Epigenetic control of early neurodegenerative events in diabetic retinopathy by the histone deacetylase SIRT6

Diabetic retinopathy (DR ) is one of the common complications associated with diabetes mellitus and the leading cause of blindness worldwide. Recent research has demonstrated that DR is not only a microvascular disease but may be a result of neurodegenerative processes. Moreover, glucose‐induced neuron and glial cell damage may occur shortly after the onset of diabetes which makes the disease hard to diagnose at early stages. SIRT 6, a NAD ‐dependent sirtuin deacylase, modulates aging, energy metabolism, and neurodegeneration. In previous studies we showed that SIRT 6 deficiency causes major retinal transmission defects, changes in the expression of glycolytic genes, and elevated levels of apoptosis. Given the importance of glucose availability for retinal function and the critical role of SIRT 6 in modulating glycolysis, we aimed to analyze SIRT 6 participation in the molecular machinery that regulates the development of experimental DR . Using non‐obese diabetic mice, we determined by western blot that 2 weeks after the onset of the disease, high glucose concentrations induced retinal increase in a neovascularization promoting factor (vascular endothelial growth factor, VEGF ), and the loss of a neuroprotective factor (brain‐derived neurotrophic factor, BDNF) associated with reduced levels of SIRT 6 and increased acetylation levels of its substrates (H3K9 and H3K56) suggesting a deregulation of key neural factors. Noteworthy, retinas from CNS conditionally deleted SIRT 6 mice showed a resemblance to diabetic retinas exhibiting lower protein levels of BDNF factor and increased protein levels of VEGF . Moreover, cultured Müller glial cells subjected to high glucose concentrations exhibited decreased levels of SIRT 6 and increased levels of H3K56 acetylation. In addition, the increment of VEGF levels induced by high glucose was reverted by the over‐expression of SIRT 6 in this cell type. Accordingly, siRNA experiments showed that, when SIRT 6 was silenced, VEGF levels increased. Our findings suggest that epigenetically regulated neurodegenerative events may occur at an early diabetic stage prior to the characteristic proliferative and vascular changes observed at a later diabetic stage.

     

Effect of biocatalyst activity changes in continuous reactions in the gas phase

Summary In the gas phase bioreactions, continuous production rate depends on the biocatalyst activity and complete dehydration causes the biocatalyst to lose most of its activity. To overcome these difficulties, a theoretical method is suggested along with the new design of biocatalyst. This will be applicable and helpful for the optimization of the gas phase continuous bioreaction.Nomenclature C_A ethanol concentration [mol/mL] - C_P acetaldehyde concentration [mol/mL] - X_P acetaldehyde composition     

Fitness consequences of pheromone production and host selection strategies in a tree-killing bark beetle (Coleoptera: Curculionidae: Scolytinae)

Timing of arrival at a resource often determines an individual’s reproductive success. Tree-killing bark beetles can reproduce in healthy trees by attacking in adequate numbers to overcome host defences that could otherwise be lethal. This process is mediated by aggregation and antiaggregation pheromones. Beetles that arrive early in such a “mass attack” must contend with undiminished tree defences, and produce enough pheromones to attract more beetles, but have a head start on gallery construction and egg-laying. Beetles that arrive late may be impeded by competition and diminishing availability of phloem, but should experience fewer costs associated with pheromone production and battling tree defences. We investigated relationships between timing of arrival, body size, pheromone production and fitness in the southern pine beetle, Dendroctonus frontalis. In field experiments, we captured beetles that arrived early (pioneers) and late on slash pine trees, Pinus elliottii, and measured pheromone amounts in their hindguts. We marked gallery entrances of beetles as they landed on a tree and measured their reproductive success after the attack terminated. We found no difference in body size or pheromone amounts between early and late arrivers. Most beetles arrived at the middle of the attack sequence, and excavated longer galleries per day than early arrivers. The number of offspring produced per day by beetles that established galleries midway through mass attack was higher than those that arrived early or very late in the sequence. Our results suggest that beetles do not exhibit adaptive phenotypic plasticity in pre-landing pheromone production, depending on the extent of previous colonisation of a host. Rather, it appears that stabilising selection favours beetles that attack in the middle of the sequence, and contributes to attack synchrony. Synchronous attack on trees is essential before population booms characteristic of tree-killing bark beetles can occur in nature.     

