Ana I. Seixas

Loss of junctophilin-3 contributes to Huntington disease-like 2 pathogenesis.

Huntington disease‐like 2 (HDL2) is a progressive, late onset autosomal dominant neurodegenerative disorder, with remarkable similarities to Huntington disease (HD). HDL2 is caused by a CTG/CAG repeat expansion. In the CTG orientation, the repeat is located within the alternatively spliced exon 2A of junctophilin‐3 (JPH3), potentially encoding polyleucine and polyalanine, whereas on the strand antisense to JPH3, the repeat is in frame to encode polyglutamine. The JPH3 protein product serves to stabilize junctional membrane complexes and regulate neuronal calcium flux. We have previously demonstrated the potential pathogenic properties of JPH3 transcripts containing expanded CUG repeats. The aim of this study was to test the possibility that loss of JPH3 expression or expanded amino acid tracts also contribute to HDL2 pathogenesis.

'Costa da Morte' ataxia is spinocerebellar ataxia 36: clinical and genetic characterization

Spinocerebellar ataxia 36 has been recently described in Japanese families as a new type of spinocerebellar ataxia with motor neuron signs. It is caused by a GGCCTG repeat expansion in intron 1 of NOP56. Family interview and document research allowed us to reconstruct two extensive, multigenerational kindreds stemming from the same village (Costa da Morte in Galicia, Spain), in the 17th century. We found the presence of the spinocerebellar ataxia 36 mutation co-segregating with disease in these families in whom we had previously identified an ∼0.8 Mb linkage region to chromosome 20 p. Subsequent screening revealed the NOP56 expansion in eight additional Galician ataxia kindreds. While normal alleles contain 5–14 hexanucleotide repeats, expanded alleles range from ∼650 to 2500 repeats, within a shared haplotype. Further expansion of repeat size was frequent, especially upon paternal transmission, while instances of allele contraction were observed in maternal transmissions. We found a total of 63 individuals carrying the mutation, 44 of whom were confirmed to be clinically affected; over 400 people are at risk. We describe here the detailed clinical picture, consisting of a late-onset, slowly progressive cerebellar syndrome with variable eye movement abnormalities and sensorineural hearing loss. There were signs of denervation in the tongue, as well as mild pyramidal signs, but otherwise no signs of classical amyotrophic lateral sclerosis. Magnetic resonance imaging findings were consistent with the clinical course, showing atrophy of the cerebellar vermis in initial stages, later evolving to a pattern of olivo-ponto-cerebellar atrophy. We estimated the origin of the founder mutation in Galicia to have occurred ∼1275 years ago. Out of 160 Galician families with spinocerebellar ataxia, 10 (6.3%) were found to have spinocerebellar ataxia 36, while 15 (9.4%) showed other of the routinely tested dominant spinocerebellar ataxia types. Spinocerebellar ataxia 36 is thus, so far, the most frequent dominant spinocerebellar ataxia in this region, which may have implications for American countries associated with traditional Spanish emigration.

A novel H101Q mutation causes PKCγ loss in spinocerebellar ataxia type 14

AbstractSpinocerebellar ataxia type 14 (SCA14) is an autosomal dominant neurodegenerative disorder, first described in a Japanese family, showing linkage to chromosome 19q13.4-qter. Recently, mutations have been identified in the PRKCG gene in families with SCA14. The PRKCG gene encodes the protein kinase Cγ (PKCγ), a member of a serine/threonine kinase family involved in signal transduction important for several cellular processes, including cell proliferation and synaptic transmission. To identify the disease-causing mutation in a large group of ataxia patients, we searched for mutations in the PRKCG gene. We ascertained 366 unrelated patients with spinocerebellar ataxia, either pure or with associated features such as epilepsy, mental retardation, seizures, paraplegia, and tremor. A C-to-G transversion in exon 4, resulting in a histidine-to-glutamine change at codon 101 of the PKCγ protein, was identified in patients from a family with slowly progressive pure cerebellar ataxia. Functional studies performed in HEK293 cells transfected with normal or mutant construct showed that this mutation affects PKCγ stability or solubility, verified by time-dependent decreased protein levels in cell culture. In conclusion, the H101Q mutation causes slowly progressive uncomplicated ataxia by interfering with PKCγ stability or solubility, which consequently may cause in either case a decrease in the overall PKCγ-dependent phosphorylation.

