SPG7 mutational screening in spastic paraplegia patients supports a dominant effect for some mutations and a pathogenic role for p.A510V.

Mutations in the SPG7 gene were initially reported in patients with autosomal recessive hereditary spastic paraplegia (HSP). Recent works suggested a dominant effect for some SPG7 mutations. To characterize the SPG7 mutational spectrum in a large cohort of Spanish HSP patients, we sequenced the whole SPG7 gene in a total of 285 Spastic Paraplegia patients. Large gene rearrangements were also ascertained in some patients. We found a total of 14 SPG7 mutations (12 new) in 14 patients; 2 were large deletions. All the mutation carriers had an adult onset age but only five (35%) had a complicated phenotype. We identified a single mutation in 13 patients. Familial analysis suggested a dominant inheritance for one (p.Leu78*) of these mutations. Carriers of the rare p.A510V variant were significantly more frequent in patients vs healthy controls (3% vs 1%), suggesting a pathogenic role for this SPG7 variant. We reported a high frequency of patients with only one SPG7 mutation, and a putative pathogenic role for the p.A510V variant.

First HPSE2 missense mutation in urofacial syndrome.

Urofacial syndrome (UFS) describes the combination of urological problems and an inverted facial expression upon attempts to smile. Seventeen independent familial cases from different ethnicities have been described so far. Some of these have been linked to chromosome 10q. Very recently, homozygous loss-of-function mutations affecting the gene HPSE2 were identified in nine cases. Here, we describe a consanguineous UFS family from Pakistan with three of six siblings affected. We establish linkage to the chromosome 10q critical region and identify two non-synonymous HPSE2 variants. In silico analysis and screening of controls defines c.631T>C (p.Y211H) as a novel benign SNP and c.1628A>T (p.N543I) as the disease-causing mutation. Our study exemplifies the challenges in proper clinical diagnosis of UFS and, thereby, supports the hypothesis of the disease being under diagnosed. By identifying the first HPSE2 missense mutation it also provides a starting point for studies aimed at functionally understanding the unusual combination of symptoms as characterizing UFS.

Comprehensive clinical, biochemical and genetic screening reveals four distinct GBA genotypes as underlying variable manifestation of Gaucher disease in a single family.

Gaucher disease (GD) is a lysosomal storage disorder that is associated with bi-allelic pathogenic variants in GBA. Its wide clinical spectrum, ranging from mild organomegaly to significant skeletal and neurological involvement, is partially explained by genotype-phenotype correlations. We present a family, in which all members over two generations presented with at least splenomegaly. Comprehensive clinical, biochemical and genetic workup was required to diagnose GD, which is caused by as many as four distinct GBA genotypes.

[Hereditary spastic paraplegia type 4 (SPG4) in Russian patients]. / Nasledstvennaia spasticheskaia paraplegiia 4-go tipa u rossiiskikh bol'nykh.

AIM: To investigate molecular, clinical and genealogical characteristics of SPG4 in a first representative Russian group, to estimate SPG4 proportion among all DNA-diagnosed spastic paraplegias. MATERIAL AND METHODS: Fifty unrelated Russian families with SPG4 detected in the course of clinical and molecular studies of spastic paraplegias were studied. Clinical, genealogical and several molecular methods were used, i.e. Sanger sequencing of SPAST, massive parallel sequencing MPS (panel 'hereditary paraplegias') and multiplex ligation-dependent amplification MLPA. RESULTS: SPG4 proportion was 56% among all DNA verified SPG cases (90 families/14 forms) and 68% in subgroup of dominant SPG. In 50 families, 43 different SPAST mutations were detected, of which 21 were novel; percentage of large rearrangements was 30% (13 mutations in 15 families). Four mutations were detected in two families each, nonsense mutation c.1291C>T (p.Arg431*) in 4 unrelated families. Proportion of familial cases was 68%, pedigrees with 'missing' disease in elderly carriers pointed to incomplete penetrance. Age of onset varied from one year to 58 years, middle-age onset was common but the proportion of early-onset cases, particularly in male index cases, was also high. Onset age showed marked intrafamilial differences (more than 10 years in 14 pedigrees, up to 50 year in one) and between families with identical mutations. Insidious onset, slow development with most patients ambulant and 'uncomplicated' phenotype were typical. Cases with additional signs were: a family with ataxia in both patients, two families with epilepsy in one of SPG4 patients; three families with mild mental deficiency in one of SPG4 patients. A case described separately is a 29-year-old male patient with indeterminate myalgia and no SPG signs in whom SPAST previously reported mutation p.Ala430Thr de novo was an unexpected MPS finding. CONCLUSION: SPG4 substantially predomimates in SPG structure in Russian families as practically everywhere else. Half of 43 detected SPAST mutations are novel, the proportion of large rearrangements is 30% higher than in most of studies. Clinical inter- and intrafamilial variability concerns mostly age of onset. SPG4 is not exclusively adult-onset as was thought earlier.

