A novel SACS mutation results in non-ataxic spastic paraplegia and peripheral neuropathy.

BACKGROUND AND PURPOSE: Mutations in the SACS gene are commonly associated with autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS), a complex neurodegenerative disorder characterized by progressive degeneration of the cerebellum and spinal cord tracts. The aim of this study was to identify the genetic cause of the disease in an Italian family with spastic paraplegia and peripheral neuropathy. METHODS: Affected subjects were subjected to a comprehensive neurological examination including electromyography and brain magnetic resonance imaging. Genetic studies included exclusion of known disease genes, genome-wide linkage analysis using high density single nucleotide polymorphism genotyping and candidate gene sequencing. RESULTS: Molecular analyses revealed a novel missense mutation in the SACS gene (c.11,104A>G) occurring in a homozygous state in patients and absent in 700 Italian control chromosomes. The mutation led to the amino acid substitution p.Thr3702Ala in the sacsin protein, in a possible protein-protein interaction site of UBE3A binding domain. CONCLUSION: This study broadens the genetic spectrum of SACS mutations and expands the clinical ARSACS phenotype suggesting that the SACS gene can be considered in patients with non-canonical ARSACS clinical presentations.

Familial temporal lobe epilepsy with psychic auras associated with a novel LGI1 mutation.

BACKGROUND: Autosomal dominant lateral temporal epilepsy (ADLTE) is characterized by focal seizures with auditory features or aphasia. Mutations in the LGI1 gene have been reported in up to 50% of ADLTE pedigrees. We report a family with temporal lobe epilepsy characterized by psychic symptoms associated with a novel LGI1 mutation. METHODS: All participants were personally interviewed and underwent neurologic examination and video-EEG recordings. LGI1 exons were sequenced by standard methods. Mutant cDNA was transfected into human embryonic kidney 293 cells; both cell lysates and media were analyzed by Western blot. In silico modeling of the Lgi1 protein EPTP domain was carried out using the structure of WD repeat protein and manually refined. RESULTS: Three affected family members were ascertained, 2 of whom had temporal epilepsy with psychic symptoms (déjà vu, fear) but no auditory or aphasic phenomena, while the third had complex partial seizures without any aura. In all patients, we found a novel LGI1 mutation, Arg407Cys, which did not hamper protein secretion in vitro. Mapping of the mutation on a 3-dimensional protein model showed that this mutation does not induce large structural rearrangements but could destabilize interactions of Lgi1 with target proteins. CONCLUSIONS: The Arg407Cys is the first mutation with no effect on Lgi1 protein secretion. The uncommon, isolated psychic symptoms associated with it suggests that ADLTE encompasses a wider range of auras of temporal origin than hitherto reported.

Adding structural information to the von Hippel-Lindau (VHL) tumor suppressor interaction network.

The von Hippel-Lindau (VHL) tumor suppressor gene is a protein interaction hub, controlling numerous genes implicated in tumor progression. Here we focus on structural aspects of protein interactions for a list of 35 experimentally verified protein VHL (pVHL) interactors. Using structural information and computational analysis we have located three distinct interaction interfaces (A, B, and C). Interface B is the most versatile, recognizing a refined linear motif present in 17 otherwise non-related proteins. It has been possible to distinguish compatible and exclusive interactions by relating pVHL function to interaction interfaces and subcellular localization. A novel hypothesis is presented regarding the possible function of the N-terminus as an inhibitor of pVHL function.

Gamma-glutamyl transferase in the cell wall participates in extracellular glutathione salvage from the root apoplast.

The molecular properties and subcellular location of bound gamma-glutamyl transferase (GGT) were studied, and an experimental setup devised to assess its functions in barley roots. Enzyme histochemistry was used to detect GGT activity at tissue level; immunocytochemistry to localize the protein at subcellular level; and modelling studies to investigate its surface charge properties. GGT activity in vivo was measured for the first time. Functions were explored by applying chemical treatments with inhibitors and the thiol-oxidizing drug diamide, performing time-course chromatographic and spectrophotometric analyses on low-molecular-weight thiols. Gamma-glutamyl transferase activity was found to be high in the root apical region and the protein was anchored to root cell wall components, probably by basic amino acid residues. The results show that GGT is essential to the recovery of apoplastic glutathione provided exogenously or extruded by oxidative treatment. It is demonstrated that GGT activity helps to salvage extracellular glutathione and may contribute to redox control of the extracellular environment, thus providing evidence of a functional role for gamma-glutamyl cycle in roots.

Large-scale prediction of protein structure and function from sequence.

The identification of novel drug targets from genomic data involves the large-scale analysis of many protein sequences. Methods for automated structure and function prediction are an essential tool for this purpose. In this review we concentrate on the recent developments in the field of protein structure prediction and how these can be used to gain hints about the function of proteins. The current state-of-the-art is highlighted through recent community-wide experiments aimed at comparing different approaches. For structure prediction this allows the identification of key improvements to increase the crucial sequence to structure alignment needed for accurate models. Function prediction is a rapidly maturing field that is still being benchmarked. Definitions for protein function are presented and available methods, mostly concentrating on functional site descriptors and structural motifs, presented.