AMPK-mediated phosphorylation on 53BP1 promotes c-NHEJ

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Validation of a high-throughput fermentation system based on online monitoring of biomass and fluorescence in continuously shaken microtiter plates

Background  

An advanced version of a recently reported high-throughput fermentation system with online measurement, called BioLector, and its validation is presented. The technology combines high-throughput screening and high-information content by applying online monitoring of scattered light and fluorescence intensities in continuously shaken microtiter plates. Various examples in calibration of the optical measurements, clone and media screening and promoter characterization are given.     

Generation of a chromosome-22-specific c-DNA library as confirmed by FISH analysis

We have recently developed a strategy for the rapid enrichment of c-DNA fragments from selected human chromosomes. Heteronuclear RNA (hn-RNA) is isolated from a somatic cell hybrid that retains a single human chromosome in a rodent background. Following c-DNA synthesis, human sequences are selectively amplified by the Alu polymerase chain reaction (Alu-PCR). Here we have applied this protocol for the selective isolation of novel c-DNAs encoded by chromosome 22. Fluorescence in situ hybridization has been used to confirm the chromosome-22-specific origin of the c-DNA fragments. Controls show DNAse-free RNase-treated hn-RNA results in no c-DNAs or Alu-PCR products. As demonstrated by competitive in situ suppression hybridization (CISS), the majority of the Alu-PCR products from hybrid GM 10027 are located on chromosome 22. Without competition, hybridization signals have also been identified on other human chromosomes. These unspecific hybridization signals result from Alu sequences and can successfully be reduced by competition with cot 1 DNA. This is the first report of the use of CISS for the localization of chromosome-specific c-DNAs.     

The deciduous dentition and dental replacement in the Eocene bat Palaeochiropteryx tupaiodon from Messel: The primitive condition and beginning of specialization of milk teeth among Chiroptera

The deciduous dentition and tooth replacement pattern of Palaeochiropteryx tupaiodon from the early Middle Eocene of Messel, near Frankfurt, Germany, are described. Ontogenetic states include fetuses to subadults. The posterior portion of the deciduous dentition (dP3-4) still shows the primitive eutherian condition of molarization, while the anterior part (dI-dC) was already engaged in the evolution of the highly derived condition found in living bats for clinging to the mother's fur. A styliform and sharp anterior dentition is considered a prerequisite in earliest chiropteran evolution. The greatly modified milk teeth of all living bats developed in different clades by parallel evolution under high selective pressure. The tiny and, at initial stages, poorly calcified teeth are substantiated by a newly developed microradiographic technique which is described in detail.     

Rewiring of purine metabolism in response to acidosis stress in glioma stem cells

Glioma stem cells (GSCs) contribute to therapy resistance and poor outcomes for glioma patients. A significant feature of GSCs is their ability to grow in an acidic microenvironment. However, the mechanism underlying the rewiring of their metabolism in low pH remains elusive. Here, using metabolomics and metabolic flux approaches, we cultured GSCs at pH 6.8 and pH 7.4 and found that cells cultured in low pH exhibited increased de novo purine nucleotide biosynthesis activity. The overexpression of glucose-6-phosphate dehydrogenase, encoded by G6PD or H6PD, supports the metabolic dependency of GSCs on nucleotides when cultured under acidic conditions, by enhancing the pentose phosphate pathway (PPP). The high level of reduced glutathione (GSH) under acidic conditions also causes demand for the PPP to provide NADPH. Taken together, upregulation of G6PD/H6PD in the PPP plays an important role in acidic-driven purine metabolic reprogramming and confers a predilection toward glioma progression. Our findings indicate that targeting G6PD/H6PD, which are closely related to glioma patient survival, may serve as a promising therapeutic target for improved glioblastoma therapeutics. An integrated metabolomics and metabolic flux analysis, as well as considering microenvironment and cancer stem cells, provide a precise insight into understanding cancer metabolic reprogramming.     

Metagenomic insights into soil microbial communities involved in carbon cycling along an elevation climosequences

Diversity and community composition of soil microorganisms along the elevation climosequences have been widely studied, while the microbial metabolic potential, particularly in regard to carbon (C) cycling, remains unclear. Here, a metagenomic analysis of C related genes along five elevations ranging from 767 to 4190 m at Mount Kilimanjaro was analysed to evaluate the microbial organic C transformation capacities in various ecosystems. The highest gene abundances for decomposition of moderate mineralizable compounds, i.e. carbohydrate esters, chitin and pectin were found at the mid-elevations with hump-shaped pattern, where the genes for decompositions of recalcitrant C (i.e. lignin) and easily mineralizable C (i.e. starch) showed the opposite trend (i.e. U-shaped pattern), due to high soil pH and seasonality in both low and high elevations. Notably, the gene abundances for the decompositions of starch, carbohydrate esters, chitin and lignin had positive relationships with corresponding C compounds, indicating the consistent responses of microbial functional profiles and metabolites to elevation climosequences. Understanding of adaptation of microbial communities, potential function and metabolites to elevation climosequences and their influencing factors provided a new insight for the regulation of terrestrial C storage.     

