Clévio Nóbrega
Biomedical and Life Sciences Researcher, PhD
Nóbrega Clévio, Carmo-Silva S, Albuquerque D, Vasconcelos-Ferreira A, Vijayakumar U-G, Mendonça L, Hirai H, Pereira de Almeida L (2015). Reestablishing Ataxin -2 downregulates translation of mutant ataxin-3 and alleviates Machado-Joseph disease. Brain (August, 2015 in press).
Machado-Joseph disease (MJD) is a progressive neurodegenerative disorder associated to the polyglutamine-expanded ataxin-3 (Atx3MUT) for which no therapy is available. Aiming at clarifying the mechanism of neurodegeneration we hypothesized that the abnormally long polyglutamine tract would interact aberrantly with ataxin-2 (Atx2), another polyglutamine protein whose function has recently been linked to translational regulation.
Using patient’s samples and cellular and animal’s models we found that in MJD a) Atx2 levels are reduced, and b) its subcellular localization is changed towards the nucleus. Restoring Atx2 levels by lentiviral-mediated overexpression: i) reduced Atx3MUT levels, ii) rescued behavior defects and neuropathology in a transgenic mouse model of MJD. Conversely, a) mutating the Atx2 motif that enables binding to its natural interactor and translation activator PABP, or b) overexpressing PABP, had opposite effects, increasing Atx3MUT translation and aggregation.
This work suggests that in MJD, Atx3MUT drives an abnormal reduction of Atx2 levels, which overactivates PABP, increases translation of Atx3MUT and other proteins and aggravates MJD. Re-establishment of Atx2 levels reduces Atx3MUT and alleviates MJD pathogenesis opening a new avenue for therapeutic intervention in this and potentially other polyglutamine disorders.
Aveleira C, Botelho M, Carmo-Silva S, Pascoal J, Ferreira-Marques M, Nóbrega Clévio, Cortes L, Valero J, Sousa-Ferreira L, Álvaro AR, Santana M, Kugler S, Pereira de Almeida L, Cavadas C (2015). Neuropepetide Y stimulates autophagy in hypothalamic neurons. Proceedings of the National Academy of Sciences USA,112:E1642-51.
Aging is characterized by autophagy impairment that contributes to age-related disease aggravation. Moreover, it was described that the hypothalamus is a critical brain area for whole-body aging development and has impact on lifespan. Neuropeptide Y (NPY) is one of the major neuropeptides present in the hypothalamus, and it has been shown that, in aged animals, the hypothalamic NPY levels decrease. Because caloric restriction (CR) delays aging, at least in part, by stimulating autophagy, and also increases hypo- thalamic NPY levels, we hypothesized that NPY could have a rele- vant role on autophagy modulation in the hypothalamus. Therefore, the aim of this study was to investigate the role of NPY on autophagy in the hypothalamus. Using both hypothalamic neuronal in vitro models and mice overexpressing NPY in the hypothalamus, we observed that NPY stimulates autophagy in the hypothalamus. Mechanistically, in rodent hypothalamic neurons, NPY increases autophagy through the activation of NPY Y1 and Y5 receptors, and this effect is tightly associated with the concerted activation of PI3K, MEK/ERK, and PKA signaling pathways. Modulation of hypothalamic NPY levels may be considered a potential strategy to produce protective effects against hypothalamic impairments associated with age and to delay aging.
Mendonça L, Nóbrega Clévio, Hirai H, Kaspar B, Pereira de Almeida L (2015) Transplantation of cerebellar neural stem cells improves motor coordination and neuropathology in a transgenic mouse model of Machado-Joseph disease. Brain, 138:320-35.
