Paraplegin gene analysis in hereditary spastic paraparesis ( HSP ) pedigrees in northeast england. (2001 Mar)
paraplegin gene analysis in hereditary spastic paraparesis ( HSP ) pedigrees in northeast england . objective : To identify the frequency and characterize the phenotype of paraplegin mutations in the hereditary spastic paraparesis ( HSP ) population in the northeast of england . background : HSP is a disorder that shows both clinical and genetic heterogeneity . To date , 13 loci have been associated with an HSP phenotype , with the causative gene having been identified in four of these . Two autosomal genes have been identified , paraplegin and spastin , and two X linked genes have been identified , l1cam ( cell adhesion molecule ) and proteolipid protein . methods : thirty HSP pedigrees from the northeast of england were analyzed for mutation in each of the 17 exons of the paraplegin gene . results : A single family with a paraplegin mutation was identified in which the paraplegin mutation co segregates with an HSP phenotype in an apparent dominant manner . The authors also describe frequent polymorphism in the paraplegin gene in both the HSP and control populations . conclusion : mutations in the paraplegin gene are not a common cause of HSP in the northeast of england . The phenotype of the paraplegin related HSP family described had several striking features including amyotrophy , raised creatine kinase , sensorimotor peripheral neuropathy , and oxidative phosphorylation defect on muscle biopsy .

Quantitative and functional analyses of spastin in the nervous system : implications for hereditary spastic paraplegia. (2008 Feb)
quantitative and functional analyses of spastin in the nervous system : implications for hereditary spastic paraplegia . spastin and P60 katanin are two distinct microtubule severing proteins . autosomal dominant mutations in the SPG4 locus corresponding to spastin are the most common cause of hereditary spastic paraplegia ( HSP ) , a neurodegenerative disease that afflicts the adult corticospinal tracts . Here we sought to evaluate whether SPG4 based HSP is best understood as a loss of function disease . using various rat tissues , we found that P60 katanin levels are much higher than spastin levels during development . In the adult , P60 katanin levels plunge dramatically but spastin levels decline only slightly . quantitative data of spastin expression in specific regions of the nervous system failed to reveal any obvious explanation for the selective sensitivity of adult corticospinal tracts to loss of spastin activity . An alternative explanation relates to the fact that the mammalian spastin gene has two start codons , resulting in a 616 amino acid protein called M1 and a slightly shorter protein called M85 . We found that M1 is almost absent from developing neurons and most adult neurons but comprises 20 25 of the spastin in the adult spinal cord , the location of the axons that degenerate during HSP . experimental expression in cultured neurons of a short dysfunctional M1 polypeptide ( but not a short dysfunctional M85 peptide ) is deleterious to normal axonal growth . In squid axoplasm , …

Structural basis of microtubule severing by the hereditary spastic paraplegia protein spastin. (2008 Jan)
structural basis of microtubule severing by the hereditary spastic paraplegia protein spastin . spastin , the most common locus for mutations in hereditary spastic paraplegias , and katanin are related microtubule severing AAA atpases involved in constructing neuronal and non centrosomal microtubule arrays and in segregating chromosomes . The mechanism by which spastin and katanin break and destabilize microtubules is unknown , in part owing to the lack of structural information on these enzymes . Here we report the X ray crystal structure of the drosophila spastin AAA domain and provide a model for the active spastin hexamer generated using small angle X ray scattering combined with atomic docking . The spastin hexamer forms a ring with a prominent central pore and six radiating arms that may dock onto the microtubule . helices unique to the microtubule severing AAA atpases surround the entrances to the pore on either side of the ring , and three highly conserved loops line the pore lumen . mutagenesis reveals essential roles for these structural elements in the severing reaction . peptide and antibody inhibition experiments further show that spastin may dismantle microtubules by recognizing specific features in the carboxy terminal tail of tubulin . collectively , our data support a model in which spastin pulls the C terminus of tubulin through its central pore , generating a mechanical force that destabilizes tubulin tubulin interactions within the microtubule lattice . Our work also provides insights into the structural defects in spastin that arise from mutations …

Human spastin has multiple microtubule related functions. (2005 Nov)
human spastin has multiple microtubule related functions . hereditary spastic paraplegias ( HSPs ) are neurodegenerative diseases caused by mutations in more than 20 genes , which lead to progressive spasticity and weakness of the lower limbs . The most frequently mutated gene causing autosomal dominant HSP is SPG4 , which encodes spastin , a protein that belongs to the family of atpases associated with various cellular activities ( AAAs ) . A number of studies have suggested that spastin regulates microtubule dynamics . We have studied the atpase activity of recombinant human spastin and examined the effect of taxol stabilized microtubules on this activity . We used spastin translated from the second ATG and provide evidence that this is the physiologically relevant form . We showed that microtubules enhance the atpase activity of the protein , a property also described for katanin , an AAA of the same spastin subgroup . furthermore , we demonstrated that human spastin has a microtubule destabilizing activity and can bundle microtubules in vitro , providing new insights into the molecular pathogenesis of HSP .