FXTAS, SCA10, and SCA17 in american patients with movement disorders

The late-onset spinocerebellar ataxias (SCAs) are rare diseases characterized by selective neurodegeneration of one or more components of the cerebellum, often with involvement of other brain regions [Margolis, 2002; Schols et al., 2004]. Fragile X-associated tremor/ataxia syndrome (FXTAS), SCA10, and SCA17 are three recently described ataxia subtypes for which the prevalence among Americans with movement disorders has not been established. Trinucleotide (CGG) repeat expansions of>200 CGG triplets in the fragile X mental retardation 1 (FMR1) gene cause fragile X syndrome, the most common form of hereditary mental retardation [Hagerman and Hagerman, 2002]. FXTAS, characterized by progressive intention tremor, ataxia, and hyperintensities of the middle cerebellar peduncles on T2weighted MRIs, was recently reported in males older than 50 years carryingFMR1 premutation alleles of 50–200 triplets [Hagerman et al., 2001]. FXTAS was initially described in families with children affected with fragile X syndrome, and subsequent studies of similar families suggested that the penetrance of FXTAS is age-related and may be as high as 75% in male carriers aged 80 years or older [Jacquemont et al., 2004]. Five women with FMR1 premutation expansions and a milder form of FXTAS have recently been reported [Hagerman et al., 2004]. About 1 in 260 females and 1 in 813 males are carriers for the FMR1 premutation expansion [Dombrowski et al., 2002], suggesting that FXTAS might explain a substantial number of apparently sporadic cases of late-onset ataxia. The frequency of FXTAS in patients with movement disorders ranges from 0% to 4% in European [Macpherson et al., 2003; Zuhlke et al., 2004; Brussino et al., 2005; Van Esch et al., 2005] and Asian [Tan et al., 2004] populations. In the only American study to date, 1 of 167 men older than 50 referred for genetic testing carried a premutation [Milunsky and Maher, 2004]. Spinocerebellar ataxia type 10 (SCA10), thus far reported only in Mexican and Brazilian families, is characterized by cerebellar ataxia and seizures [Matsuura et al., 2000; Rasmussen et al., 2001; Teive et al., 2004]. SCA10 is caused by an expanded pentanucleotide (ATTCT) repeat in an intron of the SCA10 gene. The SCA10 repeat normally contains 10– 22 pentamers, while expanded alleles range in length from hundreds to thousands of pentanucleotides. Spinocerebellar ataxia type 17 (SCA17) is characterized by progressive cerebellar ataxia associated with dementia, though the phenotypic variability is quite extensive, with some cases resembling Parkinson disease or Huntington disease (HD). SCA17 is caused by a CAG expansion in the TBP gene [Koide et al., 1999; Nakamura et al., 2001]. Normal repeat length is 25–42 triplets, and the expanded length is 46–63 triplets, with intermediate alleles having an uncertain significance. To examine the frequency of FXTAS and SCAs 10 and 17 among Americans with movement disorders, we studied a sample of 233 unrelated individuals ascertained from patients referred to the Neurogenetics Test Laboratory at Johns Hopkins University for genetic testing for SCA or HD. Each patient provided informed consent for genetic testing and record review as approved by the Johns Hopkins University School of Medicine Institutional Review Board. Clinical information and family history were reviewed and cases with clinical or radiological evidence of basal ganglia or/ and cerebellar disease were included in the study (Table I). This group of patients is clinically heterogeneous, with mild to severe disease of varying duration, in most cases affecting the cerebellum. Individuals with a predominantly Parkinsonian presentation were excluded. All patients tested negative for the DRPLA; HD; HDL2; and SCA1, 2, 3, 6, 7, 8, and 12 mutations. Most cases were ascertained prior to the identification of FXTAS as a clinical syndrome. CCG repeat length in the FMR1 gene was detected by polymerase chain reaction (PCR) of 300 ng of genomic DNA from each patient using the GC-Rich PCR System (Roche, Indianapolis, IN) and previously described primers [Brown et al., 1993] modified for fluorescent detection, and automated genotyping (Applied Biosystems, Inc., Foster City, CA). For women in whom only a single allele was detected, Southern analysis [Garcia Arocena et al., 2003] was used to exclude the existence of an expanded allele. SCA10 expansions are typically too long to be detected by standard PCR. We therefore used the repeat-primed PCR method as previously described [Cagnoli et al., 2004]. An expansion appears as a smear in agarose gels and as a large series of discrete peaks on automatic genotyping, clearly distinguishable from normal alleles. The CAG repeat length in TBP, the gene associated with SCA17, was assessed using a standard PCR assay [Koide et al., 1999]. We analyzed 140 females and 93 males (Table I), with a mean age of 57.8 17.1 years. Approximately 46% of cases were sporadic, and 75% had evidence of cerebellar involvement. No SCA10 or SCA17 expansions were detected. Of the 140 women, 108 were heterozygous for normal length FMR1 repeats, and Southern analysis suggested the others were homozygotes. No premutation expansions were detected (95% CI: 0%, 1.3%), suggesting that FXTAS is uncommon in American women referred for genetic testing. Among men, 64 of whom were 50 years or older, one premutation expansion of approximately 130 CGGs was detected. Repeat length was confirmed by an outside laboratory (DNA Diagnostics Laboratory, Yale School of Medicine, New Haven, CT). The premutation carrier presented at age 50 with a 2-year history of right hand tremor, noticeable when he was writing or Grant sponsor: Portuguese Foundation for Science and Technology (FCT) and FEDER (to I.S.); Grant number: POCTI/MGI/ 34517/00; Grant sponsor: Luso-American Foundation (a scholarship to A.I.S.); Grant sponsor: National Ataxia Foundation and National Organization for Rare Disorders (to T.M.); Grant sponsor: NIH; Grant number: NS38054 and NS16375.

