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Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disorder. While there are five FDA-approved drugs for treating this disease, each has only modest benefits. To design new and more effective therapies for ALS, particularly for sporadic ALS of unknown and diverse etiologies, we must identify key, convergent mechanisms of disease pathogenesis. This review focuses on the origin and effects of glutamate-mediated excitotoxicity in ALS (the cortical hyperexcitability hypothesis), in which increased glutamatergic signaling causes motor neurons to become hyperexcitable and eventually die. We characterize both primary and secondary contributions to excitotoxicity, referring to processes taking place at the synapse and within the cell, respectively. 'Primary pathways' include upregulation of calcium-permeable AMPA receptors, dysfunction of the EAAT2 astrocytic glutamate transporter, increased release of glutamate from the presynaptic terminal, and reduced inhibition by cortical interneurons—all of which have been observed in ALS patients and model systems. 'Secondary pathways' include changes to mitochondrial morphology and function, increased production of reactive oxygen species, and endoplasmic reticulum (ER) stress. By identifying key targets in the excitotoxicity cascade, we emphasize the importance of this pathway in the pathogenesis of ALS and suggest that intervening in this pathway could be effective for developing therapies for this disease. [ABSTRACT FROM AUTHOR]

Background: Senataxin (SETX) is a DNA/RNA helicase critical for neuron survival. SETX mutations underlie two inherited neurodegenerative diseases: Ataxia with Oculomotor Apraxia type 2 (AOA2) and Amyotrophic Lateral Sclerosis type 4 (ALS4). Methods: This review examines SETX key cellular processes and we hypothesize that SETX requires SUMO posttranslational modification to function properly. Results: SETX is localized to distinct foci during S‐phase of the cell cycle, and these foci represent sites of DNA polymerase/RNA polymerase II (RNAP) collision, as they co‐localize with DNA damage markers 53BP1 and H2AX. At such sites, SETX directs incomplete RNA transcripts to the nuclear exosome for degradation via interaction with exosome component 9 (Exosc9), a key component of the nuclear exosome. These processes require SETX SUMOylation. SETX was also recently localized within stress granules (SGs), and found to regulate SG disassembly, a process that similarly requires SUMOylation. Conclusion: SETX undergoes SUMO modification to function at S‐phase foci in cycling cells to facilitate RNA degradation. SETX may regulate similar processes in non‐dividing neurons at sites of RNAP II bidirectional self‐collision. Finally, SUMOylation of SETX appears to be required for SG disassembly. This SETX function may be crucial for neuron survival, as altered SG dynamics are linked to ALS disease pathogenesis. In addition, AOA2 point mutations have been shown to block SETX SUMOylation. Such mutations induce an ataxia phenotype indistinguishable from those with SETX null mutation, underscoring the importance of this modification. [ABSTRACT FROM AUTHOR]

Senataxin is a large 303 kDa protein linked to neuron survival, as recessive mutations cause Ataxia with Oculomotor Apraxia type 2 (AOA2), and dominant mutations cause amyotrophic lateral sclerosis type 4 (ALS4). Senataxin contains an amino-terminal protein-interaction domain and a carboxy-terminal DNA/RNA helicase domain. In this study, we focused upon the common ALS4 mutation, L389S, by performing yeast two-hybrid screens of a human brain expression library with control senataxin or L389S senataxin as bait. Interacting clones identified from the two screens were collated, and redundant hits and false positives subtracted to yield a set of 13 protein interactors. Among these hits, we discovered a highly specific and reproducible interaction of L389S senataxin with a peptide encoded by the antisense sequence of a brain-specific non-coding RNA, known as BCYRN1. We further found that L389S senataxin interacts with other proteins containing regions of conserved homology with the BCYRN1 reverse complement-encoded peptide, suggesting that such aberrant protein interactions may contribute to L389S ALS4 disease pathogenesis. As the yeast two-hybrid screen also demonstrated senataxin self-association, we confirmed senataxin dimerization via its amino-terminal binding domain and determined that the L389S mutation does not abrogate senataxin self-association. Finally, based upon detection of interactions between senataxin and ubiquitin–SUMO pathway modification enzymes, we examined senataxin for the presence of ubiquitin and SUMO monomers, and observed this post-translational modification. Our senataxin protein interaction study reveals a number of features of senataxin biology that shed light on senataxin normal function and likely on senataxin molecular pathology in ALS4. [ABSTRACT FROM AUTHOR]

