João Vasconcelos

Increased transcript diversity: novel splicing variants of Machado–Joseph Disease gene (ATXN3)

Machado–Joseph disease (MJD) is a late-onset neurodegenerative disorder that presents clinical heterogeneity not completely explained by its causative mutation. MJD is caused by an expansion of a CAG tract at exon 10 of the ATXN3 gene (14q32.1), which encodes for ataxin-3. The main goal of this study was to analyze the occurrence of alternative splicing at the ATXN3 gene, by sequencing a total of 415 cDNAs clones (from 20 MJD patients and 14 controls). Two novel exons are described for the ATXN3 gene. Fifty-six alternative splicing variants, generated by four types of splicing events, were observed. From those variants, 50 were not previously described, and 26 were only found in MJD patients samples. Most of the variants (85.7%) present frameshift, which leads to the appearance of premature stop codons. Thirty-seven of the observed variants constitute good targets to nonsense-mediated decay, the remaining are likely to be translated into at least 20 different isoforms. The presence of ataxin-3 domains was assessed, and consequences of domain disruption are discussed. The present study demonstrates high variability in the ATXN3 gene transcripts, providing a basis for further investigation on the contribution of alternative splicing to the MJD pathogenic process, as well as to the larger group of the polyglutamine disorders.

The APOE ε2 Allele Increases the Risk of Earlier Age at Onset in Machado-Joseph Disease

OBJECTIVE To investigate a modulating effect of the apolipoprotein E (APOE) polymorphism on age at onset of Machado-Joseph disease (MJD). DESIGN We collected blood samples from 192 patients with MJD and typed the APOE polymorphism. Patients The 192 patients with MJD included 59 from the Azores, 73 from mainland Portugal, and 60 from Brazil. SETTING Academic research center. RESULTS Cases with the ε2/ε3 genotype had an earlier onset compared with those with the ε3/ε3 or the ε3/ε4 genotype. In this series of patients, the presence of an APOE ε2 allele implies a decrease of nearly 5 years in the age at onset. When combining several other predictors in a general linear model, namely, the presence/absence of the APOE ε2 allele, with the size of the (CAG)(n) in expanded alleles, the model was significantly improved and the explanation of onset variance was raised from 59.8% to 66.5%. Furthermore, the presence of the ε2 allele was associated with an onset before age 39 years (odds ratio, 5.00; 95% CI, 1.18-21.14). CONCLUSION The polymorphism at the APOE gene plays a role as a genetic modifier of MJD phenotype.

Analysis of segregation patterns in Machado–Joseph disease pedigrees

AbstractMachado-Joseph disease (MJD), also known as spinocerebellar ataxia type 3 (SCA3), is an autosomal dominant neurodegenerative disorder of late onset, which is considered the most common form of SCA worldwide. The main goal of this study was to investigate the presence of segregation ratio distortion (SRD) during transmissions of ATXN3 alleles by MJD patients, evaluating the putative role of SRD in the epidemiological representation of the disease. Sixty-two complete sibships, each with one clinically affected parent, totalling 330 transmissions were selected according to defined criteria and used for segregation analysis. Onset data from MJD patients with Azorean origin was used for residual risk estimates according to different ages. Residual risk values were applied to unaffected offspring to calculate the probability of inheriting the expanded allele. The proportion of offspring that received the expanded or the normal allele from the affected parent was calculated to determine the presence of SRD during transmissions of ATXN3 alleles by MJD patients. Segregation of ATXN3 alleles was in accordance with the expected Mendelian proportions (χ2 = 0.982, P = 0.322). However, there was a tendency favouring the transmission of the normal alleles. Thus, SRD is not a potential mechanism on the basis of MJD epidemiological representation.

Parkinsonian phenotype in Machado-Joseph disease (MJD/SCA3): a two-case report

BackgroundMachado-Joseph disease (MJD), or spinocerebellar ataxia type 3 (SCA3), is an autosomal dominant neurodegenerative disorder of late onset, which is caused by a CAG repeat expansion in the coding region of the ATXN3 gene. This disease presents clinical heterogeneity, which cannot be completely explained by the size of the repeat tract. MJD presents extrapyramidal motor signs, namely Parkinsonism, more frequently than the other subtypes of autosomal dominant cerebellar ataxias. Although Parkinsonism seems to segregate within MJD families, only a few MJD patients develop parkinsonian features and, therefore, the clinical and genetic aspects of these rare presentations remain poorly investigated. The main goal of this work was to describe two MJD patients displaying the parkinsonian triad (tremor, bradykinesia and rigidity), namely on what concerns genetic variation in Parkinsons disease (PD) associated loci (PARK2, LRRK2, PINK1, DJ-1, SNCA, MAPT, APOE, and mtDNA tRNAGlnT4336C).Case presentationPatient 1 is a 40 year-old female (onset at 30 years of age), initially with a pure parkinsonian phenotype (similar to the phenotype previously reported for her mother). Patient 2 is a 38 year-old male (onset at 33 years of age), presenting an ataxic phenotype with parkinsonian features (not seen either in other affected siblings or in his father). Both patients presented an expanded ATXN3 allele with 72 CAG repeats. No PD mutations were found in the analyzed loci. However, allelic variants previously associated with PD were observed in DJ-1 and APOE genes, for both patients.ConclusionsThe present report adds clinical and genetic information on this particular and rare MJD presentation, and raises the hypothesis that DJ-1 and APOE polymorphisms may confer susceptibility to the parkinsonian phenotype in MJD.

