Size‐based interactions and trophic transfer efficiency are modified by fish predation and cyanobacteria blooms in Lake Mývatn,Iceland

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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.

     

Streptacidiphilus jiangxiensis sp. nov., a novel actinomycete isolated from acidic rhizosphere soil in China

The taxonomic position of three acidophilic actinomycetes isolated from acidic rhizosphere soil was established using a polyphasic approach. The morphological and chemical properties of the isolates were found to be consistent with their assignment to the genus Streptacidiphilus. Almost complete 16S rRNA gene sequences determined for the strains were aligned with corresponding sequences of representatives of the genera Kitasatospora, Streptacidiphilus and Streptomyces and phylogenetic trees inferred using three tree-making algorithms. The organisms formed a distinct subclade within the Streptacidiphilus 16S rRNA gene tree. They also shared nearly identical phenotypic profiles and rep-PCR fingerprint patterns that readily distinguished them from representatives of the established species of Streptacidiphilus. It is evident from the genotypic and phenotypic data that the three isolates form a new species in the genus Streptacidiphilus. The name proposed for this new species is Streptacidiphilus jiangxiensis, the type strain is isolate 33214T (= AS 4.1857T = JCM 12277T).     

Genetic and environmental factors in the resistance of Drosophila subobscura adults to high temperature shock

Summary Survival time following a high temperature shock of Drosophila subobscura adults in dry air has shown great variability. This experimental condition involved desiccation as the first cause of death. Here survival is studied under saturation humidity, so that the mortality may be imputed only to thermic stress. We analyze the influence of culture temperature and crowding on resistance for different sex and age of the adults. The results show strong influences of these environmental factors on heat shock resistance and show interactions with the age and sex of the adults. We suggest that these facts could be due to acclimatization and/or to adaptation. The acclimatization would occur during development and would affect physiological processes related to aging of the flies. The adaptation would take place for selection, acting through differential mortality before the heat shock. Of course, other processes could be significant. Whatever the causal explanation, it will be necessary in any future research related with heat shock resistance to take these factors into account.     

Genetic and environmental factors in the resistance of Drosophila subobscura adults to high temperature shock

Summary We have carried out two equivalent selection experiments to increase and decrease heat shock resistance of Drosophila subobscura adults, using an indirect selection method that avoids excessive consanguinity. Heat shock was 33±0.5 °C at saturation humidity. Control lines showed a rapid change of the physiological trait as a consequence of laboratory culture conditions, expressed as a decrease both in heat shock resistance and in the initial population variability for heat shock resistance. Thus, this reduction of variability seems to consist in the loss of those combinations of genes that confer high resistance to heat shock. After eight generations of selection, the selected lines were differentiated from their respective control lines, and the selection response obtained was similar in resistant and sensitive lines. Differences in survival of progeny of reciprocal crosses between selected lines suggest that inheritance of heat resistance may depend in part on the origin of egg cytoplasm.     

Succination is Increased on Select Proteins in the Brainstem of the NADH dehydrogenase (ubiquinone) Fe-S protein 4 (Ndufs4) Knockout Mouse,a Model of Leigh Syndrome