Novel mutations in spastin gene and absence of correlation with age at onset of symptoms. (2000 Nov)
novel mutations in spastin gene and absence of correlation with age at onset of symptoms . autosomal dominant hereditary spastic paraplegia is genetically heterogeneous , with at least five loci identified by linkage analysis . recently , mutations in spastin were identified in SPG4 , the most common locus for dominant hereditary spastic paraplegia that was previously mapped to chromosome 2p22 . We identified five novel mutations in the spastin gene in five families with SPG4 mutations from north america and tunisia and showed the absence of correlation between the predicted mutant spastin protein and age at onset of symptoms .

Mutations of SPG4 are responsible for a loss of function of spastin , an abundant neuronal protein localized in the … (2002 Dec)
mutations of SPG4 are responsible for a loss of function of spastin , an abundant neuronal protein localized in the nucleus . mutations of spastin are responsible for the most common autosomal dominant form of hereditary spastic paraplegia ( AD HSP ) , a disease characterized by axonal degeneration of corticospinal tracts and posterior columns . generation of polyclonal antibodies specific to spastin has revealed two isoforms of 75 and 80 kDa in both human and mouse tissues with a tissue specific variability of the isoform ratio . spastin is an abundant protein in neural tissues and immunolabeling experiments have shown that spastin is expressed in neurons but not in glial cells . these data indicate that axonal degeneration linked to spastin mutations is caused by a primary defect of neurons . protein and transcript analyses of patients carrying either nonsense or frameshift spastin mutations revealed neither truncated protein nor mutated transcripts , providing evidence that these mutations are responsible for a loss of spastin function . identifying agents able to induce the expression of the non mutated spastin allele should represent an attractive therapeutic strategy in this disease .

The molecular genetics of non ALS motor neuron diseases. (2006 Nov)
The molecular genetics of non ALS motor neuron diseases . hereditary disorders of voluntary motor neurons are individually relatively uncommon , but have the potential to provide significant insights into motor neuron function in general and into the mechanisms underlying the more common form of sporadic amyotrophic lateral sclerosis . recently , mutations in a number of novel genes have been associated with lower motor neuron ( hspb1 , hspb8 , GARS , dynactin ) , upper motor neuron ( spastin , atlastin , paraplegin , hsp60 , kif5a , nipa1 ) or mixed ALS like phenotypes ( alsin , senataxin , VAPB , bscl2 ) . In comparison to sporadic ALS these conditions are usually associated with slow progression , but as experience increases , a wide variation in clinical phenotype has become apparent . At the molecular level common themes are emerging that point to areas of specific vulnerability for motor neurons such as axonal transport , endosomal trafficking and RNA processing . We review the clinical and molecular features of this diverse group of genetically determined conditions and consider the implications for the broad group of motor neuron diseases in general .

Two novel mutations in the spastin gene ( SPG4 ) found by dhplc mutation analysis. (2004 Oct)
Two novel mutations in the spastin gene ( SPG4 ) found by dhplc mutation analysis . The most common form of autosomal dominant hereditary spastic paraplegia is caused by mutations in the gene encoding spastin ( SPG4 ) , a member of the AAA family of atpases . In the current study , we designed a denaturing high performance liquid chromatography based protocol for the analysis of the SPG4 gene . using this method , we detected two novel missense mutations , 1375a G ( r459g ) and 1378c T ( r460c ) , one previously described five bases deletion ( 1215 1219del ) and three polymorphic changes . This study suggests that denaturing high performance liquid chromatography would be a fast and reliable tool in the investigation of the molecular defects in the SPG4 gene .