The mouse scurfy gene, Foxp3, and its human orthologue, FOXP3, which maps to Xp11.23–Xq13.3, were recently identified by positional cloning. Point mutations and microdeletions of the FOXP3 gene were found in the affected members of eight of nine families with IPEX (immune dysfunction, polyendocrinopathy, enteropathy, X-linked; OMIM 304930). We evaluated a pedigree with clinically typical IPEX in which mutations of the coding exons of FOXP3 were not detected. Our reevaluation of this pedigree identified an A→G transition within the first polyadenylation signal (AAUAAA→AAUGAA) after the stop codon. The next polyadenylation signal is not encountered for a further 5.1 kb. This transition was not detected in over 212 normal individuals (~318 X chromosomes), excluding the possibility of a rare polymorphism. We suggest that this mutation is causal of IPEX in this family by a mechanism of nonspecific degradation of the FOXP3 gene message. [ABSTRACT FROM AUTHOR]

IPEX is a fatal disorder characterized by immune dysregulation, polyendocrinopathy, enteropathy and X-linked inheritance (MIM 304930). We present genetic evidence that different mutations of the human gene FOXP3, the ortholog of the gene mutated in scurfy mice (Foxp3), causes IPEX syndrome. Recent linkage analysis studies mapped the gene mutated in IPEX to an interval of 17?20-cM at Xp11.23?Xq13.3 (refs. 1,2). [ABSTRACT FROM AUTHOR]

Presents a letter to the editor announcing the identification of a new gene for Charcot &ndash Marie &ndash Tooth neuropathy type 1.

Analyzes the mapping of X-linked syndrome of polyendocrinopathy immune dysfunction and diarrhea (XPID) genes. Analysis on the linkage map of the XPID gene on X chromosome at Xp11.23-Xq13.3; Investigation on the mutation of Wiskott-Aldrich syndrome gene; Segregation of the XPID phenotype.

Summary: The Purkinje cell degeneration (pcd) mouse is a recessive model of neurodegeneration, involving cerebellum and retina. Purkinje cell death in pcd is dramatic, as >99% of Purkinje neurons are lost in 3 weeks. Loss of function of Nna1 causes pcd, and Nna1 is a highly conserved zinc carboxypeptidase. To determine the basis of pcd, we implemented a two-pronged approach, combining characterization of loss-of-function phenotypes of the Drosophila Nna1 ortholog (NnaD) with proteomics analysis of pcd mice. Reduced NnaD function yielded larval lethality, with survivors displaying phenotypes that mirror disease in pcd. Quantitative proteomics revealed expression alterations for glycolytic and oxidative phosphorylation enzymes. Nna proteins localize to mitochondria, loss of NnaD/Nna1 produces mitochondrial abnormalities, and pcd mice display altered proteolytic processing of Nna1 interacting proteins. Our studies indicate that Nna1 loss of function results in altered bioenergetics and mitochondrial dysfunction. [Copyright &y& Elsevier]

Senataxin (SETX) is a DNA-RNA helicase whose C-terminal region shows homology to the helicase domain of the yeast protein Sen1p. Genetic discoveries have established the importance of SETX for neural function, as recessive mutations in the SETX gene cause Ataxia with Oculomotor Apraxia type 2 (AOA2) (OMIM: 606002), which is the third most common form of recessive ataxia, after Friedreich's ataxia and Ataxia-Telangiectasia. In addition, rare, dominant SETX mutations cause a juvenile-onset form of Amyotrophic Lateral Sclerosis (ALS), known as ALS4. SETX performs a number of RNA regulatory functions, including maintaining RNA transcriptome homeostasis. Over the last decade, altered RNA regulation and aberrant RNA-binding protein function have emerged as a central theme in motor neuron disease pathogenesis, with evidence suggesting that sporadic ALS disease pathology may overlap with the molecular pathology uncovered in familial ALS. Like other RNA processing proteins linked to ALS, the basis for SETX gain-of-function motor neuron toxicity remains ill-defined. Studies of yeast Sen1p and mammalian SETX protein have revealed a range of important RNA regulatory functions, including resolution of R-loops to permit transcription termination, and RNA splicing. Growing evidence suggests that SETX may represent an important genetic modifier locus for sporadic ALS. In cycling cells, SETX is found at nuclear foci during the S/G 2 cell-cycle transition phase, and may function at sites of collision between components of the replisome and transcription machinery. While we do not yet know which SETX activities are most critical to neurodegeneration, our evolving understanding of SETX function will undoubtedly be crucial for not only understanding the role of SETX in ALS and ataxia disease pathogenesis, but also for delineating the mechanistic biology of fundamentally important molecular processes in the cell.

