blood samples using standard methods. The coding exons of TARDBP (exons 2–6) were amplified by PCR. Primer sequences were described previously3 but to facilitate the cycle sequencing analysis, the M13 forward primer sequence or the M13 reverse primer sequence was attached to the 5′ end of the sense primers or the antisense primers. Controls were analysed for the identified mutation using a Taqman based custom genotyping assay run on a 7300 real time PCR system.

Thirty-one families were included in this study and mutations in SOD1 were identified in 11 (six families with the G93C mutation, four with the D90A and one L38V). In the other 26 subjects from 20 families with FALS, SOD1 mutations were excluded by sequencing. Thereafter, the TARDBP gene was sequenced in the same individuals. A heterozygous missense mutation, 931 A>G, in exon 6, leading to the amino acid change M311V, was identified in one subject (fig 1, subject III:1). In exon 6 of TARDBP, this methionine is evolutionary well conserved from Homo sapiens to Xenopus tropicalis, as shown in fig 1C. We designed a Taqman based custom allelic discrimination assay to screen 601 controls for this mutation and found it to be absent in this healthy population.

Figure 1 The TAR DNA binding protein 43 (TDP-43) M311V mutation in exon 6. (A) Pedigree of family FALS18 shows affected individuals. M311V mutation was identified in subject III:1. (B) DNA sequence illustrating the single base substitution 931 A>G resulting in changing a methionine to a valine. (C) Alignment of TARDBP sequences in different species illustrates the highly conserved region in which the M311V mutation (which is indicated in red) is localised. ALS, amyotrophic lateral sclerosis.

The family in which the M311V mutation was detected was Caucasian, of Belgian origin, and affected members are shown in fig 1. Interestingly, subject III:1 presented at the age of 50 years with bulbar symptoms of dysarthria and dysphagia, and emotional lability suggesting frontal disturbances, but no formal cognitive assessment was performed. Within 19 months she rapidly progressed and became anarthric and required a feeding tube. Subsequently, she died after a disease course of 30 months. No autopsy was performed.

Her mother originally complained of weakness in the left arm and leg at the age of 38 years and afterwards developed bulbar symptoms. She died 3 years after her initial symptoms at the age of 41 years and no DNA of this patient was extracted. Detailed clinical information of the other affected members was not available. The siblings III:2 and III:4 have been neurologically examined at regular intervals and found to be normal.

The presence of ubiquitin positive tau negative neuronal cytoplasmic inclusions in motor neurons is a pathological hallmark of ALS. Interestingly, these inclusions contain TDP-43.2 Their pathophysiological relevance remains to be established but it is hypothesised that under pathological conditions, TDP-43 is drained away from the nucleus and trapped into the cytoplasmic compartment, resulting in loss of nuclear function.2

The mutation identified in our cohort of patients with FALS is novel and resulted in an amino acid change (M311V) in the hnRNP binding domain of TARDBP.

Since the other affected members in this reported family had deceased before DNA collection, we were not able to show segregation. However, the exon 6 931A>G mutation was not observed in 601 control individuals (1202 chromosomes) from the same population. The 95% upper confidence limit for the frequency of this variant in the control population is 0.0025. Previously, no variants in exon 6 were observed on sequencing of >1400 healthy Caucasian individuals.4 5 Furthermore, the M311 residue is evolutionary conserved between species. The high degree of conservation and the clustering in the same functional domain with several previously reported pathogenic TARDP mutations strongly support the M311V mutation described here as a novel pathogenic mutation causing ALS.

In conclusion, in our Belgian cohort of FALS patients, SOD1 mutations were responsible for 35% of all cases while TARDBP was only identified in one family and therefore seemed to account for just a minority of FALS (3%) in this population. Other reported populations have shown both higher and lower rates of TARDBP mutations, suggesting that the indication for sequencing of TARDBP (for instance exon 6) might be population dependent.


  • Competing interests: None.

  • Funding: This study was supported by grants from the University of Leuven. WR is supported through the E von Behring Chair for Neuromuscular and Neurodegenerative Disorders, and by the Interuniversity Attraction Poles (IUAP) program P6/43 of the Belgian Federal Science Policy Office. AG and PVD are postdoctoral fellows of the Research Foundation Flanders (FWO-Vlaanderen). SB and BD are senior clinical investigators of the Fund for Scientific Research Flanders (FWO-F). All authors had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis.

  • Patient consent: Obtained.

  • Ethics approval: The study was approved by the local ethics committee.


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