Interaction of two hereditary spastic paraplegia gene products , spastin and atlastin , suggests a common pathway for axonal maintenance. (2006 Jul)
interaction of two hereditary spastic paraplegia gene products , spastin and atlastin , suggests a common pathway for axonal maintenance . hereditary spastic paraplegia ( HSP ) is a neurodegenerative disorder that is characterized by retrograde axonal degeneration that primarily affects long spinal neurons . The disease is clinically heterogeneous , and there are 20 genetic loci identified . Here , we show a physical interaction between spastin and atlastin , two autosomal dominant HSP gene products . spastin encodes a microtubule ( MT ) severing AAA atpase ( atpase associated with various activities ) , and atlastin encodes a golgi localized integral membrane protein gtpase . atlastin does not regulate the enzymatic activity of spastin . We also identified a clinical mutation in atlastin outside of the gtpase domain that prevents interaction with spastin in cells . therefore , we hypothesize that failure of appropriate interaction between these two HSP gene products may be pathogenetically relevant . these data indicate that at least a subset of HSP genes may define a cellular biological pathway that is important in axonal maintenance .

Large deletion involving the 5 UTR in the spastin gene caused mild phenotype of autosomal dominant hereditary spastic paraplegia. (2005 Feb)
large deletion involving the 5 UTR in the spastin gene caused mild phenotype of autosomal dominant hereditary spastic paraplegia . hereditary spastic paraplegia ( HSP ) due to mutations in the spastin gene ( SPG4 ) located to 2p22 p21 is the most common form of autosomal dominant ( AD ) HSP . We performed PCR based direct sequencing of SPG4 , followed by a linkage analysis and subsequent southern blot analysis in large japanese kindred where 20 of 33 members were evaluated neurologically , and consequently 6 were affected with HSP . clinical evaluation showed that the mean age at disease onset of the patients was older and the disability was less severe than those of previously reported typical patients with SPG4 mutations . direct sequencing of genomic DNA and RT PCR product did not show a SPG4 mutation despite of a strong linkage to the SPG4 locus at 2p . southern blot analysis suggested a deletion involving the 5 UTR of SPG4 . further sequence analysis confirmed a heterozygous 2307 bp deletion spanning from the 5 UTR to intron 1 of SPG4 . The results suggested that transcription of the mutated allele starts from an authentic initiation site , but lacks an authentic translational start site of exon 1 because of a deficient splice donor site and coding region . The abnormal transcripts may result in rapid RNA decay . The novel refractory mutation we identified widens the spectrum of SPG4 mutations . these findings suggest that structural …

Hereditary spastic paraparesis : disrupted intracellular transport associated with spastin mutation. (2003 Dec)
hereditary spastic paraparesis : disrupted intracellular transport associated with spastin mutation . The commonest cause of hereditary spastic paraplegia ( HSP ) is mutation in the spastin gene . Both the normal function of spastin in the central nervous system and the mechanism by which mutation in spastin causes axonal degeneration are unknown . One hypothesis is that mutant spastin disrupts microtubule dynamics , causing an impairment of organelle transport on the microtubule network , which leads to degeneration in the distal parts of long axons . To study this neuronal and non neuronal cells were transfected with either wild type or mutant spastin proteins . We demonstrated evidence of a transient interaction of wild type spastin with microtubules , with resulting disassembly of microtubules , supporting a role for wild type spastin as a microtubule severing protein . mutant spastin demonstrated an abnormal interaction with microtubules , colocalizing with but no longer severing microtubules . The abnormal interaction of mutant spastin with microtubules was demonstrated to be associated with an abnormal perinuclear clustering of mitochondria and peroxisomes , suggestive of an impairment of kinesin mediated intracellular transport . Our findings indicate that an abnormal interaction of mutant spastin with microtubules , which disrupts organelle transport on the microtubule cytoskeleton , is likely to be the primary disease mechanism in HSP caused by missense mutations in the spastin gene .

The hereditary spastic paraplegia protein spastin interacts with the escrt III complex associated endosomal protein chmp1b. (2004 Dec)
The hereditary spastic paraplegia protein spastin interacts with the escrt III complex associated endosomal protein chmp1b . Pure hereditary spastic paraplegia is characterized by length dependent degeneration of the distal ends of long axons . mutations in spastin are the most common cause of the condition . We set out to investigate the function of spastin using a yeast two hybrid approach to identify interacting proteins . using full length spastin as bait , we identified chmp1b , a protein associated with the escrt ( endosomal sorting complex required for transport ) III complex , as a binding partner . several different approaches confirmed the physiological relevance of the interaction in mammalian cells . epitope tagged chmp1b and spastin showed clear cytoplasmic co localization in Cos 7 and PC12 cells . chmp1b and spastin interacted specifically in vitro and in vivo in beta lactamase protein fragment complementation assays , and spastin co immunoprecipitated with chmp1b . The interaction was mediated by a region of spastin lying between residues 80 and 196 and containing a microtubule interacting and trafficking domain . expression of epitope tagged chmp1b in mammalian cells prevented the development of the abnormal microtubule phenotype associated with expression of atpase defective spastin . these data point to a role for spastin in intracellular membrane traffic events and provide further evidence to support the emerging recognition that defects in intracellular membrane traffic are a significant cause of motor neuron pathology .