Machado-Joseph disease is a neurodegenerative disease without effective treatment. Patients with Machado-Joseph disease exhibit significant motor impairments such as gait ataxia, associated with multiple neuropathological changes including mutant ATXN3 inclusions, marked neuronal loss and atrophy of the cerebellum. Thus, an effective treatment of symptomatic patients with Machado-Joseph disease may require cell replacement, which we investigated in this study. For this purpose, we injected cerebellar neural stem cells into the cerebellum of adult Machado-Joseph disease transgenic mice and assessed the effect on the neuropath- ology, neuroinflammation mediators and neurotrophic factor levels and motor coordination. We found that upon transplantation into the cerebellum of adult Machado-Joseph disease mice, cerebellar neural stem cells differentiate into neurons, astrocytes and oligodendrocytes. Importantly, cerebellar neural stem cell transplantation mediated a significant and robust alleviation of the motor behaviour impairments, which correlated with preservation from Machado-Joseph disease-associated neuropathology, namely reduction of Purkinje cell loss, reduction of cellular layer shrinkage and mutant ATXN3 aggregates. Additionally, a significant reduction of neuroinflammation and an increase of neurotrophic factors levels was observed, indicating that transplantation of cerebellar neural stem cells also triggers important neuroprotective effects. Thus, cerebellar neural stem cells have the potential to be used as a cell replacement and neuroprotective approach for Machado-Joseph disease therapy.
Albuquerque D, Stice E, Rodríguez-López R, Manco L, Nóbrega Clévio (2015). Current review of genetics of human obesity: from molecular mechanisms to an evolutionary perspective. Mol. Genet. Genomics, 290:1191-221.
It is well-known that obesity is a complex multifactorial and heterogeneous condition with an important genetic component. Recently, major advances in obesity research emerged concerning the molecular mechanisms contributing to the obese condition. This review outlines several studies and data concerning the genetics and other important factors in the susceptibility risk to develop obe- sity. Based in the genetic etiology three main categories of obesity are considered: monogenic, syndromic, and com- mon obesity. For the monogenic forms of obesity, the gene causing the phenotype is clearly identified, whereas for the common obesity the loci architecture underlying the pheno- type is still being characterized. Given that, in this review we focus mainly in this obesity form, reviewing loci found until now by genome-wide association studies related with the susceptibility risk to develop obesity. Moreover, we also detail the obesity-related loci identified in children and in different ethnic groups, trying to highlight the complexity of the genetics underlying the common obese phenotype. Importantly, we also focus in the evolutionary hypotheses that have been proposed trying to explain how natural selection favored the spread of genes that increase the risk for an obese phenotype and how this predisposition to obe- sity evolved. Other factors are important in the obesity con- dition, and thus, we also discuss the epigenetic mechanisms involved in the susceptibility and development of obesity. Covering all these topics we expect to provide a complete and recent perspective about the underlying mechanisms involved in the development and origin of obesity. Only with a full understanding of the factors and mechanisms contributing to obesity, it will be possible to provide and allow the development of new therapeutic approaches to this condition.
Nóbrega Clévio*, Nascimento-Ferreira I*, Onofre I, Albuquerque D, Hirai H, Déglon N, Pereira de Almeida L (2014) RNA interference mitigates motor and neuropathological deficits in a cerebellar mouse model of Machado-Joseph disease. PlosOne, 9(8):e100086. *Equal contribution
Machado-Joseph disease or Spinocerebellar ataxia type 3 is a progressive fatal neurodegenerative disorder caused by the polyglutamine-expanded protein ataxin-3. Recent studies demonstrate that RNA interference is a promising approach for the treatment of Machado-Joseph disease. However, whether gene silencing at an early time-point is able to prevent the appearance of motor behavior deficits typical of the disease when initiated before onset of the disease had not been explored. Here, using a lentiviral-mediated allele-specific silencing of mutant ataxin-3 in an early pre-symptomatic cerebellar mouse model of Machado-Joseph disease we show that this strategy hampers the development of the motor and neuropathological phenotypic characteristics of the disease. At the histological level, the RNA-specific silencing of mutant ataxin-3 decreased formation of mutant ataxin-3 aggregates, preserved Purkinje cell morphology and expression of neuronal markers while reducing cell death. Importantly, gene silencing prevented the development of impairments in balance, motor coordination, gait and hyperactivity observed in control mice. These data support the therapeutic potential of RNA interference for Machado-Joseph disease and constitute a proof of principle of the beneficial effects of early allele-specific silencing for therapy of this disease.