Interest in senataxin biology began in 2004 when mutations were first identified in what was then a novel protein. Dominantly inherited mutations were documented in rare juvenile-onset, motor neuron disease pedigrees in a familial form of amyotrophic lateral sclerosis (ALS4), while recessive mutations were found to cause a severe early-onset ataxia with oculomotor apraxia (AOA2) that is actually the second most common recessive ataxia after Freidreich's ataxia. From earlier studies of sen1p, the yeast ortholog of senataxin, a range of important RNA processing functions have been attributed to this protein. Like sen1p, senataxin contains a helicase domain to interact with RNA and an amino-terminal domain for critical protein interactions. Senataxin also joins a group of important proteins responsible for maintaining RNA transcriptome homeostasis, including FUS, TDP-43, and SMN that can all cause familial forms of motor neuron disease (MND). Independent of this association, senataxin is gaining attention for its role in maintaining genomic stability. Senataxin has been shown to resolve R-Loop structures, which form when nascent RNA hybridizes to DNA, displacing the non-transcribed strand. But in cycling cells, senataxin is also found at nuclear foci during the S/G2 cell-cycle phase, and may function at sites of specific collision between components of the replisome and transcription machinery. Which of these important processes is most critical to prevent neurodegeneration remains unknown, but our evolving understanding of these processes will be crucial not only for understanding senataxin's role in neurological disease, but also in a number of fundamentally important cellular functions.

One of the major challenges facing the long term survival of neurons is their requirement to maintain efficient axonal transport over long distances. In humans as large, long-lived vertebrates, the machinery maintaining neuronal transport must remain efficient despite the slow accumulation of cell damage during aging. Mutations in genes encoding proteins which function in the transport system feature prominently in neurologic disorders. Genes known to cause such disorders and showing traditional Mendelian inheritance have been more readily identified. It has been more difficult, however, to isolate factors underlying the complex genetics contributing to the more common idiopathic forms of neurodegenerative disease. At the heart of neuronal transport is the rail network or scaffolding provided by neuron specific microtubules (MTs). The importance of MT dynamics and stability is underscored by the critical role tau protein plays in MT-associated stabilization versus the dysfunction seen in Alzheimer's disease, frontotemporal dementia and other tauopathies. Another example of the requirement for tight regulation of MT dynamics is the need to maintain balanced levels of post-translational modification of key MT building-blocks such as α-tubulin. Tubulins require extensive polyglutamylation at their carboxyl-terminus as part of a novel post-translational modification mechanism to signal MT growth versus destabilization. Dramatically, knock-out of a gene encoding a deglutamylation family member causes an extremely rapid cell death of Purkinje cells in the ataxic mouse model, pcd . This review will examine a range of neurodegenerative conditions where current molecular understanding points to defects in the stability of MTs and axonal transport to emphasize the central role of MTs in neuron survival.

Mutation of the SIMPLE gene (small integral membrane protein of the lysosome/late endosome) is the molecular basis of Charcot-Marie-Tooth disease type 1C (CMT1C), a demyelinating peripheral neuropathy. Although the precise function of SIMPLE is unknown, prior reports suggest it localizes to the lysosome/late endosome. Furthermore, murine Simple interacts with Nedd4 (neural precursor cell expressed, developmentally downregulated 4), an E3 ubiquitin ligase that is important for regulating lysosomal degradation of plasma membrane proteins. To bring insights into the biochemical function of human SIMPLE, we confirmed that human SIMPLE interacts with NEDD4 and also report a novel interaction with tumor susceptibility gene 101 (TSG101), a class E vacuolar sorting protein. TSG101 is known to function downstream of NEDD4, sorting ubiquitinated substrates into multivesicular bodies (MVBs), which then deliver their cargo into the lysosomal lumen for degradation. Given the interaction with NEDD4 and TSG101, and the localization of SIMPLE along the lysosomal degradation pathway, we hypothesize that SIMPLE plays a role in the lysosomal sorting of plasma membrane proteins. We examine three CMT1C-associated SIMPLE mutations and show that they do not affect the interaction with NEDD4 or TSG101, nor do they lead to altered subcellular localization., ((c) 2005 Wiley-Liss, Inc.)