Spastin and microtubules : functions in health and disease. (2007 Sep)
spastin and microtubules : functions in health and disease . SPG4 , the gene encoding for spastin , a member of the atpases associated with various cellular activities ( AAA ) family , is mutated in around 40 of cases of autosomal dominant hereditary spastic paraplegia ( AD HSP ) . This group of neurodegenerative diseases is characterized by a progressive spasticity and lower limb weakness with degeneration of terminal axons in cortico spinal tracts and dorsal columns . spastin has two main domains , a microtubule interacting and endosomal trafficking ( MIT ) domain at the N terminus and the C terminus AAA domain . early studies suggested that spastin interacts with microtubules similarly to katanin , a member of the same subgroup of AAA . recent evidence confirmed that spastin possesses microtubule severing activity but can also bundle microtubules in vitro . understanding the physiologic and pathologic involvement of these activities and their regulation is critical in the study of HSP .

Identification and expression analysis of spastin gene mutations in hereditary spastic paraplegia. (2001 Apr)
identification and expression analysis of spastin gene mutations in hereditary spastic paraplegia . Pure hereditary spastic paraplegia ( SPG ) type 4 is the most common form of autosomal dominant hereditary SPG , a neurodegenerative disease characterized primarily by hyperreflexia and progressive spasticity of the lower limbs . It is caused by mutations in the gene encoding spastin , a member of the AAA family of atpases . We have screened the spastin gene for mutations in 15 families consistent with linkage to the spastin gene locus , SPG4 , and have identified 11 mutations , 10 of which are novel . Five of the mutations identified are in noninvariant splice junction sequences . reverse transcription PCR analysis of mRNA from patients shows that each of these five mutations results in aberrant splicing . One mutation was found to be leaky , or partially penetrant ; that is , the mutant allele produced both mutant ( skipped exon ) and wild type ( full length ) transcripts . This phenomenon was reproduced in in vitro splicing experiments , with a minigene splicing vector construct only in the context of the endogenous splice junctions flanking the splice junctions of the skipped exon . In the absence of endogenous splice junctions , only mutant transcript was detected . The existence of at least one leaky mutation suggests that relatively small differences in the level of wild type spastin expression can have significant functional consequences . This may account , at least in …

Spectrum of SPG4 mutations in a large collection of north american families with hereditary spastic paraplegia. (2002 Feb)
spectrum of SPG4 mutations in a large collection of north american families with hereditary spastic paraplegia . background : hereditary spastic paraplegia ( HSP ) is a neurodegenerative disease characterized by progressive spasticity and weakness of the lower limbs . The most common form of HSP is caused by mutations in the SPG4 gene , which codes for spastin , an adenosine triphosphatase with various cellular activities ( AAA ) protein family member . objective : To investigate a large collection of predominantly north american patients with HSP for mutations in the spastin encoding gene , SPG4 . methods : DNA from 76 unrelated affected individuals was studied for mutations by single stranded conformational polymorphism analysis and direct sequencing . Each new variant identified was then analyzed in 80 control subjects to determine whether the variant is a common polymorphism or a rare mutation . All DNA samples were amplified by polymerase chain reaction , followed by electrophoresis and autoradiography . results : We identified 8 novel mutations and 5 previously reported mutations in 15 affected individuals . The novel mutations are 4 missense , 1 nonsense , 1 frameshift , and 2 splice mutations . Two polymorphisms ( one in an affected individual ) were also identified . conclusions : Our collection of families with HSP is different on a genetic level from those previously described . The percentage of our families with a SPG4 mutation is 10 lower than the 40 estimate of families with autosomal dominant HSP …

Autosomal dominant hereditary spastic paraplegia : report of a large italian family with r581x spastin mutation. (2007 Aug)
autosomal dominant hereditary spastic paraplegia : report of a large italian family with r581x spastin mutation . We describe a large kindred with a typical pure form of autosomal dominant hereditary spastic paraplegia ( adhsp ) . On the basis of maximum LOD score of 1 . 94 at theta ( max ) 0 with marker d2s367 , we obtained suggestive evidence for linkage of adhsp to SPG4 locus . denaturing high performance liquid chromatography ( dhplc ) and direct sequence analysis allowed us to identify a nonsense mutation ( 1741 C T ) in exon 17 of the spastin gene . This transition , carried by all the affected family members and two apparently healthy individuals , lead to truncation of the last 36 amino acids in the C terminus of the protein . these results confirm the existence of mutation in the SPG4 gene with a reduced penetrance , indicating that other genetic or environmental factors are required to trigger full blown disease .