Albuquerque D, Estévez MN, Víbora PB, Giralt PS, Balsera AM, Cortés PG, López MJ, Luego LM, Gervasini G, Hernández SB, Arroyo-Díez J, Vacas MA, Nóbrega Clévio, Manco L, Rodríguez-López R (2014) Novel variants in the MC4R and LEPR genes among severly obese children from the Iberian population. Ann Human Genet, 78:195-207.
We screened for mutations in the MC4R and LEPR genes and investigated the genotype-phenotype correlation in obese individuals belonging to families with evident hereditary patterns of severe and early-onset obesity among the Iberian population.
A total of 202 unrelated and severely obese patients since childhood, were enrolled in the study. Bidirectional sequencing of the MC4R gene was carried out in all patients; the LEPR gene was sequenced in 15 individuals based on additional clinical signals. Segregation analysis and/or genotype-phenotype description was performed for subjects with the new mutations and with presumably functional variants.
Ten MC4R gene mutations were identified in the heterozygous state in 10 patients. Mutations p.R147G and p.G323E are new and mutations p.R7H, p.G32E, p.H76R, p.V103I, p.S127L, p.T150I, p.I251L and p.G252S were previously described. A new dinucleotide insertion -77_-76insTA in the promoter region of the LEPR gene was found in the heterozygous state in one patient.
The new p.R147G and the previously published p.R7H, p.S127L, p.T150I and p.G252S MC4R mutations, cosegregate with obesity in our patients and were predicted to be deleterious. For the novel MC4R p.G323E and LEPR -77_-76insTA gene mutations, the genotype-phenotype correlation and bioinformatic analysis did not clarify whether these mutations are indeed implicated in obesity
Albuquerque D, Nóbrega Clévio, Rodriguez-López R, Manco L (2014) Association study of common polymorphisms in MSRA, TFAP2B, MC4R, NRXN3, PPARGC1A, TMEM18, SEC16B, HOXB5 and OLFM4 genes with obesity-related traits amons Portuguese children. J Hum Genet, 59:307-13.
At least 52 genetic loci were associated with obesity-related traits. However, little is known about the genetic basis of obesity among children. This study aims to test whether 10 polymorphisms in obesity-related genes methionine sulfoxide reductase A (MSRA), transcription factor AP-2 beta (TFAP2B), melanocortin 4 receptor (MC4R), neurexin 3 (NRXN3), peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PPARGC1A), transmembrane protein 18 (TMEM18), homolog of
S. cerevisiae Sec16 (SEC16B), homeobox B5 (HOXB5) and olfactomedin 4 (OLFM4) are associated with the risk of obesity in Portuguese children. A total of 730 children aging from 6 to 12 years old, recruited randomly from public schools in Portugal, were analysed. Anthropometric measurements were obtained and children were classified into three phenotypic groups, normal weight (n 1⁄4 256), overweight (n 1⁄4 320) and obese (n 1⁄4 154), according to the International Obesity Task Force cutoffs. Polymorphisms were genotyped by allelic discrimination TaqMan assays. The MC4R rs12970134 polymorphism was nominally associated with body mass index (BMI) (P 1⁄4 0.035), BMI Z-score (P 1⁄4 0.043) and waist circumference (P 1⁄4 0.020), and borderline associated with weight (P 1⁄4 0.053). Near nominal associations were also found for the PPARGC1A rs8192678 polymorphism with weight (P 1⁄4 0.061), and for the MSRA rs545854 polymorphism with BMI (P 1⁄4 0.055) and BMI Z-score
(P 1⁄4 0.056). Furthermore, logistic regression showed that MC4R rs12970134 and TFAP2B rs987237 were nominally, respectively, associated (P 1⁄4 0.029) and borderline associated (P 1⁄4 0.056) with the obese phenotype. This study highlighted the possible association of MC4R, PPARGC1A, MSRA and TFAP2B polymorphisms with several obesity-related traits in a sample of Portuguese children. The two significant associated TFAP2B rs987237 and MC4R rs12970134 polymorphisms showed an opposite direction of effect to that in the original reports
Nascimento-Ferreira I*, Nóbrega Clévio*, Ferreira-Vasconcelos A, Onofre I, Albuquerque D, Aveleira C, Hirai H, Déglon N, Pereira de Almeida L (2013) Beclin-1 mitigates motor and neuropathological deficits in genetic mouse models of Machado-Joseph disease. Brain, 136:2173-88. *Equal contribution.