Haplotype diversity and somatic instability in normal and expanded SCA8 alleles

Spinocerebellar ataxia type 8 (SCA8) is an autosomal dominant late‐onset neurodegenerative disorder, belonging to the group of diseases caused by trinucleotide repeat expansions. SCA8 remains one of the most intriguing SCAs, regarding the reduced disease penetrance, and the high instability and poorly understood functional meaning of the (CTA)n(CTG)n expansion. We performed haplotype and sequencing analysis in a large region, encompassing the repeat, in four SCA8 and 20 control Portuguese families. The results from the haplotype study including the combined repeat and six SNP markers showed two different haplotypes, AG‐Exp‐GTTG and AG‐Exp‐CTTG, in the SCA8 families. Among the control population, these were also the most frequent, in a total of five haplotypes found unequally distributed throughout repeat sizes. From cloning fragments of control, unstable normal and expanded chromosomes, eleven different base substitutions were identified in exon A of the SCA8 gene. In some instances, somatic variability in repeat size or base composition was found for a same chromosome, regardless of its normal or expanded nature. In conclusion, our results in Portuguese families with ataxia show that SCA8 expansions arose in common backgrounds; in addition, this region seems to be unstable beyond the repeat.

Neuropathology and Cellular Pathogenesis of Spinocerebellar Ataxia Type 12.

SCA12 is a progressive autosomal‐dominant disorder, caused by a CAG/CTG repeat expansion in PPP2R2B on chromosome 5q32, and characterized by tremor, gait ataxia, hyperreflexia, dysmetria, abnormal eye movements, anxiety, depression, and sometimes cognitive impairment. Neuroimaging has demonstrated cerebellar and cortical atrophy. We now present the neuropathology of the first autopsied SCA12 brain and utilize cell models to characterize potential mechanisms of SCA12 neurodegeneration.

Jmy regulates oligodendrocyte differentiation via modulation of actin cytoskeleton dynamics

During central nervous system development, oligodendrocytes form structurally and functionally distinct actin‐rich protrusions that contact and wrap around axons to assemble myelin sheaths. Establishment of axonal contact is a limiting step in myelination that relies on the oligodendrocytes ability to locally coordinate cytoskeletal rearrangements with myelin production, under the control of a transcriptional differentiation program. The molecules that provide fine‐tuning of actin dynamics during oligodendrocyte differentiation and axon ensheathment remain largely unidentified. We performed transcriptomics analysis of soma and protrusion fractions from rat brain oligodendrocyte progenitors and found a subcellular enrichment of mRNAs in newly‐formed protrusions. Approximately 30% of protrusion‐enriched transcripts encode proteins related to cytoskeleton dynamics, including the junction mediating and regulatory protein Jmy, a multifunctional regulator of actin polymerization. Here, we show that expression of Jmy is upregulated during myelination and is required for the assembly of actin filaments and protrusion formation during oligodendrocyte differentiation. Quantitative morphodynamics analysis of live oligodendrocytes showed that differentiation is driven by a stereotypical actin network‐dependent “cellular shaping” program. Disruption of actin dynamics via knockdown of Jmy leads to a program fail resulting in oligodendrocytes that do not acquire an arborized morphology and are less efficient in contacting neurites and forming myelin wraps in co‐cultures with neurons. Our findings provide new mechanistic insight into the relationship between cell shape dynamics and differentiation in development.

Evolvability of the actin cytoskeleton in oligodendrocytes during central nervous system development and aging

The organization of actin filaments into a wide range of subcellular structures is a defining feature of cell shape and dynamics, important for tissue development and homeostasis. Nervous system function requires morphological and functional plasticity of neurons and glial cells, which is largely determined by the dynamic reorganization of the actin cytoskeleton in response to intrinsic and extracellular signals. Oligodendrocytes are specialized glia that extend multiple actin-based protrusions to form the multilayered myelin membrane that spirally wraps around axons, increasing conduction speed and promoting long-term axonal integrity. Myelination is a remarkable biological paradigm in development, and maintenance of myelin is essential for a healthy adult nervous system. In this review, we discuss how structure and dynamics of the actin cytoskeleton is a defining feature of myelinating oligodendrocytes’ biology and function. We also review “old and new” concepts to reflect on the potential role of the cytoskeleton in balancing life and death of myelin membranes and oligodendrocytes in the aging central nervous system.