Isoform-specific increase of spastin stability by N-terminal missense variants including intragenic modifiers of SPG4 hereditary spastic paraplegia.

Hereditary spastic paraplegia (HSP) is a neurodegenerative disorder selectively affecting axons of spinal cord motoneurons. Classical mutations in the most frequent HSP gene SPAST (spastin protein) act through haploinsufficiency by abolishing the activity of a C-terminal ATPase domain or by interfering with expression from the affected allele. N-terminal missense variants have been suggested to represent rare polymorphisms, to cause unusually mild phenotypes, and to aggravate the effect of a classical mutation. We confirm these associations for p.S44L but do not detect two other variants (p.E43Q; p.P45Q) in HSP patients and controls. We show that neither of several disease mechanisms associated with classical SPAST mutations applies to the N-terminal variants. Instead, all three alterations enhance the stability of one of two alternative spastin isoforms. Their phenotypic effect may thus not be mediated by haploinsufficiency but by increasing isoform competition for interacting proteins, substrates or oligomerization partners.

Control of foot differentiation in Hydra: in vitro evidence that the NK-2 homeobox factor CnNK-2 autoregulates its own expression and uses pedibin as target gene.

The foot of the simple metazoan Hydra is a highly dynamic body region of constant tissue movement, cell proliferation, determination and differentiation. Previously, two genes have been shown to participate in the development and differentiation of this body region: homeodomain factor CnNK-2 and signal peptide pedibin [Dev. Biol. 180 (1996) 473; Development 126 (1999) 517; Development 122 (1996) 1941; Mech. Dev. 106 (2001) 37]. CnNk-2 functions as transcriptional regulator and is responsive to changes in the positional value while the secreted peptide pedibin serves as "extrinsic" positional signal. Exposure of polyps to pedibin increases the spatial domain of CnNK-2 expression towards the gastric region, indicating that positional signals are integrated at the cis-regulatory region of CnNK-2. In the present study, to elucidate the molecular basis of the interaction of CnNK-2 and pedibin, we characterized the 5' regulatory regions of both genes. Within the CnNK-2 5' upstream region, electrophoretic mobility shift assays showed that putative NK-2 binding motifs are specifically bound by both nuclear protein from Hydra foot and by recombinant CnNK-2, suggesting that CnNK-2 might autoregulate its own expression. This is the first indication for an autoregulatory circuit in Hydra. In addition, we also identified NK-2 binding sites in the cis-regulatory region of the pedibin gene, indicating that this gene is one of the targets of the transcription factor CnNK-2. On the basis of these results, we present a model for the regulatory interactions required for patterning the basal end of the single axis in Hydra which postulates that CnNK-2 together with pedibin orchestrates foot specific differentiation.

Morphological and electrophysiological features of mature neurons in differentiated skin-derived precursor cells.

In vitro modelling of neuronal pathologies is, in particular, demanding and a lot of efforts have been undertaken to differentiate skin derived precursor cells into neuronal cells. However, so far all attempts did not result in cells with functional features of neurons like the ability to generate action potentials or synaptic activity. Here, we report that simple modifications of the protocols result in neuronal cells that display action potentials and synaptic activity. We think that our observation is an important step to model individual neuronal pathologies in vitro.

Dementia in SPG4 hereditary spastic paraplegia: clinical, genetic, and neuropathologic evidence.