Machado-Joseph disease or spinocerebellar ataxia type 3, the most common dominantly-inherited spinocerebellar ataxia, results from translation of the polyglutamine-expanded and aggregation prone ataxin 3 protein. Clinical manifestations include cere- bellar ataxia and pyramidal signs and there is no therapy to delay disease progression. Beclin 1, an autophagy-related protein and essential gene for cell survival, is decreased in several neurodegenerative disorders. This study aimed at evaluating if lentiviral-mediated beclin 1 overexpression would rescue motor and neuropathological impairments when administered to pre- and post-symptomatic lentiviral-based and transgenic mouse models of Machado-Joseph disease. Beclin 1-mediated significant improvements in motor coordination, balance and gait with beclin 1-treated mice equilibrating longer periods in the Rotarod and presenting longer and narrower footprints. Furthermore, in agreement with the improvements observed in motor function beclin 1 overexpression prevented neuronal dysfunction and neurodegeneration, decreasing formation of polyglutamine-expanded aggregates, preserving Purkinje cell arborization and immunoreactivity for neuronal markers. These data show that overexpres- sion of beclin 1 in the mouse cerebellum is able to rescue and hinder the progression of motor deficits when administered to pre- and post-symptomatic stages of the disease.
Nóbrega Clévio*, Nascimento-Ferreira I*, Onofre I, Albuquerque D, Hirai H, Déglon N, Pereira de Almeida L (2013) Silencing mutant ataxin-3 rescues motor deficits and neuropathology in Machado-Joseph disease transgenic mice. PLoS One 8(1):e52396. *Equal contribution.
Machado-Joseph disease (MJD) or spinocerebellar ataxia type 3 (SCA3) is an autosomal dominantly-inherited neurodegenerative disorder caused by the over-repetition of a CAG codon in the MJD1 gene. This expansion translates into a polyglutamine tract that confers a toxic gain-of-function to the mutant protein – ataxin-3, leading to neurodegeneration in specific brain regions, with particular severity in the cerebellum. No treatment able to modify the disease progression is available. However, gene silencing by RNA interference has shown promising results. Therefore, in this study we investigated whether lentiviral-mediated allele-specific silencing of the mutant ataxin-3 gene, after disease onset, would rescue the motor behavior deficits and neuropathological features in a severely impaired transgenic mouse model of MJD. For this purpose, we injected lentiviral vectors encoding allele-specific silencing-sequences (shAtx3) into the cerebellum of diseased transgenic mice expressing the targeted C-variant of mutant ataxin-3 present in 70% of MJD patients. This variation permits to discriminate between the wild-type and mutant forms, maintaining the normal function of the wild-type allele and silencing only the mutant form. Quantitative analysis of rotarod performance, footprint and activity patterns revealed significant and robust alleviation of gait, balance (average 3-fold increase of rotarod test time), locomotor and exploratory activity impairments in shAtx3-injected mice, as compared to control ones injected with shGFP. An important improvement of neuropathology was also observed, regarding the number of intranuclear inclusions, calbindin and DARPP-32 immunoreactivity, fluorojade B and Golgi staining and molecular and granular layers thickness. These data demonstrate for the first time the efficacy of gene silencing in blocking the MJD-associated motor-behavior and neuropathological abnormalities after the onset of the disease, supporting the use of this strategy for therapy of MJD.
