DNAJB6, a Key Factor in Neuronal Sensitivity to Amyloidogenesis.

CAG-repeat expansions in at least eight different genes cause neurodegeneration. The length of the extended polyglutamine stretches in the corresponding proteins is proportionally related to their aggregation propensity. Although these proteins are ubiquitously expressed, they predominantly cause toxicity to neurons. To understand this neuronal hypersensitivity, we generated induced pluripotent stem cell (iPSC) lines of spinocerebellar ataxia type 3 and Huntington's disease patients. iPSC generation and neuronal differentiation are unaffected by polyglutamine proteins and show no spontaneous aggregate formation. However, upon glutamate treatment, aggregates form in neurons but not in patient-derived neural progenitors. During differentiation, the chaperone network is drastically rewired, including loss of expression of the anti-amyloidogenic chaperone DNAJB6. Upregulation of DNAJB6 in neurons antagonizes glutamate-induced aggregation, while knockdown of DNAJB6 in progenitors results in spontaneous polyglutamine aggregation. Loss of DNAJB6 expression upon differentiation is confirmed in vivo, explaining why stem cells are intrinsically protected against amyloidogenesis and protein aggregates are dominantly present in neurons.

The S/T-Rich Motif in the DNAJB6 Chaperone Delays Polyglutamine Aggregation and the Onset of Disease in a Mouse Model.

Expanded CAG repeats lead to debilitating neurodegenerative disorders characterized by aggregation of proteins with expanded polyglutamine (polyQ) tracts. The mechanism of aggregation involves primary and secondary nucleation steps. We show how a noncanonical member of the DNAJ-chaperone family, DNAJB6, inhibits the conversion of soluble polyQ peptides into amyloid fibrils, in particular by suppressing primary nucleation. This inhibition is mediated by a serine/threonine-rich region that provides an array of surface-exposed hydroxyl groups that bind to polyQ peptides and may disrupt the formation of the H bonds essential for the stability of amyloid fibrils. Early prevention of polyQ aggregation by DNAJB6 occurs also in cells and leads to delayed neurite retraction even before aggregates are visible. In a mouse model, brain-specific coexpression of DNAJB6 delays polyQ aggregation, relieves symptoms, and prolongs lifespan, pointing to DNAJB6 as a potential target for disease therapy and tool for unraveling early events in the onset of polyQ diseases.

ABC-transporter CFTR folds with high fidelity through a modular, stepwise pathway.

The question how proteins fold is especially pointed for large multi-domain, multi-spanning membrane proteins with complex topologies. We have uncovered the sequence of events that encompass proper folding of the ABC transporter CFTR in live cells by combining kinetic radiolabeling with protease-susceptibility assays. We found that CFTR folds in two clearly distinct stages. The first, co-translational, stage involves folding of the 2 transmembrane domains TMD1 and TMD2, plus one nucleotide-binding domain, NBD1. The second stage is a simultaneous, post-translational increase in protease resistance for both TMDs and NBD2, caused by assembly of these domains onto NBD1. Our assays probe every 2-3 residues (on average) in CFTR. This in-depth analysis at amino-acid level allows detailed analysis of domain folding and importantly also the next level: assembly of the domains into native, folded CFTR. Defects and changes brought about by medicines, chaperones, or mutations also are amenable to analysis. We here show that the well-known disease-causing mutation F508del, which established cystic fibrosis as protein-folding disease, caused co-translational misfolding of NBD1 but not TMD1 nor TMD2 in stage 1, leading to absence of stage-2 folding. Corrector drugs rescued stage 2 without rescuing NBD1. Likewise, the DxD motif in NBD1 that was identified to be required for export of CFTR from the ER we found to be required already upstream of export as CFTR mutated in this motif phenocopies F508del CFTR. The highly modular and stepwise folding process of such a large, complex protein explains the relatively high fidelity and correctability of its folding.

Transmembrane Helices 7 and 8 Confer Aggregation Sensitivity to the Cystic Fibrosis Transmembrane Conductance Regulator.