Charcot-Marie-Tooth neuropathy type 1C (CMT1C) is an autosomal dominant demyelinating peripheral neuropathy caused by missense mutations in the small integral membrane protein of lysosome/late endosome (SIMPLE) gene. To investigate the prevalence of SIMPLE mutations, we screened a cohort of 152 probands with various types of demyelinating or axonal and pure motor or sensory inherited neuropathies. SIMPLE mutations were found only in CMT1 patients, including one G112S and one W116G missense mutations. A novel I74I polymorphism was identified, yet no splicing defect of SIMPLE is likely. Haplotype analysis of STR markers and intragenic SNPs linked to the gene demonstrated that families with the same mutation are unlikely to be related. The clustering of the G112S, T115N, and W116G mutations within five amino acids suggests this domain may be critical to peripheral nerve myelination. Electrophysiological studies showed that CMT1C patients from six pedigrees (n = 38) had reduced nerve conduction velocities ranging from 7.5 to 27.0m/sec (peroneal). Two patients had temporal dispersion of nerve conduction and irregularity of conduction slowing, which is unusual for CMT1 patients. We report the expression of SIMPLE in various cell types of the sciatic nerve, including Schwann cells, the affected cell type in CMT1C.

Joubert syndrome (JS) is a rare autosomal recessive malformation syndrome, involving dysgenesis of the cerebellar vermis with accompanying brainstem malformations (comprising the molar tooth sign). JS is characterized by hypotonia, developmental delay, intermittent hyperpnea and apnea, and abnormal eye movements. A single locus for JS was previously identified on 9q34 in a consanguineous family of Arabian origin. However, linkage to this locus has subsequently been shown to be rare. We have ascertained 35 JS pedigrees for haplotype segregation analysis of genetic loci for genes with a putative role in cerebellar development. We examined the ZIC1 gene as a functional candidate for JS as Zic1(-/-) null mice have a phenotype reminiscent of JS. We undertook mutational analysis of ZIC1 by standard mutational analysis (dideoxy-fingerprinting (ddf)) of 47 JS probands, and fully sequenced the coding region in five of these probands. By these means, ZIC1 was excluded from playing a causal role in most cases of JS as no disease-associated mutations were identified. Further, linkage to the ZIC1 genetic locus (3q24) was excluded in 21 of 35 pedigrees by haplotype segregation analysis of closely spaced markers. The remaining 14 of 35 pedigrees were consistent with linkage. However, this number does not significantly depart from that expected by random chance (16.5) for this cohort. Therefore, this systematic approach has been validated as a means to prioritize functional candidate genes and enables us to confine mutational analysis to only those probands whose segregation is consistent with linkage to any given locus., (Copyright 2003 Wiley-Liss, Inc.)

Joubert syndrome (JS) is a rare autosomal recessive malformation syndrome involving agenesis or dysgenesis of the cerebellar vermis with accompanying brainstem malformations. JS is further characterized by hypotonia, developmental delay, intermittent hyperpnea, and abnormal eye movements. The biochemical and molecular basis of JS remains unknown, although several genes that are crucial in the development of the cerebellum have been proposed as attractive candidate genes. JS is clinically heterogeneous; this, together with previous linkage analyses, suggests that there may also be genetic heterogeneity. A locus for JS was previously identified on chromosome 9q34 by linkage analysis in a consanguineous family of Arabian origin. A putative second JS locus was recently suggested when a deletion on chromosome 17p11.2 was observed in a patient with Smith-Magenis syndrome and JS phenotype. We have investigated a cohort of apparently unrelated North American JS pedigrees for association with the loci on chromosomes 9q34 and 17p11.2 and excluded them in all cases where data were informative. Analysis of an additional 21 unrelated JS patients showed no evidence of homozygosity at the 9q34 and 17p11.2 loci that would suggest inheritance of founder JS mutation(s) or unreported consanguinity. Together, these data suggest that one or more major loci for JS remain to be identified. Consequently, we undertook mutation analysis of several functional candidate genes, EN1, EN2, and FGF8, in a total of 26 unrelated JS patients. Our data suggest that all of these genes may be excluded from a direct pathogenic role in JS. The BARHL1 gene, which localizes to chromosome 9q34 and has previously been proposed as a strong positional candidate gene for JS, was also investigated and excluded from involvement in JS that is linked to chromosome 9q34., (Copyright 2002 Wiley-Liss, Inc.)