Costa PM, Cardoso AL, Nóbrega Clévio, Pereira de Almeida L, Bruce JN, Cannol P, Pedroso de Lima MC (2013) MicroRNA-21 silencing enhances the cytotoxic effect of the antiangiogenic drug sunitinib in gliobastoma. Hum Mol Genet 22:904-18.
Highly malignant glioblastoma (GBM) is characterized by high genetic heterogeneity and infiltrative brain in- vasion patterns, and aberrant miRNA expression has been associated with hallmark malignant properties of GBM. The lack of effective GBM treatment options prompted us to investigate whether miRNAs would con- stitute promising therapeutic targets toward the generation of a gene therapy approach with clinical signifi- cance for this disease. Here, we show that microRNA-21 (miR-21) is upregulated and microRNA-128 (miR-128) is downregulated in mouse and human GBM samples, a finding that is corroborated by analysis of a large set of human GBM data from The Cancer Genome Atlas. Moreover, we demonstrate that oligonucleotide- mediated miR-21 silencing in U87 human GBM cells resulted in increased levels of the tumor suppressors PTEN and PDCD4, caspase 3/7 activation and decreased tumor cell proliferation. Cell exposure to pifithrin, an inhibitor of p53 transcriptional activity, reduced the caspase activity associated with decreased miR-21 expression. Finally, we demonstrate for the first time that miR-21 silencing enhances the antitumoral effect of the tyrosine kinase inhibitor sunitinib, whereas no therapeutic benefit is observed when coupling miR- 21 silencing with the first-line drug temozolomide. Overall, our results provide evidence that miR-21 is uni- formly overexpressed in GBM and constitutes a highly promising target for multimodal therapeutic approaches toward GBM.
Nóbrega Clévio*, Nascimento-Ferreira I*, Onofre I, Albuquerque D, Conceição M, Déglon N, Pereira de Almeida L (2013) Overexpression of mutant ataxin-3 in mouse cerebellum induces ataxia and cerebellar neuropathology. Cerebellum, 12:441-55 *Equal contribution.
Machado-Joseph disease (MJD), also known as spinocerebellar ataxia type 3 (SCA3) is a fatal, dominant neurodegenerative disorder caused by the polyglutamine-expanded protein ataxin-3. Clinical manifestations include cerebellar ataxia and pyramidal signs culminating in severe neuronal degeneration. Currently, there is no therapy able to modify of disease progression.
In the present study, we aimed at investigating one of the most severely attained brain regions in the disorder - the cerebellum, and the behavioral defects associated with the neuropathology in this region. For this purpose we injected lentiviral vectors encoding full-length human mutant ataxin-3 in the mouse cerebellum of three weeks old C57/BL6 mice.
We show that circumscribed expression of human mutant ataxin-3 in the cerebellum mediates within a short time frame - 6 weeks, the development of a behavioral phenotype including reduced motor coordination, ataxic wide-based gait and hyperactivity. Furthermore, expression of mutant ataxin-3 resulted in the accumulation of intranuclear inclusions, neuropathological abnormalities and neuronal death.
This data shows that lentiviral-based expression of mutant ataxin-3 in the mouse cerebellum induces localized neuropathology, which is sufficient to generate a behavioral ataxic phenotype. Moreover, this approach provides a physiologically relevant, cost- and time-effective animal model to gain further insights into the pathogenesis of MJD and for the evaluation of experimental therapeutics of MJD.