The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is a large multi-spanning membrane protein that is susceptible to misfolding and aggregation. We have identified here the region responsible for this instability. Temperature-induced aggregation of C-terminally truncated versions of CFTR demonstrated that all truncations up to the second transmembrane domain (TMD2), including the R region, largely resisted aggregation. Limited proteolysis identified a folded structure that was prone to aggregation and consisted of TMD2 and at least part of the Regulatory Region R. Only when both TM7 (TransMembrane helix 7) and TM8 were present, TMD2 fragments became as aggregation-sensitive as wild-type CFTR, in line with increased thermo-instability of late CFTR nascent chains and in silico prediction of aggregation propensity. In accord, isolated TMD2 was degraded faster in cells than isolated TMD1. We conclude that TMD2 extended at its N-terminus with part of the R region forms a protease-resistant structure that induces heat instability in CFTR and may be responsible for its limited intracellular stability.

CAG Repeat Size Influences the Progression Rate of Spinocerebellar Ataxia Type 3.

OBJECTIVE: In spinocerebellar ataxia type 3/Machado-Joseph disease (SCA3/MJD), the expanded cytosine adenine guanine (CAG) repeat in ATXN3 is the causal mutation, and its length is the main factor in determining the age at onset (AO) of clinical symptoms. However, the contribution of the expanded CAG repeat length to the rate of disease progression after onset has remained a matter of debate, even though an understanding of this factor is crucial for experimental data on disease modifiers and their translation to clinical trials and their design. METHODS: Eighty-two Dutch patients with SCA3/MJD were evaluated annually for 15 years using the International Cooperative Ataxia Rating Scale (ICARS). Using linear growth curve models, ICARS progression rates were calculated and tested for their relation to the length of the CAG repeat expansion and to the residual age at onset (RAO): The difference between the observed AO and the AO predicted on the basis of the CAG repeat length. RESULTS: On average, ICARS scores increased 2.57 points/year of disease. The length of the CAG repeat was positively correlated with a more rapid ICARS progression, explaining 30% of the differences between patients. Combining both the length of the CAG repeat and RAO as comodifiers explained up to 47% of the interpatient variation in ICARS progression. INTERPRETATION: Our data imply that the length of the expanded CAG repeat in ATXN3 is a major determinant of clinical decline, which suggests that CAG-dependent molecular mechanisms similar to those responsible for disease onset also contribute to the rate of disease progression in SCA3/MJD. ANN NEUROL 2021;89:66-73.

The effect of seed-dispersal timing on seedling recruitment is modulated by environmental conditions that vary across altitude in a threatened palm.

BACKGROUND AND AIMS: The timing of seed dispersal determines the environmental conditions that plants face during early life stages. In seasonal environments, selection is expected to favour dispersal timing that is matched to environmental conditions suitable for successful recruitment. Our aim here was to test whether the timing of seed dispersal influences seedling establishment success in two populations of Euterpe edulis that are located at contrasting altitudes, have different seed-dispersal phenologies and are subjected to distinct climatic conditions. METHODS: We sowed E. edulis seeds in contrasting altitudes on different dates, and monitored seed germination, emergence and seedling establishment at each altitude over 4 years. At the high-altitude site, five seed-dispersal cohorts were established during the natural dispersal period. At the low-altitude site, three seed-dispersal cohorts were established during natural dispersal, and two were established either before or after natural dispersal. KEY RESULTS: At the high-altitude site, seed-dispersal timing did not affect seed germination, seedling emergence or seedling establishment success. In contrast, at the low-altitude site, late seed dispersal near the end of the wet season resulted in a lower probability of seedling establishment, possibly due to the exposure of seeds, germinants and seedlings to unfavourable drought conditions. In addition, at the low-altitude site, the natural seed-dispersal period was poorly matched to favourable environmental conditions for seedling establishment. CONCLUSIONS: The greater effect of seed-dispersal timing on seedling establishment at the low-altitude site is probably related to a more seasonal and drought-prone environment that favours a restricted period of seed dispersal. The magnitude of the effect of dispersal timing on seedling establishment success was modulated by environmental conditions that vary across altitude. Furthermore, reproductive phenology appears to be subject to more intense selection at the lower limit of the altitudinal range, due to a more restrictive window of opportunity for successful seedling establishment.

Alternative plant designs: consequences for community assembly and ecosystem functioning.

