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Haloferax volcanii uses extracellular DNA as a source for carbon, nitrogen, and phosphorous. However, it can also grow to a limited extend in the absence of added phosphorous, indicating that it contains an intracellular phosphate storage molecule. As Hfx. volcanii is polyploid, it was investigated whether DNA might be used as storage polymer, in addition to its role as genetic material. It could be verified that during phosphate starvation cells multiply by distributing as well as by degrading their chromosomes. In contrast, the number of ribosomes stayed constant, revealing that ribosomes are distributed to descendant cells, but not degraded. These results suggest that the phosphate of phosphate-containing biomolecules (other than DNA and RNA) originates from that stored in DNA, not in rRNA. Adding phosphate to chromosome depleted cells rapidly restores polyploidy. Quantification of desiccation survival of cells with different ploidy levels showed that under phosphate starvation Hfx. volcanii diminishes genetic advantages of polyploidy in favor of cell multiplication. The consequences of the usage of genomic DNA as phosphate storage polymer are discussed as well as the hypothesis that DNA might have initially evolved in evolution as a storage polymer, and the various genetic benefits evolved later.
DNA damage in oocytes induces a switch of the quality control factor TAp63α from dimer to tetramer
(2011)
TAp63a, a homolog of the p53 tumor suppressor, is a quality control factor in the female germline. Remarkably, already undamaged oocytes express high levels of the protein, suggesting that TAp63a’s activity is under tight control of an inhibitory mechanism. Biochemical studies have proposed that inhibition requires the C-terminal transactivation inhibitory domain. However, the structural mechanism of TAp63a inhibition remains unknown. Here, we show that TAp63a is kept in an inactive dimeric state. We reveal that relief of inhibition leads to tetramer formation with ~20-fold higher DNA affinity. In vivo, phosphorylation-triggered tetramerization of TAp63a is not reversible by dephosphorylation. Furthermore, we show that a helix in the oligomerization domain of p63 is crucial for tetramer stabilization and competes with the transactivation domain for the same binding site. Our results demonstrate how TAp63a is inhibited by complex domain-domain interactions that provide the basis for regulating quality control in oocytes.
DNA methylation reader MECP2 : cell type- and differentiation stage-specific protein distribution
(2014)
Background: Methyl-CpG binding protein 2 (MECP2) is a protein that specifically binds methylated DNA, thus regulating transcription and chromatin organization. Mutations in the gene have been identified as the principal cause of Rett syndrome, a severe neurological disorder. Although the role of MECP2 has been extensively studied in nervous tissues, still very little is known about its function and cell type specific distribution in other tissues.
Results: Using immunostaining on tissue cryosections, we characterized the distribution of MECP2 in 60 cell types of 16 mouse neuronal and non-neuronal tissues. We show that MECP2 is expressed at a very high level in all retinal neurons except rod photoreceptors. The onset of its expression during retina development coincides with massive synapse formation. In contrast to astroglia, retinal microglial cells lack MECP2, similar to microglia in the brain, cerebellum, and spinal cord. MECP2 is also present in almost all non-neural cell types, with the exception of intestinal epithelial cells, erythropoietic cells, and hair matrix keratinocytes. Our study demonstrates the role of MECP2 as a marker of the differentiated state in all studied cells other than oocytes and spermatogenic cells. MECP2-deficient male (Mecp2−/y) mice show no apparent defects in the morphology and development of the retina. The nuclear architecture of retinal neurons is also unaffected as the degree of chromocenter fusion and the distribution of major histone modifications do not differ between Mecp2−/y and Mecp2wt mice. Surprisingly, the absence of MECP2 is not compensated by other methyl-CpG binding proteins. On the contrary, their mRNA levels were downregulated in Mecp2−/y mice.
Conclusions: MECP2 is almost universally expressed in all studied cell types with few exceptions, including microglia. MECP2 deficiency does not change the nuclear architecture and epigenetic landscape of retinal cells despite the missing compensatory expression of other methyl-CpG binding proteins. Furthermore, retinal development and morphology are also preserved in Mecp2-null mice. Our study reveals the significance of MECP2 function in cell differentiation and sets the basis for future investigations in this direction.
Bacterial autotransporters represent a diverse family of proteins that autonomously translocate across the inner membrane of Gram-negative bacteria via the Sec complex and across the outer bacterial membrane. They often possess exceptionally long N-terminal signal sequences. We analyzed 90 long signal sequences of bacterial autotransporters and members of the two-partner secretion pathway in silico and describe common domain organization found in 79 of these sequences. The domains are in agreement with previously published experimental data. Our algorithmic approach allows for the systematic identification of functionally different domains in long signal sequences. Keywords: bacterial autotransporter, sequence analysis, pattern, protein targeting, signal peptide, protein trafficking
Targeting signals direct proteins to their extra- or intracellular destination such as the plasma membrane or cellular organelles. Here we investigated the structure and function of exceptionally long signal peptides encompassing at least 40 amino acid residues. We discovered a two-domain organization ("NtraC model") in many long signals from vertebrate precursor proteins. Accordingly, long signal peptides may contain an N-terminal domain (N-domain) and a C-terminal domain (C-domain) with different signal or targeting capabilities, separable by a presumably turn-rich transition area (tra). Individual domain functions were probed by cellular targeting experiments with fusion proteins containing parts of the long signal peptide of human membrane protein shrew-1 and secreted alkaline phosphatase as a reporter protein. As predicted, the N-domain of the fusion protein alone was shown to act as a mitochondrial targeting signal, whereas the C-domain alone functions as an export signal. Selective disruption of the transition area in the signal peptide impairs the export efficiency of the reporter protein. Altogether, the results of cellular targeting studies provide a proof-of-principle for our NtraC model and highlight the particular functional importance of the predicted transition area, which critically affects the rate of protein export. In conclusion, the NtraC approach enables the systematic detection and prediction of cryptic targeting signals present in one coherent sequence, and provides a structurally motivated basis for decoding the functional complexity of long protein targeting signals.