Albuquerque D, Nóbrega Clévio, Manco L (2013) Association of FTO polymorphisms with obesity and obseity-related outcomes in Portuguese children. PLoS One 8(1):e54370.
Background: Several studies have reported an association between single nucleotide polymorphisms in the first intron of the FTO gene and body mass index (BMI) or obesity. However, this association has not yet been studied among the Portuguese population. This study aims to assess the association of three FTO polymorphisms (rs1861868, rs1421085 and rs9939609) with obesity-related outcomes in a sample of Portuguese children.
Methods: We examined a total of 730 children, 256 normal-weight (55.9% girls), 320 overweight (45.3% girls) and 154 obese (53.2% girls), aging from 6 to 12-years-old, recruited randomly from public schools in the central region of Portugal. DNA samples were genotyped for the three polymorphisms by allelic discrimination TaqMan assay. Association of the FTO polymorphisms with several anthropometric traits was investigated. Additionally, we tested association with the risk of obesity using overweight and obese vs. normal-weight children.
Results: We found significant associations of rs9939609 and rs1421085 polymorphisms with weight, BMI, BMI Z-score, waist circumference and hip circumference, even after age and gender adjustment (p,0.05 in all traits). For rs1861868 polymorphism, marginally significant associations were obtained with weight (p=0.081) and BMI (p=0.096) after adjustment for age and gender. In case-control studies, both rs9939609 and rs1421085 polymorphisms were significantly associated with obesity (OR 1.97; 95% CI, 1.08–3.59; p = 0.026; OR 2.11; 95% CI, 1.17–3.81; p = 0.013, respectively) but not with overweight (p.0.05). Haplotype analyses identified two combinations (ACA and GCA) associated with a higher risk of obesity (OR 1.53; 95% CI, 1.06–2.22; p = 0.023; OR 1.73; 95% CI, 1.06–2.87; p = 0.030, respectively).
Conclusions: This study provides the first evidence for the association of FTO polymorphisms with anthropometric traits and risk of obesity in Portuguese children.
Albuquerque D, Nóbrega Clévio, Manco L (2012) The lactase persistence -13190 C>T polymorphism shows indication of association with abdominal obesity among Portuguese children. Acta Paediatr., 102(4):e153-7.
Aim: The -13910C>T single nucleotide polymorphism located upstream of the lactase gene (LCT) was found tightly associated with lactase persistence in European populations. Recently, it was also associated with body mass index (BMI) and obesity in European adults. The aim of this study was to test the association of -13910C>T polymorphism with obesity-related traits and risk of obesity in children.
Methods: We genotyped 580 Portuguese children (6–12-year-olds) for the -13910C>T polymorphism using TaqMan probes by real-time PCR. Anthropometric measurements were assessed in all children. Obesity was defined according to the International Obesity Task Force (IOTF) cut-offs and abdominal obesity using the sex and age-specific ! 90th waist circumference percentile.
Results: We found indication for an association between the-13910*T allele and children abdominal obesity (odds ratio [OR] = 1.41; 95% confidence intervals [CI]: 1.03–1.94;
p = 0.030). Under the dominant model, the indicative association was observed between the LCT-13910 CT/TT genotypes and abdominal obesity, remaining significant after adjustment for age and gender (OR = 1.65; 95% CI: 1.04–2.60; p = 0.029). No association was detected with the risk of obesity (p = 0.350).
Conclusion: Our results suggest that the -13910C>T polymorphism may predispose to abdominal obesity in Portuguese children. The association with BMI or risk of obesity, previously observed in adults, was not confirmed.
Albuquerque D, Nóbrega Clévio, Samouda H, Manco L (2012) Assessment of obesity and abdominal obesity among Portuguese children. Acta Med Port. 25(3):169-73.
background: Childhood obesity is a major public health issue in developed countries, and frequently proceeds into adulthood. The aim of this study was to estimate the prevalence of obesity and abdominal fat distribution in 6-12 years old children from the central region of Portugal, providing new data about trends on prevalence, epidemiology and evolution in obesity.