BACKGROUND: Alternative organism designs (i.e. the existence of distinct combinations of traits leading to the same function or performance) are a widespread phenomenon in nature and are considered an important mechanism driving the evolution and maintenance of species trait diversity. However, alternative designs are rarely considered when investigating assembly rules and species effects on ecosystem functioning, assuming that single trait trade-offs linearly affect species fitness and niche differentiation. SCOPE: Here, we first review the concept of alternative designs, and the empirical evidence in plants indicating the importance of the complex effects of multiple traits on fitness. We then discuss how the potential decoupling of single traits from performance and function of species can compromise our ability to detect the mechanisms responsible for species coexistence and the effects of species on ecosystems. Placing traits in the continuum of organism integration level (i.e. traits hierarchically structured ranging from organ-level traits to whole-organism traits) can help in choosing traits more directly related to performance and function. CONCLUSIONS: We conclude that alternative designs have important implications for the resulting trait patterning expected from different assembly processes. For instance, when only single trade-offs are considered, environmental filtering is expected to result in decreased functional diversity. Alternatively, it may result in increased functional diversity as an outcome of alternative strategies providing different solutions to local conditions and thus supporting coexistence. Additionally, alternative designs can result in higher stability of ecosystem functioning as species filtering due to environmental changes would not result in directional changes in (effect) trait values. Assessing the combined effects of multiple plant traits and their implications for plant functioning and functions will improve our mechanistic inferences about the functional significance of community trait patterning.

Genetic risk factors for modulation of age at onset in Machado-Joseph disease/spinocerebellar ataxia type 3: a systematic review and meta-analysis.

OBJECTIVES: To perform a systematic review and meta-analysis of genetic risk factors for age at onset (AO) in spinocerebellar ataxia type 3/Machado-Joseph disease (SCA3/MJD). METHODS: Two authors independently reviewed reports on the mathematical relationship between CAG length at the expanded ATXN3 allele (CAGexp), and other genetic variants if available, and AO. Publications from January 1994 to September 2017 in English, Portuguese or Spanish and indexed in MEDLINE (PubMed), LILACS or EMBASE were considered. Inclusion criteria were reports with >20 SCA3/MJD carriers with molecular diagnosis performed by capillary electrophoresis. Non-overlapping cohorts were determined on contact with corresponding authors. A detailed analysis protocol was registered at the PROSPERO database prior to data extraction (CRD42017073071). RESULTS: Eleven studies were eligible for meta-analysis, comprising 10 individual-participant (n=2099 subjects) and two aggregated data cohorts. On average, CAGexp explained 55.2% (95% CI 50.8 to 59.0; p<0.001) of AO variability. Population-specific factors accounted for 8.3% of AO variance. Cohorts clustered into distinct geographic groups, evidencing significantly earlier AO in non-Portuguese Europeans than in Portuguese/South Brazilians with similar CAGexp lengths. Presence of intermediate ATXN2 alleles (27-33 CAG repeats) significantly correlated with earlier AO. Familial factors accounted for ~10% of AO variability. CAGexp, origin, family effects and CAG length at ATXN2 together explained 73.5% of AO variance. CONCLUSIONS: Current evidence supports genetic modulation of AO in SCA3/MJD by CAGexp, ATXN2 and family-specific and population-specific factors. Future studies should take these into account in the search for new genetic modifiers of AO, which could be of therapeutic relevance.

Ophthalmological and Neurologic Manifestations in Pre-clinical and Clinical Phases of Spinocerebellar Ataxia Type 7.

Spinocerebellar ataxia type 7 (SCA7) is a polyglutamine disease that progressively affects the cerebellum, brainstem, and retina. SCA7 is quite rare, and insights into biomarkers and pre-clinical phases are still missing. We aimed to describe neurologic and ophthalmological findings observed in symptomatic and pre-symptomatic SCA7 subjects. Several neurologic scales, visual acuity, visual fields obtained by computer perimetry, and macular thickness in optical coherence tomography (mOCT) were measured in symptomatic carriers and at risk relatives. Molecular analysis of the ATXN7 was done blindly in individuals at risk. Thirteen symptomatic carriers, 3 pre-symptomatic subjects, and 5 related controls were enrolled. Symptomatic carriers presented scores significantly different from those of controls in most neurologic and ophthalmological scores. Gradual changes from controls to pre-symptomatic and then to symptomatic carriers were seen in mean (SD) of visual fields - 1.34 (1.15), - 2.81 (1.66). and - 9.56 (7.26); mOCT - 1.11 (2.6), - 3.48 (3.54), and - 7.73 (2.56) Z scores; and "Spinocerebellar Ataxia Functional Index (SCAFI)" - 1.16 (0.28), 0.65 (0.56), and - 0.61 (0.44), respectively. Visual fields and SCAFI were significantly correlated with time to disease onset (pre-symptomatic)/disease duration (symptomatic carriers). Visual fields, mOCT, and SCAFI stood out as candidates for state biomarkers for SCA7 since pre-symptomatic stages of disease.