Selection in monoculture vs. mixture alters plant metabolic fingerprints

Aims In grassland biodiversity experiments, positive biodiversity effects on primary productivity increase over time. Recent research has shown that differential selection in monoculture and mixed-species communities leads to the rapid emergence of monoculture and mixture types, adapted to their own biotic community. We used eight plant species selected for 8 years in such a biodiversity experiment to test if monoculture and mixture types differed in metabolic profiles using infrared spectroscopy.Methods Fourier transform infrared spectroscopy (FTIR) was used to assess metabolic fingerprints of leaf samples of 10 individuals of each species from either monocultures or mixtures. The FTIR spectra were analyzed using multivariate procedures to assess (i) whether individuals within species could be correctly assigned to monoculture or mixture history based on the spectra alone and (ii) which parts of the spectra drive the group assignment, i.e. which metabolic groups were subject to differential selection in monocultures vs. mixtures.Important findings Plant individuals within each of the eight species could be classified as either from monoculture or mixture selection history based on their FTIR spectra. Different metabolic groups were differentially selected in the different species; some of them may be related to defense of pathogens accumulating more strongly in monocultures than in mixtures. The rapid selection of the monoculture and mixture types within the eight study species could have been due to a sorting-out process based on large initial genetic or epigenetic variation within the species.     

Integrity of mating behaviors and seasonal reproduction in coyotes (Canis latrans) following treatment with estradiol benzoate

Coyotes (Canis latrans) are seasonally monestrous and form perennial pair-bonds. Breeding is dominated by each pack's alpha male and female, and both sexes share responsibility for territory defense and pup-rearing. They are also opportunistic predators on domestic livestock and pets. But while dominant adults have been implicated as primary killers, depredation is reduced when coyotes are without pups. Contraception, therefore, may represent a non-lethal solution for conflicts between coyotes and humans. Steroid hormones successfully control fertility in some species, but have been considered contraindicated in wildlife and canids in particular; specific concerns include possible induction of aberrant behavior, or uterine and hematopoietic pathologies. Herein we describe a study examining the physiological effectiveness, health safety, and behavioral consequences following treatment of estrous coyotes with exogenous estrogen. We treated captive adult female coyotes in estrus with 0.01 mg/kg estradiol benzoate (EB), either before (n = 5) or immediately after ovulation (n = 6), then documented reproductive outcome, physiological variables and behavioral responses, during and after treatment. Pregnancy was averted in six females treated after ovulation, suggesting that appropriate timing of treatment proved crucial. A transient suppression of sexual behavior was observed, and in some cases, estrus appeared slightly lengthened. However, neither ovulation nor mating behavior was fully suppressed. Importantly, non-pregnant females (and their mates) displayed diestrous socio-sexual behavior similar to pregnant coyotes (behavioral pseudopregnancy). Furthermore, non-pregnant coyotes did not mate again until the next native breeding season, and we observed no deleterious physiological effects during diestrus or subsequent ovarian cycles.     