Methods: Weight, height and waist circumference were measured in a random representative sample of 1,433 children (747 girls and 686 boys) from public schools in 2011. International Obesity Task Force (IOTF) cut-offs were used to define overweight and obesity. Abdominal obesity was estimated using the sex and age-specific ≥ 90th waist circumference percentile and waist-to-height ratio cut-off. results: The prevalence of overweight and obesity among children was 33.0%; 10.7% were obese. Overweight was significantly higher in boys than in girls (p = 0.044), whereas no gender differences was found in obesity (10.6 % in boys and 10.7% in girls, p = 0.571). The prevalence of abdominal obesity based on waist circumference was similar in girls and boys (3.8% vs. 3.9% respectively; p = 0.924), but significantly higher in boys than in girls based on waist-to-height ratio (28.1% vs. 19.4%, respectively; p = 0.009). Comparison with previous studies showed a slightly increase in overweight/obesity in children of central Portugal in the last 10 years, reaching values of 40.0% prevalence in the 7-9 years old.
conclusion: In conclusion, this study shows a very high prevalence of overweight/obesity and abdominal obesity among Portuguese children, following the trend of other southern European countries. Thus, it is of the utmost importance the development of preventive and treatment strategies.
Sousa-Ferreira L, Garrido M, Nascimento-Ferreira I, Nóbrega Clévio, Santos-Carvalho A, Álvaro AR, Rosmaninho-Salgado J, Kaster M, Kügler S, de Almeida LP, Cavadas C (2011) Moderate long-term modulation of Neuropeptide Y in the Arcuate Nucleus induces energy balance alterations in adult rats. PLoS ONE, 6(7):e22333.
Neuropeptide Y (NPY) produced by arcuate nucleus (ARC) neurons has a strong orexigenic effect on target neurons. Hypothalamic NPY levels undergo wide-ranging oscillations during the circadian cycle and in response to fasting and peripheral hormones (from 0.25 to 10-fold change). The aim of the present study was to evaluate the impact of a moderate long-term modulation of NPY within the ARC neurons on food consumption, body weight gain and hypothalamic neuropeptides. We achieved a physiological overexpression (3.6-fold increase) and down-regulation (0.5-fold decrease) of NPY in the rat ARC by injection of AAV vectors expressing NPY and synthetic microRNA that target the NPY, respectively. Our work shows that a moderate overexpression of NPY was sufficient to induce diurnal over-feeding, sustained body weight gain and severe obesity in adult rats. Additionally, the circulating levels of leptin were elevated but the immunoreactivity (ir) of ARC neuropeptides was not in accordance (POMC-ir was unchanged and AGRP-ir increased), suggesting a disruption in the ability of ARC neurons to response to peripheral metabolic alterations. Furthermore, a dysfunction in adipocytes phenotype was observed in these obese rats. In addition, moderate down-regulation of NPY did not affect basal feeding or normal body weight gain but the response to food deprivation was compromised since fasting- induced hyperphagia was inhibited and fasting-induced decrease in locomotor activity was absent. These results highlight the importance of the physiological ARC NPY levels oscillations on feeding regulation, fasting response and body weight preservation, and are important for the design of therapeutic interventions for obesity that include the NPY.
Khadem M, Camacho R, Nóbrega Clévio (2011) Studies of the species barrier between Drosophila subobscura and D. madeirensis V: the importance of sex-linked inversion in preserving species identity. J Evol Biol 24:1263-1273.