Synthesis and preclinical evaluation of [18F]AlF-NODA-MP-C6-CTHRSSVVC as a PET tracer for CD163-positive tumor-infiltrating macrophages.

Positron emission tomography (PET) can provide information about tumor-associated macrophage (TAM) infiltration, as long as a suitable tracer is available. This study aimed to evaluate the radiolabeled peptide [18F]AlF-NODA-MP-C6-CTHRSSVVC as a potential PET tracer for imaging of the CD163 receptor, which is expressed on M2-type tumor-associated macrophages. The conjugated peptide NODA-MP-C6-CTHRSSVVC was labeled with aluminum [18F]fluoride. Tracer binding and its biodistribution were evaluated in an in vitro binding assay and in healthy BALB/c mice, respectively. In addition, different treatments with cyclophosphamide in tumor-bearing mice were used to assess whether the tracer could detect differences in CD163 expression caused by differential TAM infiltration. After 7 days of treatment, animals were injected with [18F]AlF-NODA-MP-C6-CTHRSSVVC, and a 60-min dynamic PET scan was performed, followed by an ex vivo biodistribution study. [18F]AlF-NODA-MP-C6-CTHRSSVVC was prepared in 23 ± 6 % radiochemical yield and showed approximately 50 % of specific receptor-mediated binding in an in vitro binding assay on human CD163-expressing tissue homogenates. No CD163-mediated binding of [18F]AlF-NODA-MP-C6-CTHRSSVVC was detected by PET under normal physiological conditions in healthy BALB/c mice. On the other hand, CD163-positive xenograft tumors were clearly visualized with PET and a positive correlation was found between CD163 levels and the [18F]AlF-NODA-MP-C6-CTHRSSVVC tumor-to-muscle ratio (TMR) obtained from the PET images (Pearson r = 0.76, p = 0.002). No significant differences in the CD163 protein level and in the tracer uptake between treatment groups were found in the tumors. Taken together, [18F]AlF-NODA-MP-C6-CTHRSSVVC appears a promising candidate PET tracer for M2-type TAM, as it binds specifically to CD163 in vitro and its tumor uptake correlates well with CD163 expression in vivo.

Biogenesis of [Fe-S] cluster in Firmicutes: an unexploited field of investigation.

Iron-sulfur clusters (ISC) ([Fe-S]) are evolutionarily ancient and ubiquitous inorganic prosthetic groups present in almost all living organisms, whose biosynthetic assembly is dependent on complex protein machineries. [Fe-S] clusters are involved in biologically important processes, ranging from electron transfer catalysis to transcriptional regulatory roles. Three different systems involved in [Fe-S] cluster assembly have already been characterized in Proteobacteria, namely, the nitrogen fixation system, the ISC system and the sulfur assimilation system. Although they are well described in various microorganisms, these machineries are poorly characterized in members of the Firmicutes phylum, to which several groups of pathogenic bacteria belong. Recently, several research groups have made efforts to elucidate the biogenesis of [Fe-S] clusters at the molecular level in Firmicutes, and many important characteristics have been described. Considering the pivotal role of [Fe-S] clusters in a number of biological processes, the review presented here focuses on the description of the biosynthetic machineries for [Fe-S] cluster biogenesis in prokaryotes, followed by a discussion on recent results observed for Firmicutes [Fe-S] cluster assembly.

Are leaf physiological traits related to leaf water isotopic enrichment in restinga woody species?

