DNA Sequences Proximal to Human Mitochondrial DNA Deletion Breakpoints Prevalent in Human Disease Form G-quadruplexes, a Class of DNA Structures Inefficiently Unwound by the Mitochondrial Replicative Twinkle Helicase

Contains fulltext : 138677.pdf (publisher's version ) (Open Access)Mitochondrial DNA deletions are prominent in human genetic disorders, cancer, and aging. It is thought that stalling of the mitochondrial replication machinery during DNA synthesis is a prominent source of mitochondrial genome instability; however, the precise molecular determinants of defective mitochondrial replication are not well understood. In this work, we performed a computational analysis of the human mitochondrial genome using the "Pattern Finder" G-quadruplex (G4) predictor algorithm to assess whether G4-forming sequences reside in close proximity (within 20 base pairs) to known mitochondrial DNA deletion breakpoints. We then used this information to map G4P sequences with deletions characteristic of representative mitochondrial genetic disorders and also those identified in various cancers and aging. Circular dichroism and UV spectral analysis demonstrated that mitochondrial G-rich sequences near deletion breakpoints prevalent in human disease form G-quadruplex DNA structures. A biochemical analysis of purified recombinant human Twinkle protein (gene product of c10orf2) showed that the mitochondrial replicative helicase inefficiently unwinds well characterized intermolecular and intramolecular G-quadruplex DNA substrates, as well as a unimolecular G4 substrate derived from a mitochondrial sequence that nests a deletion breakpoint described in human renal cell carcinoma. Although G4 has been implicated in the initiation of mitochondrial DNA replication, our current findings suggest that mitochondrial G-quadruplexes are also likely to be a source of instability for the mitochondrial genome by perturbing the normal progression of the mitochondrial replication machinery, including DNA unwinding by Twinkle helicase

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Efeito do estresse climático sobre os parâmetros produtivos e fisiológicos de ovinos Santa Inês mantidos em confinamento na região litorânea do Nordeste do Brasil Effects of environmental stress on physiological parameters of feedlot sheep in the Northeast of Brazil

O presente estudo foi conduzido objetivando-se avaliar a influência do estresse climático sobre o desempenho produtivo e as respostas fisiológicas de ovinos da raça Santa Inês em confinamento. Foram avaliados dois ambientes (sombra e sol) e duas dietas com duas relações volumoso:concentrado (70C:30V: 70% de concentrado e 30% de volumoso; 30C:70V: 30% de concentrado e 70% de volumoso). Os animais mantidos à sombra e alimentados com dieta contendo alto teor de concentrado (70C:30V) apresentaram maior consumo de matéria seca (1258 g/animal/dia) e de proteína bruta (0,8% do peso vivo [PV] e 18 g/PV0,75) e maior ganho de peso (247 g/animal/dia). A temperatura nos ambientes de sol e sombra durante a tarde foi de 32,1 e 30,6:C e os valores do índice de temperatura e umidade (ITU), 82,3 e 81,1, respectivamente. Os animais alimentados com alto teor de concentrado (70C:30V) apresentaram maior freqüência respiratória (FR), tanto à sombra quanto ao sol (87 e 71 mov/min, respectivamente). Durante a tarde, a temperatura retal (TR) dos animais foi maior (39,1:C) que pela manhã (38,9:C). Porém, a TR mais elevada (39,3:C) foi observada nos animais mantidos à sombra e alimentados com alto percentual de concentrado (70C:30V). Independentemente do ambiente, os animais alimentados com alto teor de concentrado (70C:30V) apresentaram maior TR (39,2:C) do que aqueles alimentados com reduzido teor de concentrado (30C:70V) (38,8:C). O tipo de dieta teve efeito sobre a susceptibilidade dos animais ao estresse causado pelas condições ambientais críticas durante o experimento. Os animais da raça Santa Inês mostraram-se sensíveis ao estresse ambiental, uma vez que apresentaram menor desempenho produtivo, quando expostos a condições de ausência de sombra.<br>A study was conducted to determine the effect of environmental stress on physiological criteria of feedlot sheep. Treatments consisted of two different housing conditions (shade and no shade) and two levels of concentrate in the diet: high concentrate (70% of concentrate and 30% of roughage-70C:30R); low concentrate (30% of concentrate and 70% of roughage-30C:70R). Animals raised under shade and fed a high concentrate diet (70C:30R) had greater dry mater intake (1258 g/animal/day) and crude protein (0.8% of body weight (BW) and 18 g/BW0.75), as well as weight gain (247 g/animal/day). Air temperature in the afternoon was 30.6:C, under shade, and 32.1:C, under sun and values of temperature humidity index(THI), 82.3 and 81.1, respectively. Animals fed more concentrate (70C:30R) had also higher respiratory frequency (RF), either raised under shade (87 mov/min) or under the sun (71 mov/min.). Rectal temperature (RT) was higher in the afternoon (39.1:C) than in the morning (38.9:C), but animals raised under shade and fed diets with 70% concentrate (70C:30R) had the highest value of RT (39.3:C). Regardless of type of housing conditions, the animals that received more concentrate (70C:30R) had greater rectal temperature (39.2:C) than those fed a diet with less concentrate (30C:70R) (38.8:C). Therefore, the type of diet influenced the response of the animals to housing conditions. Moreover, Santa Inês sheep are susceptible to environmental stress because had lower performance when raised under no shade