Novel candidate blood-based transcriptional biomarkers of Machado-Joseph disease.

Machado‐Joseph disease (or spinocerebellar ataxia type 3) is a late‐onset polyglutamine neurodegenerative disorder caused by a mutation in the ATXN3 gene, which encodes for the ubiquitously expressed protein ataxin‐3. Previous studies on cell and animal models have suggested that mutated ataxin‐3 is involved in transcriptional dysregulation. Starting with a whole‐transcriptome profiling of peripheral blood samples from patients and controls, we aimed to confirm abnormal expression profiles in Machado‐Joseph disease and to identify promising up‐regulated genes as potential candidate biomarkers of disease status.

Nystagmus as an early ocular alteration in Machado-Joseph disease (MJD/SCA3)

BackgroundMachado-Joseph disease (MJD), also named spinocerebellar ataxia type 3 (SCA3) is the most common autosomal dominant ataxia worldwide. Although nystagmus is one of the most frequently reported ocular alterations in MJD patients its behaviour during the course of the disease, namely in its early stages, has only recently started to be investigated. The main goal of this work was to characterize the frequency of nystagmus in symptomatic and presymptomatic carriers of the MJD mutation, and investigate its usefulness as an early indicator of the disease.MethodsWe conducted an observational study of Azorean MJD family members, comprising a total of 158 subjects which underwent neurological evaluation. Sixty eight were clinically and molecularly diagnosed with MJD, 48 were confirmed asymptomatic carriers and 42 were confirmed non-carriers of the MJD mutation. The frequency of nystagmus was calculated for the 3 groups.ResultsNystagmus was present in 88% of the MJD patients. Seventeen percent of the at-risk subjects with a carrier result in the molecular test and none of the 42 individuals who received a non-carrier test result displayed nystagmus (p < 0.006). Although not reaching statistical significance, symptomatic subjects showing nystagmus had a tendency for a higher length of the CAG tract in the expanded allele, when compared to individuals who did not have nystagmus.ConclusionsThe frequency of nystagmus in asymptomatic carriers and its absence in non-carriers of the mutation, suggests that nystagmus may appear before gait disturbance and can thus be considered an early sign of MJD.

The (CAG) n tract of Machado-Joseph Disease gene (ATXN3): a comparison between DNA and mRNA in patients and controls

Machado–Joseph disease (MJD) is an autosomal dominant neurodegenerative disorder of late onset (occurring at a mean age of 40.2 years). The clinical manifestation of MJD is dependent on the presence of an expansion of the (CAG)n motif within exon 10 of the ATXN3 gene, located at 14q32.1. The variance in onset of MJD is only partially correlated (∼50–80%) with the extension of the CAG tract in genomic DNA (gDNA). The main aim of this work was to determine whether there are discrepancies in the size of the (CAG)n tract between gDNA and mRNA, and to establish whether there is a better association between age at onset and repeat size at the mRNA level. We typed gDNA and cDNA samples for the (CAG)n tract totalizing 108 wild-type and 52 expanded ATXN3 alleles. In wild-type alleles no differences were found between gDNA and cDNA. In expanded alleles, the CAG repeat size in gDNA was not always directly transcribed into the mRNA; on average there were differences of +1 repeat at the cDNA level. The slight discrepancies obtained were insufficient to cause significant differences in the distribution of the expanded alleles, and therefore no improvement in onset variance explanation was obtained with mRNA.

Psychological well-being and family satisfaction levels five years after being confirmed as a carrier of the Machado-Joseph disease mutation.

The present study on long-term outcome of presymptomatic testing for Machado-Joseph disease (MJD) aimed to evaluate the psychological well-being and the familial satisfaction of subjects that 5 years prior received an unfavorable result in the predictive testing (PT). The study included 47 testees of Azorean origin (23 from the island of Flores and 24 from S. Miguel) that completed the fourth evaluation session of the MJD protocol, and undertook a neurological examination at the moment of participation in the study. Nearly 50% of testees were symptomatic at the time of the study. Psychological well-being of the 47 participants was evaluated using the Psychological General Well-Being Index (PGWB). The family satisfaction scale by adjectives was applied to obtain information on family dynamics. The average PGWB score of the total participants was of 73.3, a value indicative of psychological well-being. Nearly half of the testees presented scores indicating psychological well-being, whereas scores indicating moderate (28.9%) or severe (23.7%) stress were found in the remaining. The average score in the PGWB scale was lower in symptomatic than in asymptomatic subjects; moreover, the distinct distribution of the well-being categories seen in the two groups shows an impact of the appearance of first symptoms on the psychological state. Motives for undertaking the test, provided 5 years prior, failed to show an impact in well-being. The average score for familial satisfaction was of 134, a value compatible with high familial satisfaction, which represented the most frequent category (59.6%). Results demonstrate that well-being and family satisfaction need to be monitored in confirmed carriers of the MJD mutation. The inclusion of acceptance studies, after PT, as well as the development of acceptance training actions, should be of major importance to anticipate the possibility of psychological damage.