Monitoring enzyme catalysis in the multimeric state: Direct observation of Arthrobacter 4-hydroxybenzoyl-coenzyme A thioesterase catalytic complexes using time-resolved electrospray ionization mass spectrometry

The ability to examine real-time reaction kinetics for multimeric enzymes in their native state may offer unique insights into understanding the catalytic mechanism and its interplay with three-dimensional structure. In this study, we have used a time-resolved electrospray mass spectrometry approach to probe the kinetic mechanism of 4-hydroxybenzoyl-coenzyme A (4-HBA-CoA) thioesterase from Arthrobacter sp. strain SU in the millisecond time domain. Intact tetrameric complexes of 4-HBA-CoA thioesterase with up to four natural substrate (4-HBA-CoA) molecules bound were detected at times as early as 6 ms using an online rapid-mixing device directly coupled to an electrospray ionization time-of-flight mass spectrometer. Species corresponding to the formation of a folded tetramer of the thioesterase at charge states 16+, 17+, 18+, and 19+ around m/z 3800 were observed and assigned as individual tetramers of thioesterase and noncovalent complexes of the tetramers with up to four substrate and/or product molecules. Real-time evaluation of the reaction kinetics was accomplished by monitoring change in peak intensity corresponding to the substrate and product complexes of the tetrameric protein. The mass spectral data suggest that product 4-HBA is released from the active site of the enzyme prior to the release of product CoA following catalytic turnover. This study demonstrates the utility of this technique to provide additional molecular details for an understanding of the individual enzyme states during the thioesterase catalysis and ability to observe real-time interactions between enzyme and substrates and/or products in the millisecond time range.     

Calcium influences the stability and conformation of rotavirus SAH glycoprotein VP7 expressed in Dictyostelium discoideum

We have previously reported expression of the rotavirus outer capsid glycoprotein, VP7, in the relatively new expression host, Dictyostelium discoideum. To optimise yields of recombinant VP7, we examined the role of Ca²⁺ since stability of both VP7 and mature rotavirus during a rotavirus infection are calcium-dependent. Low micromolar levels of free extracellular Ca²⁺ were required to maximise yields of VP7 in D. discoideum whilst levels of VP7 were reduced following depletion of intracellular Ca²⁺ reserves using A23187 and EGTA. Immunoblot analysis suggested that VP7 was being degraded in an intracellular compartment. Immunoprecipitation with a conformation-dependent neutralising antibody confirmed that EGTA-induced Ca²⁺ chelation alters the conformation of VP7. These results suggest that stability of VP7 is dependent on maintaining adequate levels of intracellular Ca²⁺ and that conformational changes in VP7 which occur following depletion of Ca²⁺ reserves induce rapid proteolysis of the protein. Since these results establish conditions for expressing optimal levels of VP7 in the correct conformation they have important implications for the development of a subunit vaccine based on recombinant VP7.     

No Evidence for a Difference in Neuropsychological Profile among Carriers and Noncarriers of the FMR1 Premutation in Adults under the Age of 50

The 5′ untranslated region of the fragile X mental retardation gene, FMR1, contains a polymorphic CGG repeat. Expansions of this repeat are associated with a spectrum of disorders. Full mutation alleles, repeats ≥ 200, are associated with fragile X syndrome. Premutation alleles, repeats of ~55–199, are associated with a tremor-ataxia syndrome most commonly in older males and primary ovarian insufficiency in females. However, the neuropsychological impact of carrying a premutation allele is presently unclear in younger adults. In this study, we analyzed neuropsychological scores for 138 males and 506 females ascertained from the general population and from families with a history of fragile X syndrome. Subjects were age 18–50 years and had varying repeat lengths. Neuropsychological scores were obtained from measures of general intelligence, memory, and executive functioning, including attention. Principal component analysis followed by varimax rotation was used to create independent factors for analysis. These factors were modeled for males and females separately via a general linear model that accounted for correlation among related subjects. All models were adjusted for potential confounders, including age at testing, ethnicity, and household income. Among males, no repeat length associations were detected for any factor. Among females, only a significant association with repeat length and self-report attention (p < 0.01) was detected, with premutation carriers self-reporting significantly more attention-related problems compared to noncarriers. No significant interactions between repeat length and age were detected. Overall, these results indicate the lack of a global neuropsychological impact of carrying a premutation allele among adults under the age of 50.