Dinoflagellate cyst biostratigraphy of the Opalinuston Formation (Middle Jurassic) in the Aalenian type area in southwest Germany and north Switzerland

Feist‐Burkhardt, S. & Pross, J. 2010: Dinoflagellate cyst biostratigraphy of the Opalinuston Formation (Middle Jurassic) in the Aalenian type area in southwest Germany and north Switzerland. Lethaia, Vol. 43, pp. 10–31. In order to provide a detailed dinoflagellate cyst stratigraphy of the Lower Aalenian Opalinuston Formation from the Aalenian type area, 68 samples from four boreholes and one outcrop section were analysed. The sample localities are Hausen an der Fils and Wittnau in southwest Germany, Weiach in north Switzerland and Mont Russelin in the Swiss Jura Mountains. Dinoflagellate cyst assemblages were recovered from the Late Toarcian Aalensis Zone to the Late Aalenian Murchisonae Zone. The samples yielded rich, well‐preserved and diverse assemblages with 51 dinoflagellate cyst taxa identified in total. The dinoflagellate cyst distribution data obtained from this study allow a high‐resolution biostratigraphical subdivision of the lowermost Middle Jurassic Opalinuston Formation into four palynostratigraphical units. First and last occurrences, acmes and consistent presence of the species Batiacasphaera sp. A, Evansia cf. granochagrinata, Kallosphaeridium praussii, Nannoceratopsis triangulata, Phallocysta? frommernensis and Wallodinium laganum were selected as the criteria for defining these units. The obtained high‐resolution palynostratigraphical scheme provides a basis for establishing and further refining early Middle Jurassic biostratigraphy in the Boreal and Tethyan realms. □Aalenian, biostratigraphy, dinoflagellate cysts, Germany, Jurassic, Switzerland, Toarcian.     

Analysis of the mitochondrial genome of Schistocerca gregaria gregaria (Orthoptera: Acrididae)

In the present study, we present the full sequence of the mitochondrial genome of the African desert locust Schistocerca gregaria gregaria. The size of 15625 bp reported matches very well with mitochondrial genomes of other Orthopteriodea. The mitochondrial genome comprises 13 protein‐coding genes, two ribosomal RNAs and 22 t‐RNAs with two t‐RNA (trnD and trnK) rearrangements that are typical for the taxon Caelifera. We compared the sequence with 12 mitochondrial genes of Schistocerca gregaria flaviventris and Schistocerca americana and used some of these data to construct phylogenetic trees, which confirm the close relationship between the two subspecies S. g. flaviventris and S. g. gregaria. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 296–305.     

Hydrogen postconditioning promotes survival of rat retinal ganglion cells against ischemia/reperfusion injury through the PI3K/Akt pathway

Retinal ischemia/reperfusion injury (IRI) plays a crucial role in the pathophysiology of various ocular diseases. Our previous study have shown that postconditioning with inhaled hydrogen (H₂) (HPC) can protect retinal ganglion cells (RGCs) in a rat model of retinal IRI. Our further study aims to investigate potential mechanisms underlying HPC-induced protection. Retinal IRI was performed on the right eyes of rats and was followed by inhalation of 67% H₂ mixed with 33% oxygen immediately after ischemia for 1?h daily for one week. RGC density was counted using haematoxylin and eosin (HE) staining, retrograde labelling with cholera toxin beta (CTB) and TUNEL staining, respectively. Visual function was assessed using flash visual evoked potentials (FVEP) and pupillary light reflex (PLR). The phosphorylated Akt was analysed by RT-PCR and western blot. The results showed that administration of HPC significantly inhibited the apoptosis of RGCs and protected the visual function. Simultaneously, HPC treatment markedly increased the phosphorylations of Akt. Blockade of PI3K activity by inhibitors (LY294002) dramatically abolished its anti-apoptotic effect and lowered both visual function and Akt phosphorylation levels.Taken together, our results demonstrate that HPC appears to confer neuroprotection against retinal IRI via the PI3K/Akt pathway.