BACKGROUND: Cognitive impairment and dementia has been reported in autosomal dominant hereditary spastic paraparesis (HSP) linked to the SPG4 locus. There has only been one postmortem examination described; not all accept that progressive cognitive decline is a feature of this disorder. OBJECTIVE: A family with SPG4-HSP known to have a deletion of exon 17 in the spastin gene (SPG4delEx17) was cognitively assessed over a 7-year period. The index family member died and a postmortem examination was performed. METHODS: Thirteen family members older than 40 years were clinically and cognitively assessed using the Cambridge Cognitive Assessment over a 7-year period. The presence of SPG4delEx17 was assessed; a neuropathologic examination of the brain of the index family member was performed. RESULTS: Cognitive decline occurred in 6 of the 13 family members and in all 4 older than 60 years. Two genetic deletions were identified: SPG4delEx17 in 12 of the 13 family members and a deletion of SPG6 (SPG6del) in 5. Eight individuals had the SPG4delEx17 deletion only; 4 had evidence of progressive cognitive impairment. Four family members had both SPG4delEx17 and SPG6del; 2 of these had cognitive impairment. One family member with the SPG6del alone had neither HSP nor cognitive impairment. The index case with both deletions died with dementia; the brain showed widespread ubiquitin positivity within the neocortex and white matter. CONCLUSION: Cognitive decline and dementia is a feature of SPG4-HSP due to a deletion of exon 17 of the spastin gene.

Unexpected pathogenic mechanism of a novel mutation in the coding sequence of SPG4 (spastin).

The authors report a nucleotide substitution (c.1216A>G) in SPG4 (spastin) causing hereditary spastic paraplegia. This apparent missense mutation in the ATPase domain confers aberrant, in-frame splicing and results in destabilization of mutated transcript. Mutated protein is deficient in microtubule-severing activity but, unlike neighboring mutations, shows regular subcellular localization. The authors' data point to haploinsufficiency rather than a dominant negative effect as the disease-causing mechanism for this mutation.

High frequency of partial SPAST deletions in autosomal dominant hereditary spastic paraplegia.

BACKGROUND: Hereditary spastic paraplegia (HSP) is a genetically heterogeneous neurodegenerative disease. The most frequent cause of autosomal dominant HSP is mutation of SPAST (SPG4 locus), but additional pedigrees remain mutation negative by conventional screening despite linkage to SPG4. OBJECTIVE: To determine the frequency of genomic copy number aberrations of SPAST in autosomal dominant HSP. METHODS: We developed and validated a multiplex ligation-dependent probe amplification assay targeting SPAST and SPG3A, another gene frequently involved in autosomal dominant HSP. In a multicenter study we subsequently investigated 65 index patients with autosomal dominant HSP, all of whom had previously been screened negative for SPAST mutations. Independent secondary samples, additional family members, and cDNA were analyzed to confirm positive findings. RESULTS: Aberrant MLPA profiles were identified in 12 cases (18%). They exclusively affect SPAST, represent deletions, segregate with the disease, and are largely pedigree specific. Internal SPAST deletions entail expression of correspondingly shortened transcripts, which vary in stability. Age at onset in SPAST deletion carriers does not differ from that associated with other SPAST mutations. CONCLUSIONS: Partial SPAST deletions, but not SPAST amplifications and SPG3A copy number aberrations, represent an underestimated cause of autosomal dominant hereditary spastic paraplegia. Partial SPAST deletions are likely to act via haploinsufficiency.

A reinterpretation of the infrared linear dichroism of oriented nucleic acid films and a calculation of some effective partial changes on the ribose phosphate backbone.

In this paper we show, based on symmetry considerations, that structural information cannot be obtained from the linear infrared dichroism of the dioxy vibrations of the phosphate group of nucleic acids. Consequently, the discrepancies between the results of x-ray structure measurements and linear dichroism measurements are not meaningful. The linear dichroism measurements are instead important for a calculation of transition dipole moments that involve both the vibrations of all the atoms of the nucleotide and their charges. Independent information on either the atomic displacements contributing to a given vibration or the atomic charges permits a refinement of the unknown quantities. Based on the molecular dynamics calculations of Prohofsky et al., atomic charges of DNA are calculated to reproduce the observed linear dichroism results. Some of the resulting charges are unexpected and may reflect the inadequacy of the molecular dynamic calculation.