Diatoms are thought to provide about 40% of total global photosynthesis and diatoms of the genus Coscinodiscus are an important, sometimes dominant, cosmopolitan component of the marine diatom community. The oomycete parasitoid Lagenisma coscinodisci is widespread in the northern hemisphere on its hosts in the genus Coscinodiscus. Because of its potential ecological importance, it would be a suitable pathogen model to investigate plankton/parasite interactions, but the species cannot be cultivated on media without its host, so far. Thus, it was the aim of this study to explore the potential of dual culture of host and pathogen in the laboratory and to optimise cultivation to ensure a long-term cultivation of the pathogen. Here, we report successful cultivation of a single spore strain of L. coscinodisci (Isla), on several Coscinodiscus species and strains, as well as the establishment of a cultivation routine with Coscinodiscus granii (CGS1 and CG36), which enabled us to maintain the single spore strain for more than 3 years in 6 cm Petri dishes and 10 ml tissue culture flasks. This opens up the opportunity to study the processes and mechanism in plankton/parasitoid interactions under controlled conditions.
Enzymes involved in tRNA maturation are essential for cytosolic, mitochondrial, and plastid protein synthesis and are therefore localized to these different compartments of the cell. Interestingly, only one isoform of tRNA nucleotidyltransferase (responsible for adding the 3′-terminal cytidine–cytidine–adenosine to tRNAs) has been identified in plants. The present study therefore explored how signals contained on this enzyme allow it to be distributed among the different cell compartments. It is demonstrated that the N-terminal portion of the protein acts as an organellar targeting signal and that differential use of multiple in-frame start codons alters the localization of the protein. Moreover, it is shown that the mature domain has a major impact on the distribution of the protein within the cell. These data indicate that regulation of dual localization involves not only specific N-terminal signals, but also additional factors within the protein or the cell.
Length variations of repetitive sequences in different AT-rich loop-coding regions of mitochondrial 16S rDNA in two gastropod species were discovered during intraspecific haplotype surveys. Examination of the discrete length variation of the basic repeat unit in a phylogenetic framework led to the conclusion of a microsatellite-like mutational dynamic. The observations suggest that the presence of a repetitive sequence structure alone is sufficient to trigger this dynamic.
Membranes are central for cells as borders to the environment or intracellular organelle definition. They are composed of and harbor different molecules like various lipid species and sterols, and they are generally crowded with proteins. The membrane system is very dynamic and components show lateral, rotational and translational diffusion. The consequence of the latter is that phase separation can occur in membranes in vivo and in vitro. It was documented that molecular dynamics simulations of an idealized plasma membrane model result in formation of membrane areas where either saturated lipids and cholesterol (liquid-ordered character, Lo) or unsaturated lipids (liquid-disordered character, Ld) were enriched. Furthermore, current discussions favor the idea that proteins are sorted into the liquid-disordered phase of model membranes, but experimental support for the behavior of isolated proteins in native membranes is sparse. To gain insight into the protein behavior we built a model of the red blood cell membrane with integrated glycophorin A dimer. The sorting and the dynamics of the dimer were subsequently explored by coarse-grained molecular dynamics simulations. In addition, we inspected the impact of lipid head groups and the presence of cholesterol within the membrane on the dynamics of the dimer within the membrane. We observed that cholesterol is important for the formation of membrane areas with Lo and Ld character. Moreover, it is an important factor for the reproduction of the dynamic behavior of the protein found in its native environment. The protein dimer was exclusively sorted into the domain of Ld character in the model red blood cell plasma membrane. Therefore, we present structural information on the glycophorin A dimer distribution in the plasma membrane in the absence of other factors like e.g. lipid anchors in a coarse grain resolution.
Déterminants de l’utilisation de Acacia auriculiformis comme bois d’œuvre en Afrique de l’Ouest
(2020)
Acacia auriculiformis, un bois énergie, suscite de plus en plus des intérêts de bois d’œuvre au niveau des industriels de bois au Bénin. L’appréciation des performances de l’espèce dans les usines et en plantation est déterminante pour la vul- garisation de l’espèce comme alternative pour mitiger la déforestation en lien avec la demande en bois d’œuvre. L’objectif principal de ce travail est donc d’évaluer les conditions entourant l’adoption de Acacia auriculiformis comme espèce de bois d’œuvre au Bénin, Afrique de l’Ouest. Au total, 154 usines de bois et 25 plantations ont été enquêtées dans les zones abritant les plantations à A. auriculiformis. A. auriculiformis est l’espèce la plus fréquente dans les usines de bois (81%) suivie de Afzelia africana (55%), Tectona grandis (47%) et Khaya senegalensis (47%). Les superficies des plantations à A. auriculi- formis ont augmenté entre 1999 et 2019. Les connaissances sur l’utilisation de ce bois sont variables dans la zone d’étude. Le bois de A. auriculiformis est apprécié comme bois d’œuvre parce qu’il présente une couleur esthétique, un séchage rapide, une facilité de mise en œuvre, une imprégnabilité élevée, une densité moyenne à élevée et un bel aspect après mise en œuvre. Cependant, son bois fournit beaucoup de sciure, a beaucoup de nœuds et présente une déformabilité moyenne. Sa disponibili- té et son accessibilité sont les principaux facteurs justifiant la préférence de l’espèce par les industriels de bois d’œuvre. Cette forme d’utilisation de l’espèce est également remarquée au Togo, en Côte d’Ivoire. L’espèce présente une bonne perspective d’utilisation comme bois d’œuvre.