During plant-transpiration, water molecules having the lighter stable isotopes of oxygen and hydrogen evaporate and diffuse at a faster rate through the stomata than molecules having the heavier isotopes, which cause isotopic enrichment of leaf water. Although previous models have assumed that leaf water is well-mixed and isotopically uniform, non-uniform stomatal closure, promoting different enrichments between cells, and different pools of water within leaves, due to morpho-physiological traits, might lead to inaccuracies in isotopic models predicting leaf water enrichment. We evaluate the role of leaf morpho-physiological traits on leaf water isotopic enrichment in woody species occurring in a coastal vegetation of Brazil known as restinga. Hydrogen and oxygen stable isotope values of soil, plant stem and leaf water and leaf traits were measured in six species from restinga vegetation during a drought and a wet period. Leaf water isotopic enrichment relative to stem water was more homogeneous among species during the drought in contrast to the wet period suggesting convergent responses to deal to temporal heterogeneity in water availability. Average leaf water isotopic enrichment relative to stem water during the drought period was highly correlated with relative apoplastic water content. We discuss this observation in the context of current models of leaf water isotopic enrichment as a function of the Péclet effect. We suggest that future studies should include relative apoplastic water content in isotopic models.

Randomized Noninferiority Trial of Radiation Exposure During Coronary Angiography: the Transradial and Transfemoral Approach by EXPERienced Operators in Daily rouTine (EXPERT) Trial.

BACKGROUND: The transradial approach (TRA) to coronary angiography reduces vascular complications but is associated with greater radiation exposure than the transfemoral approach (TFA). It is unknown whether exposure remains higher when TRA is performed by experienced operators. METHODS: Patients were randomly, prospectively assigned to TRA or TFA. The primary end point was patient radiation dose; secondary end points were the physician radiation dose and 30-day major adverse cardiac event rate. Coronary angiography was performed by experienced operators using a standardized protocol. RESULTS: Clinical and procedural characteristics were similar between the TRA (n = 150) and TFA (n = 149) groups, and they had comparable mean (SD) radiation doses for patients (616.51 [252] vs 585.57 [225] mGy; P = .13) and physicians (0.49 [0.3] vs 0.46 [0.29] mSv; P = .32). The mean (SD) fluoroscopy time (3.52 [2.02] vs 3.13 [2.46] min; P = .14) and the mean (SD) dose area product (35,496.5 [15,670] vs 38,313.4 [17,764.9] mGy·cm2; P = .2) did not differ. None of the following factors predicted higher radiation doses: female sex (hazard ratio [HR], 0.69 [95% CI, 0.38-1.3]; P = .34), body mass index >25 (HR, 0.84 [95% CI, 0.43-1.6]; P = .76), age >65 years (HR, 1.67 [95% CI, 0.89-3.1]; P = .11), severe valve disease (HR, 1.37 [95% CI, 0.52-3.5]; P = .68), or previous coronary artery bypass graft (HR, 0.6; 95% CI, 0.2-1.8; P = .38). CONCLUSION: TRA for elective coronary angiography is noninferior to TFA when performed by experienced operators.

Variation in DNA Repair System Gene as an Additional Modifier of Age at Onset in Spinocerebellar Ataxia Type 3/Machado-Joseph Disease.

Spinocerebellar ataxia type 3, or Machado-Joseph disease (SCA3/MJD), is caused by an expansion of CAG repeats, which is inversely correlated to age at onset (AO) of symptoms. However, on average, just 55.2% of variation in AO can be explained by expansion length. Additional modulators, such as polymorphic CAG tract in ATXN2 gene, can raise to 63.0% of the variation in AO. A sequence variation (rs3512) in FAN1 gene has previously been shown to be associated with late AO in Huntington's disease and polyglutaminopathies associated to ataxia. In the present study, genotype frequency of rs3512 was demonstrated in a cohort of SCA3/MJD patients from South Brazil, and these data were correlated to AO. The disease started 2.44 years earlier in subjects with the G/G genotype when compared to those subjects carrying the same CAGexp length at the ATXN3 gene and other genotypes (C/G and C/C) at rs3512. Placing together data on rs3512 genotype with data on CAG tract in ATXN2, AO of patients with G/G genotype was 2.58 years earlier, and a delay of 4.25 years was observed in patients that carry a short ATXN2 allele. Data presented here add further insights on the contribution of other factors in AO of SCA3/MJD beyond the causal mutation. Thus, well-known modifiers can help to unveil new ones and, as a whole, to better elucidate the mechanisms behind disease onset.

Protein Quality Control Pathways at the Crossroad of Synucleinopathies.