Spastic paraplegia caused by a novel mutation in the spastin gene ( 1207c G , p361r ) clinical features of … (2008 Feb)
spastic paraplegia caused by a novel mutation in the spastin gene ( 1207c G , p361r ) clinical features of a patient without family history A 52 year old man with no apparent family history of neurodegenerative diseases developed gait disturbance at age 47 . neurological examination at aged 52 revealed spastic paraplegia , generalized hyperreflexia , decreased of vibration sense in the lower limbs , and pollakisuria . ocular symptoms , deafness , cerebellar ataxia , extrapyramidal signs , mental deterioration , dementia , peripheral neuropathy , retinal pigment degeneration , ichthyosis and syndactyly were absent . MRI of the brain was normal . A pure form of hereditary spastic paraplegia was diagnosed . genetic analysis revealed a novel missense mutation in the spastin gene ( 1207c G , p361r ) . The clinical features of this patient were consistent with those of patient with the pure form of SPG4 . Gene analysis should be considered for patients with spastic paraplegia even in the absence of any family history .

Clinical signs and symptoms in a large hereditary spastic paraparesis pedigree with a novel spastin mutation. (2004 Jun)
clinical signs and symptoms in a large hereditary spastic paraparesis pedigree with a novel spastin mutation . The most common form of autosomal dominant hereditary spastic paraparesis ( HSP ) , SPG4 , is caused by mutations in the spastin gene on chromosome 2p . This disease is characterized by intra and interfamilial phenotypic variation . To determine the predictive values of clinical signs and symptoms in SPG4 , we examined 43 members of a large pedigree with autosomal dominant HSP . We then identified the genetic etiology of the disorder in this family , a novel nonsense mutation in exon 1 of spastin , carried by 24 of the examined family members . The best clinical predictors of positive gene status were the presence of hyperreflexia in the lower extremities , 2 beats of ankle clonus , pes cavus , bladder symptoms and increased tone in the legs . The mean age of onset was 32 . 2 / 7 . 4 years , but the age of onset was earlier in children from 10 of 12 child parent gene positive pairs , with a mean difference of 10 . 8 / 3 . 3 years . The finding of leg weakness was especially common in older onset affected family member with leg hyperreflexia . these results suggest that specific clinical signs and symptoms may be of value in differentiating individuals affected with SPG4 from family members with nonspecific neurological findings .

Novel spastin mutations and their expression analysis in two italian families. (2003 Aug)
novel spastin mutations and their expression analysis in two italian families . mutations in spastin cause the most common form of pure autosomal dominant hereditary spastic paraparesis ( SPG4 ) . Here , we report two italian families affected with SPG4 linked HSP harboring two novel spastin mutations . SSCP / sequencing analysis of the spastin gene showed a single base pair deletion causing a frame shift in one family ( 1442delt ) and a missense mutation ( 1726t C ) resulting in a leucine to proline amino acid change ( l534p ) in the other family . total RNA from the mutant and the wild type spastin allele in muscle biopsies from patients from the two affected families was quantitated . RNA expression was almost absent from the spastin allele harboring the single base pair deletion , while it was nearly normal for the spastin allele harboring the missense mutation . these data suggest that varying spastin RNA levels are found in out of frame and missense spastin mutations and imply different mechanisms involved in the molecular pathology of SPG4 linked HSP .

Structural characterization of the MIT domain from human vps4b. (2005 Jul)
structural characterization of the MIT domain from human vps4b . The microtubule interacting and trafficking ( MIT ) domain is a small protein module of unknown function that is conserved in proteins of diverse function , such as Vps4 , sorting nexin 15 ( snx15 ) , and spastin . One non synonymous single nucleotide polymorphism was reported , which results in a ile58 to Met ( I58M ) substitution in hvps4b . Here , we have determined the solution structure of the MIT domain isolated from the NH ( 2 ) terminus of human vps4b , an AAA atpase involved in multivesicular body formation . The MIT domain adopts an up and down three helix bundle . comparison with the sequences of other MIT domains clearly shows that the residues involved in inter helical contacts are well conserved . The ile58 to Met substitution resulted a substantial thermal instability . In addition , we found a shallow crevice between helices A and C that may serve as a protein binding site . We propose that the MIT domain serves as a putative adaptor domain for the escrt III complex involved in endosomal trafficking .