Elevated fumarate concentrations as a result of Krebs cycle inhibition lead to increases in protein succination, an irreversible post-translational modification that occurs when fumarate reacts with cysteine residues to generate S-(2-succino)cysteine (2SC). Metabolic events that reduce NADH re-oxidation can block Krebs cycle activity; therefore we hypothesized that oxidative phosphorylation deficiencies, such as those observed in some mitochondrial diseases, would also lead to increased protein succination. Using the Ndufs4 knockout (Ndufs4 KO) mouse, a model of Leigh syndrome, we demonstrate for the first time that protein succination is increased in the brainstem (BS), particularly in the vestibular nucleus. Importantly, the brainstem is the most affected region exhibiting neurodegeneration and astrocyte and microglial proliferation, and these mice typically die of respiratory failure attributed to vestibular nucleus pathology. In contrast, no increases in protein succination were observed in the skeletal muscle, corresponding with the lack of muscle pathology observed in this model. 2D SDS-PAGE followed by immunoblotting for succinated proteins and MS/MS analysis of BS proteins allowed us to identify the voltage-dependent anion channels 1 and 2 as specific targets of succination in the Ndufs4 knockout. Using targeted mass spectrometry, Cys⁷⁷ and Cys⁴⁸ were identified as endogenous sites of succination in voltage-dependent anion channels 2. Given the important role of voltage-dependent anion channels isoforms in the exchange of ADP/ATP between the cytosol and the mitochondria, and the already decreased capacity for ATP synthesis in the Ndufs4 KO mice, we propose that the increased protein succination observed in the BS of these animals would further decrease the already compromised mitochondrial function. These data suggest that fumarate is a novel biochemical link that may contribute to the progression of the neuropathology in this mitochondrial disease model.We previously identified the formation of S-(2-succino)cysteine (2SC)¹ (protein succination) as a result of the irreversible reaction of fumarate with reactive cysteine thiols (1, 2). Fumarate concentrations are increased during adipogenesis and adipocyte maturation (2, 3), and the excess of glucose and insulin leads to augmented protein succination in the adipose tissue of type 2 diabetic mice (4, 5). Protein succination is also specifically increased in fumarate hydratase deficient hereditary leiomyomatosis and renal cell carcinoma (HLRCC), because of the decreased conversion of fumarate to malate (6, 7). In both cases, intracellular fumarate concentrations are elevated; in fumarate hydratase deficient cells, the fumarate concentration is about 5 mm (8), whereas fumarate levels increase up to fivefold in adipocytes grown in the presence of high (30 mm) versus normal (5 mm) glucose concentrations (2). In the adipocyte the increase in fumarate and succinated proteins develops as a direct result of mitochondrial stress induced by nutrient excess. Mechanistically, excess glucose without increased ATP demand inhibits the electron transport chain resulting in an elevated NADH/NAD⁺ ratio. This inhibits NAD⁺-dependent Krebs cycle enzymes and leads to an increase in fumarate and protein succination (9). In support of this we have also shown that low concentrations of chemical uncouplers of oxidative phosphorylation (OXPHOS) can decrease fumarate concentrations and protein succination (9). The physiological consequences of protein succination include a decrease in the functionality of the target protein (8, 10–12), for example succination of adiponectin prevents the formation of multimeric complexes and reduces plasma adiponectin levels in diabetes (4). Considering the impact of glucotoxicity driven mitochondrial stress in the adipocyte, we predicted that deficiencies in OXPHOS associated with NADH accumulation would also result in increased protein succination.Mitochondrial respiratory chain disorders encompass a broad range of encephalopathies and myopathies associated with the defective assembly, activity or maintenance of the OXPHOS machinery (13), and are estimated to occur in about 1 in 5,000 live births (14). A common feature in most mitochondrial diseases (MD) is a failure to thrive because of reduced mitochondrial energy production; both the brain and muscle are usually affected because of their high dependence on oxidative metabolism (13). Leigh syndrome is one of the most common manifestations of MD and is characterized by progressive neurodegeneration with bilateral necrotizing lesions of the brainstem and basal ganglia, resulting in lactic acidosis, ataxia, seizures, dystonia, and respiratory failure (15, 16). Mutations in genes encoding the five complexes of the OXPHOS machinery can lead to Leigh syndrome; however, the majority of these mutations affect subunits of complexes I and IV (17), and both mitochondrial and nuclear encoded proteins may be affected (17–19). Complex I is a large (980 kDa) l-shaped protein assembly consisting of 45 peptides, with one flavin mononucleotide and eight iron–sulfur clusters (20). One of the first identified mutations of complex I encoded Ndufs4, a small (18 kDa) assembly protein (21–23). Ndufs4 assists in the final stages of complex I assembly, and its absence results in the formation of a smaller ∼830 kDa subcomplex that lacks the NADH dehydrogenase module and has significantly less electron shuttling activity than the intact holoenzyme (24, 25). Ndufs4 mutations are associated with brainstem deterioration in humans (26), and a recently described Ndufs4 knockout mouse (Ndufs4 KO) exhibits many of the clinical and neurological symptoms observed in human Leigh syndrome (27, 28).One of the most common clinical features of MD is lactic acidosis, derived from the accumulation of pyruvate and elevated NADH. Increased lactate or lactate:pyruvate ratios have been measured in the blood, urine, and cerebrospinal fluid of a large number of Leigh syndrome patients (15, 16). Increases in other organic acids in urine have also been reported (16), indicating that metabolic acidosis is a prominent clinical feature. Interestingly, a study designed to find new diagnostic metabolites in MD demonstrated that within certain age ranges the measurement of urinary fumarate and malate was a more useful discriminator of MD than lactate or other organic acids (29). Barshop''s findings support the hypothesis that MD derived from OXPHOS deficiencies may exhibit increased protein succination because of the accumulation of NADH and subsequently fumarate. In this study we report for the first time that protein succination is present in the brain in an animal model of Leigh syndrome, the Ndufs4 KO mouse, suggesting that this modification may be an important biochemical link between the genetic defect and the onset of neuropathology observed in Leigh syndrome.     