The pathophysiology of Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, and many others converge at alpha-synuclein (α-Syn) aggregation. Although it is still not entirely clear what precise biophysical processes act as triggers, cumulative evidence points towards a crucial role for protein quality control (PQC) systems in modulating α-Syn aggregation and toxicity. These encompass distinct cellular strategies that tightly balance protein production, stability, and degradation, ultimately regulating α-Syn levels. Here, we review the main aspects of α-Syn biology, focusing on the cellular PQC components that are at the heart of recognizing and disposing toxic, aggregate-prone α-Syn assemblies: molecular chaperones and the ubiquitin-proteasome system and autophagy-lysosome pathway, respectively. A deeper understanding of these basic protein homeostasis mechanisms might contribute to the development of new therapeutic strategies envisioning the prevention and/or enhanced degradation of α-Syn aggregates.

Strong quenching of chlorophyll fluorescence in the desiccated state in three poikilohydric and homoiochlorophyllous moss species indicates photo-oxidative protection on highly light-exposed rocks of a tropical inselberg.

The three poikilohydric and homoiochlorophyllous moss species Campylopus savannarum (C. Muell.) Mitt., Racocarpus fontinaloides (C. Muell.) Par. and Ptychomitrium vaginatum Besch. grow on sun-exposed rocks of a tropical inselberg in Brazil subject to regular drying and wetting cycles. Effective photo-oxidative protection in the light-adapted desiccated state in all three species is achieved by a reduction of ground chlorophyll fluorescence, F', to almost zero. Upon rewatering, the kinetics of the recovery of F' in air dry cushions to higher values is very fast in the first 5 min, but more than 80 min are needed until an equilibrium is reached gradually. The kinetics were not different between the three species. The three moss species, have a distinct niche occupation and form a characteristic zonation around soil vegetation islands on the rock outcrops, where C. savannarum and R. fontinaloides form an inner and outer belt, respectively, around vegetation islands and P. vaginatum occurs as small isolated cushions on bare rock. However, they were not distinguished by the reduction of F' in the dry state and the rewetting recovery kinetics and only slightly different in their photosynthetic capacity. Stable isotope ratios (delta(13)C, delta(15)N) indicate that liquid films of water limiting diffusion of CO(2) are important in determining carbon acquisition and suggest that limitation of CO(2) fixation by water films must be more pronounced over time in P. vaginatum than in the latter species. This is determined by both the micro site occupied and the form of the moss cushions.

Demographic processes limit upward altitudinal range expansion in a threatened tropical palm.

Understanding the factors that determine species' range limits is a key issue in ecology, and is fundamental for biodiversity conservation under widespread global environmental change. Elucidating how altitudinal variation affects demographic processes may provide important clues for understanding the factors limiting current and future species distributions, yet population dynamics at range limits are still poorly understood. Here, we tested the hypothesis that lower abundance at a species' upper altitudinal range limit is related to lower vital rates. We compared the dynamics of two populations of the tropical palm Euterpe edulis, located near and at the edge of its altitudinal limit of distribution in the Brazilian Atlantic Forest. Data from four annual censuses, from 2012 to 2015, were used. We used matrix population models to estimate asymptotic population growth rates and the elasticity values for the vital rates of the two populations of E. edulis. Life table response experiments were used to compare population performance by measuring the contribution of each vital rate to population growth rates. Population growth rates were not significantly different from one in either population, indicating that both populations were stable during the study period. However, the abundance of all ontogenetic stages was lower at the altitudinal range limit, which was related to decreases in some vital rates, especially fecundity. Additionally, there were higher elasticity values for the survival of immatures and reproductive individuals, compared to all other vital rates, in both populations. Synthesis. Our results show that even a small-scale environmental variation near range limits is sufficient to drive changes in the demography of this threatened palm. A minor increase in elevation approaching the limit of altitudinal distribution may reduce environmental suitability and affect population vital rates, thus contributing to setting upper altitudinal range limits for plants.

The progression rate of spinocerebellar ataxia type 2 changes with stage of disease.