Cama de frango e suplemento à base de microbiota ruminal em dietas de novilhas leiteiras: desempenho produtivo e avaliação econômica Broiler litter and supplement based on ruminal microbiota in dairy heifers diets: performance and economic evaluation

O trabalho foi conduzido para avaliar o desempenho de novilhas de rebanhos leiteiros recebendo dietas contendo cama-de-frango (CF) e suplemento à base de microbiota ruminal (SMR). Foram utilizadas 20 novilhas mestiças Holandês-zebu, com peso médio inicial de 276 kg e idade média de 13,8 meses. Os animais foram distribuídos aleatoriamente em arranjo fatorial 2x2, no qual foram estudados dois níveis de CF (15 e 30% na matéria seca) e de SMR (0 e 10 g/animal/dia), em delineamento inteiramente casualizado, com cinco repetições. Foram avaliados o consumo, o ganho de peso e a conversão alimentar e calculados os custos com alimentação, sendo composto um fluxo de caixa simulado para recria dos animais em confinamento e a pasto. Os consumos de matéria seca, matéria orgânica, proteína bruta e fibra em detergente neutro não foram influenciados pelos níveis de CF ou de SMR. O ganho de peso médio diário foi maior para as novilhas recebendo dietas com 15% de CF e 10 g de SMR (1,13 kg), em relação ao tratamento com 15% de CF sem SMR (0,85 kg). Entretanto, não houve diferenças em relação aos animais alimentados com 30% de CF com e sem SMR (1,09 e 0,97 kg). Não foram observadas diferenças para conversão alimentar da matéria seca e da fibra em detergente neutro, e houve interação entre os níveis de cama-de-frango e de suplemento para a conversão alimentar da proteína bruta. Melhor relação custo/benefício foi encontrada para o tratamento 30% de CF sem SMR, em relação ao nível de 15% sem SMR, porém não-significativa para os demais. O fluxo de caixa simulado mostrou que o sistema de recria de novilhas em confinamento foi economicamente mais vantajoso que o sistema em regime de pasto, para as condições do presente trabalho.<br>This research was carried out to evaluate the performance of dairy heifers fed with broiler litter (BL) and ruminal microbiota supplement (RMS). Twenty heifers, crossbred Holstein x zebu, with average weight and age of 276 kg and 13,8 months, respectively, were used. The animals were randomly allotted to a 2x2 factorial scheme: two levels of BL (15 and 30% in the diet dry matter) and RMS (0 and 10 g/heifer/day), in a completely randomized design, with five replications. The feed intake, weight gain, feeding efficiency and costs were evaluated, and an cash flow for feedlot and grazing systems was simulated. The intake of dry matter, organic matter, crude protein and neutral detergent fiber were not different for BL and RMS. Daily weight gain was higher for heifers fed diet with 15% BL and 10 g of RMS (1.13 kg), in relation to 15%BL without RMS (0.85 kg), but without differences for animals receiving only 30% BL or SMR (1.09 and 0.97 kg). There were no differences in dry matter and neutral detergent fiber conversions, but there was an interaction between BL and RMS levels and efficiency of crude protein utilization. Cost/benefit ratio was better for the 30% BL level, without RMS. Simulated cash flow showed that the feedlot system was economically more advantageous than grazing system, for the condition of this work

The Role of Cellular Senescence in Werner Syndrome: Toward Therapeutic Intervention in Human Premature Aging

Werner syndrome (WS) is a premature aging disorder used as a model of normal human aging. WS individuals have several characteristics of normal aging, such as cataracts, hair graying, and skin aging, but manifest these at an early age. Additionally, WS individuals have high levels of inflammatory diseases, such as atherosclerosis and type 2 diabetes. The in vivo aging in WS is associated with accelerated aging of fibroblasts in culture. The cause of the accelerated senescence is not understood, but may be due to the genomic instability that is a hallmark of WS. Genome instability results in activation of stress kinases, such as p38, and the p38-specific inhibitor SB203580, prevents the accelerated senescence seen in WS fibroblasts. However, oxidative damage plays a role, as low oxygen conditions and antioxidant treatment revert some of the accelerated senescence phenotype. The effects of oxidative stress appear to be suppressible by SB203580; however, it does not appear to be transduced by p38. As SB203580 is known to inhibit other kinases in addition to p38, this suggests that more than one kinase pathway is involved. The recent development of p38 inhibitors with different binding properties, specificities, and oral bioavailability, and of new potent and selective inhibitors of JNK and MK2, will make it possible to dissect the roles of various kinase pathways in the accelerated senescence of WS cells. If this accelerated senescence is reflective of WS aging in vivo, these kinase inhibitors may well form the basis of antiaging therapies for individuals with WS