GOLGI APPARATUS, GERL, AND LYSOSOMES OF NEURONS IN RAT DORSAL ROOT GANGLIA, STUDIED BY THICK SECTION AND THIN SECTION CYTOCHEMISTRY

New insights into the ultrastructure and phosphatase localizations of Golgi apparatus and GERL, and into the probable origin of lysosomes in the neurons of fetal dorsal root ganglia and the small neurons of adult ganglia have come from studying thick (0.5–1.0 µ) as well as thin (up to 500 A) sections by conventional electron microscopy. Tilting the thick specimens, by a goniometer stage, has helped to increase our understanding of the three-dimensional aspects of the Golgi apparatus and GERL. One Golgi element, situated at the inner aspect of the Golgi stack, displays thiamine pyrophosphatase and nucleoside diphosphatase activities. This element exhibits regular geometric arrays (hexagons) of interconnected tubules without evidence of a flattened portion (saccule or cisterna). In contrast, GERL shows acid phosphatase activity and possesses small cisternal portions and anastomosing tubules. Lysosomes appear to bud from GERL. Osmium deposits, following prolonged osmication, are found in the outer Golgi element. Serial 0.5-µ and thin sections of thiamine pyrophosphatase-incubated material demonstrate that, in the neurons studied, the Golgi apparatus is a continuous network coursing through the cytoplasm. Serial thick sections of acid phosphatase-incubated tissue suggest that GERL is also a continuous structure throughout the cytoplasm. Tubules of smooth endoplasmic reticulum, possibly part of GERL, extend into the polygonal compartments of the inner Golgi element. The possible physiological significance of a polygonal arrangement of a phosphatase-rich Golgi element in proximity to smooth ER is considered. A tentative diagram of the Golgi stack and associated endoplasmic reticulum in these neurons has been drawn.     

Genetic and environmental factors in the resistance of Drosophila subobscura adults to high temperature shock

Two replicate selection experiments to increase and decrease heat shock resistance of Drosophila subobscura adults were carried out maintaining control lines. In the present paper, the chromosomal-inversion and enzymatic polymorphism variation with selection is analyzed. The results indicate an erratic variation of chromosomal arrangement frequencies for practically all the chromosomes in the selected lines, showing a loss of the less frequent arrangements especially in sensitive lines. Only the A chromosome and the O+4 arrangement show a behaviour that may not be due to random effects, which points to the possible existence of heat shock factor(s) in these chromosomes. Similarly, an erratic variation of allele frequencies is observed for all the enzymes studied (Aph, Pept-1) except for the Hk-1 enzyme. We cannot establish the possible participation of this locus in heat shock resistance from the results obtained up to now. A significant decrease in heterozygosity is detected in sensitive lines from chromosomal-inversion polymorphism.