The X chromosome is known to exert a disproportionately large effect on characters related to post-zygotic reproductive isolation. There is also growing evidence about the important role of the chromosomal regions with reduced recombination (such as inversions) in maintaining the identity of closely related species. Using molecular markers, we examine the effect of different regions of the X chromosome on determination of hybrid traits (viability, testes size, sperm motility and morphological anomalies) in hybrid males between Drosophila madeirensis and Drosophila subobscura. The preponderant effect of a region localized inside the A2 inversion in the X chromosome in all hybrid traits is identified. Other marked regions exert a weaker influence or only influence some of the hybrid trait. Our results confirm the crucial role of sex-linked chromosomal inversion in preserving the identity of species with incomplete reproductive isolation. The specific genomic make-up of parental lines used to perform crosses has a great effect on hybrid fitness.
Alves S, Nascimento-Ferreira I, Dufour N, Hassig R, Auregan G, Nóbrega Clévio, Brouillet E, Hantraye P, Pedroso de Lima MC, Deglon N, de Almeida LP (2010) Silencing ataxin-3 mitigates degeneration in a rat model of Machado-Joseph disease: no role for wild-type ataxin-3? Hum Mol Genet 19:2380-2394.
Machado–Joseph disease or spinocerebellar ataxia type 3 (MJD/SCA3) is a fatal, autosomal dominant dis- order caused by a cytosine-adenine-guanine expansion in the coding region of the MJD1 gene. RNA interfer- ence has potential as a therapeutic approach but raises the issue of the role of wild-type ataxin-3 (WT ATX3) in MJD and of whether the expression of the wild-type protein must be maintained. To address this issue, we both overexpressed and silenced WT ATX3 in a rat model of MJD. We showed that (i) overexpression of WT ATX3 did not protect against MJD pathology, (ii) knockdown of WT ATX3 did not aggravate MJD pathology and that (iii) non-allele-specific silencing of ataxin-3 strongly reduced neuropathology in a rat model of MJD. Our findings indicate that therapeutic strategies involving non-allele-specific silencing to treat MJD patients may be safe and effective.
Nóbrega Clévio, Khadem M, Aguade M, Segarra C (2008) Genetic exchange versus genetic differentiation in a medium-sized inversion of Drosophila: the A2/Ast arrangements of Drosophila subobscura. Mol Biol Evol 25:1534-1543.
Chromosomal inversion polymorphism affects nucleotide variation at loci associated with inversions. In Drosophila subobscura, a species with a rich chromosomal inversion polymorphism and the largest recombinational map so far reported in the Drosophila genus, extensive genetic structure of nucleotide variation was detected in the segment affected by the O3 inversion, a moderately sized inversion at Muller’s element E. Indeed, a strong genetic differentiation all over O3 and no evidence of a higher genetic exchange in the center of the inversion than at breakpoints were detected. In order to ascertain, whether other polymorphic and differently sized inversions of D. subobscura also exhibited a strong genetic structure, nucleotide variation in 5 gene regions (P236, P275, P150, Sxl, and P125) located along the A2 inversion was analyzed in Ast and A2 chromosomes of D. subobscura. A2 is a medium-sized inversion at Muller’s element A and forms a single inversion loop in heterokaryotypes. The lower level of variation in A2 relative to Ast and the significant excess of low-frequency variants at polymorphic sites indicate that nucleotide variation at A2 is not at mutation–drift equilibrium. The closest region to an inversion breakpoint, P236, exhibits the highest level of genetic differentiation (FST) and of linkage disequilibrium (LD) between arrangements and variants at nucleotide polymorphic sites. The remaining 4 regions show a higher level of genetic exchange between A2 and Ast chromosomes than P236, as revealed by FST and LD estimates. However, significant genetic differentiation between the Ast and A2 arrangements was detected not only at P236 but also in the other 4 regions separated from the nearest breakpoint by 1.2–2.9 Mb. Therefore, the extent of genetic exchange between arrangements has not been high enough to homogenize nucleotide variation in the center of the A2 inversion. A2 can be considered a typical successful inversion of D. subobscura according to its relative length. Chromosomal inversion polymorphism of D. subobscura might thus cause the genome of this species to be highly structured and to harbor different gene pools that might contribute to maintain adaptations to particular environments.