BACKGROUND: Spinocerebellar ataxia type 2 (SCA2) affects several neurological structures, giving rise to multiple symptoms. However, only the natural history of ataxia is well known, as measured during the study duration. We aimed to describe the progression rate of ataxia, by the Scale for the Assessment and Rating of Ataxia (SARA), as well as the progression rate of the overall neurological picture, by the Neurological Examination Score for Spinocerebellar Ataxias (NESSCA), and not only during the study duration but also in a disease duration model. Comparisons between these models might allow us to explore whether progression is linear during the disease duration in SCA2; and to look for potential modifiers. RESULTS: Eighty-eight evaluations were prospectively done on 49 symptomatic subjects; on average (SD), study duration and disease duration models covered 13 (2.16) months and 14 (6.66) years of individuals' life, respectively. SARA progressed 1.75 (CI 95%: 0.92-2.57) versus 0.79 (95% CI 0.45 to 1.14) points/year in the study duration and disease duration models. NESSCA progressed 1.45 (CI 95%: 0.74-2.16) versus 0.41 (95% CI 0.24 to 0.59) points/year in the same models. In order to explain these discrepancies, the progression rates of the study duration model were plotted against disease duration. Then an acceleration was detected after 10 years of disease duration: SARA scores progressed 0.35 before and 2.45 points/year after this deadline (p = 0.013). Age at onset, mutation severity, and presence of amyotrophy, parkinsonism, dystonic manifestations and cognitive decline at baseline did not influence the rate of disease progression. CONCLUSIONS: NESSCA and SARA progression rates were not constant during disease duration in SCA2: early phases of disease were associated with slower progressions. Modelling of future clinical trials on SCA2 should take this phenomenon into account, since disease duration might impact on inclusion criteria, sample size, and study duration. Our database is available online and accessible to future studies aimed to compare the present data with other cohorts.

Chaperones in Polyglutamine Aggregation: Beyond the Q-Stretch.

Expanded polyglutamine (polyQ) stretches in at least nine unrelated proteins lead to inherited neuronal dysfunction and degeneration. The expansion size in all diseases correlates with age at onset (AO) of disease and with polyQ protein aggregation, indicating that the expanded polyQ stretch is the main driving force for the disease onset. Interestingly, there is marked interpatient variability in expansion thresholds for a given disease. Between different polyQ diseases the repeat length vs. AO also indicates the existence of modulatory effects on aggregation of the upstream and downstream amino acid sequences flanking the Q expansion. This can be either due to intrinsic modulation of aggregation by the flanking regions, or due to differential interaction with other proteins, such as the components of the cellular protein quality control network. Indeed, several lines of evidence suggest that molecular chaperones have impact on the handling of different polyQ proteins. Here, we review factors differentially influencing polyQ aggregation: the Q-stretch itself, modulatory flanking sequences, interaction partners, cleavage of polyQ-containing proteins, and post-translational modifications, with a special focus on the role of molecular chaperones. By discussing typical examples of how these factors influence aggregation, we provide more insight on the variability of AO between different diseases as well as within the same polyQ disorder, on the molecular level.

Phenotypic and genotypic heterogeneity of Enterococcus species isolated from food in Southern Brazil.

Enterococcus is an important group of lactic acid bacteria (LAB), which inhabits the gastrointestinal tract of humans and animals. These microorganisms can also be found in large groups of foods where they can play a beneficial role during food maturation processes or, conversely, can be used as a food contamination indicator. These microorganisms have an additional importance in various aspects of clinical microbiology. The aim of this study was to investigate and determine the phenotypic and genetic diversity in 55 enterococci isolated from different food sources. Phenotypic characteristics based upon substrate hydrolysis differences were used to identify different Enterococcus species. Analysis of the resultant data divided these species into eight Enterococcus phenotype groups. E. faecalis was the food isolate species with the greatest phenotypic variability. Fifty-five previously isolated Enterococcus strains were re-confirmed as belonging to this genus by PCR techniques. Randomly amplified polymorphic DNA (RAPD-PCR) was used to study the genetic variability using M13 primers and the resultant DNA fragments produced a database of different fingerprints. After statistical analyses of the RAPD-PCR profiles, 42 patterns were obtained and 6 different Enterococcus RAPD clusters (ERC) were identified. Genetic diversity was highest in ERC I, which grouped together approximately 40% of the E. faecium and E. faecalis isolates obtained from dairy products. Samples isolated from meat and vegetables offered the greatest genotype variability. Results of the present study suggest the presence of both phenotypic and genotypic variability within enterococci strains isolated from diverse sources of food common to Southern Brazil.