LONG-TERM DEVELOPMENTAL CHANGES IN XANTHIUM INDUCED BY GIBBERELLIC ACID

One application of gibberellic acid (GA₃) to Xanthium shoots resulted in an initial large stimulation, followed by inhibition, of internode elongation. After presumed translocation of the hormone from the locus of its application to the stem apex several morphological changes were observed. There was a significant increase in number of mitotic figures in the apical meristem and a twofold increase in volume of the apical dome. With time, the rate of leaf production was accelerated about 1.8 times. The phyllotaxis of leaf primordia initiated under the influence of GA:, changed from a (2, 3) contact parastichy pattern in control shoot to a (3, 5) pattern. Final petiole length was smaller than the control, and the absolute rate of lamina expansion decreased under prolonged treatment. Gibberellic acid had a pronounced effect on leaf morphology. GA_a induced the development of lanceolate leaves instead of typical deltoid leaves. The reduction in leaf area coincided with a 32% reduction in the average area of epidermal cells. Plastochron changes were correlated with anatomical and morphological changes during the course of leaf development.     

Genetic biodiversity in the Baltic Sea: species-specific patterns challenge management

Information on spatial and temporal patterns of genetic diversity is a prerequisite to understanding the demography of populations, and is fundamental to successful management and conservation of species. In the sea, it has been observed that oceanographic and other physical forces can constitute barriers to gene flow that may result in similar population genetic structures in different species. Such similarities among species would greatly simplify management of genetic biodiversity. Here, we tested for shared genetic patterns in a complex marine area, the Baltic Sea. We assessed spatial patterns of intraspecific genetic diversity and differentiation in seven ecologically important species of the Baltic ecosystem—Atlantic herring (Clupea harengus), northern pike (Esox lucius), European whitefish (Coregonus lavaretus), three-spined stickleback (Gasterosteus aculeatus), nine-spined stickleback (Pungitius pungitius), blue mussel (Mytilus spp.), and bladderwrack (Fucus vesiculosus). We used nuclear genetic data of putatively neutral microsatellite and SNP loci from samples collected from seven regions throughout the Baltic Sea, and reference samples from North Atlantic areas. Overall, patterns of genetic diversity and differentiation among sampling regions were unique for each species, although all six species with Atlantic samples indicated strong resistence to Atlantic-Baltic gene-flow. Major genetic barriers were not shared among species within the Baltic Sea; most species show genetic heterogeneity, but significant isolation by distance was only detected in pike and whitefish. These species-specific patterns of genetic structure preclude generalizations and emphasize the need to undertake genetic surveys for species separately, and to design management plans taking into consideration the specific structures of each species.     

Species turnover in ant assemblages is greater horizontally than vertically in the world's tallest tropical forest

Abiotic and biotic factors structure species assembly in ecosystems both horizontally and vertically. However, the way community composition changes along comparable horizontal and vertical distances in complex three‐dimensional habitats, and the factors driving these patterns, remains poorly understood. By sampling ant assemblages at comparable vertical and horizontal spatial scales in a tropical rainforest, we tested hypotheses that predicted differences in vertical and horizontal turnover explained by different drivers in vertical and horizontal space. These drivers included environmental filtering, such as microclimate (temperature, humidity, and photosynthetic photon flux density) and microhabitat connectivity (leaf area), which are structured differently across vertical and horizontal space. We found that both ant abundance and richness decreased significantly with increasing vertical height. Although the dissimilarity between ant assemblages increased with vertical distance, indicating a clear distance‐decay pattern, the dissimilarity was higher horizontally where it appeared independent of distance. The pronounced horizontal and vertical structuring of ant assemblages across short distances is likely explained by a combination of microclimate and microhabitat connectivity. Our results demonstrate the importance of considering three‐dimensional spatial variation in local assemblages and reveal how highly diverse communities can be supported by complex habitats.     

ModifiComb, a new proteomic tool for mapping substoichiometric post-translational modifications, finding novel types of modifications, and fingerprinting complex protein mixtures

A major challenge in proteomics is to fully identify and characterize the post-translational modification (PTM) patterns present at any given time in cells, tissues, and organisms. Here we present a fast and reliable method ("ModifiComb") for mapping hundreds types of PTMs at a time, including novel and unexpected PTMs. The high mass accuracy of Fourier transform mass spectrometry provides in many cases unique elemental composition of the PTM through the difference DeltaM between the molecular masses of the modified and unmodified peptides, whereas the retention time difference DeltaRT between their elution in reversed-phase liquid chromatography provides an additional dimension for PTM identification. Abundant sequence information obtained with complementary fragmentation techniques using ion-neutral collisions and electron capture often locates the modification to a single residue. The (DeltaM, DeltaRT) maps are representative of the proteome and its overall modification state and may be used for database-independent organism identification, comparative proteomic studies, and biomarker discovery. Examples of newly found modifications include +12.000 Da (+C atom) incorporation into proline residues of peptides from proline-rich proteins found in human saliva. This modification is hypothesized to increase the known activity of the peptide.     

Lysosomal fusion and SNARE function are impaired by cholesterol accumulation in lysosomal storage disorders

The function of lysosomes relies on the ability of the lysosomal membrane to fuse with several target membranes in the cell. It is known that in lysosomal storage disorders (LSDs), lysosomal accumulation of several types of substrates is associated with lysosomal dysfunction and impairment of endocytic membrane traffic. By analysing cells from two severe neurodegenerative LSDs, we observed that cholesterol abnormally accumulates in the endolysosomal membrane of LSD cells, thereby reducing the ability of lysosomes to efficiently fuse with endocytic and autophagic vesicles. Furthermore, we discovered that soluble N‐ethylmaleimide‐sensitive factor attachment protein (SNAP) receptors (SNAREs), which are key components of the cellular membrane fusion machinery are aberrantly sequestered in cholesterol‐enriched regions of LSD endolysosomal membranes. This abnormal spatial organization locks SNAREs in complexes and impairs their sorting and recycling. Importantly, reducing membrane cholesterol levels in LSD cells restores normal SNARE function and efficient lysosomal fusion. Our results support a model by which cholesterol abnormalities determine lysosomal dysfunction and endocytic traffic jam in LSDs by impairing the membrane fusion machinery, thus suggesting new therapeutic targets for the treatment of these disorders.     

Identifying protein construct variants with increased crystallization propensity--a case study

This study describes an efficient multiparallel automated workflow of cloning, expression, purification, and crystallization of a large set of construct variants for isolated protein domains aimed at structure determination by X-ray crystallography. This methodology is applied to MAPKAP kinase 2, a key enzyme in the inflammation pathway and thus an attractive drug target. The study reveals a distinct subset of truncation variants with improved crystallization properties. These constructs distinguish themselves by increased solubility and stability during a parallel automated multistep purification process including removal of the recombinant tag. High-throughput protein melting point analysis characterizes this subset of constructs as particularly thermostable. Both parallel purification screening and melting point determination clearly identify residue 364 as the optimal C terminus for the kinase domain. Moreover, all three constructs that ultimately crystallized feature this C terminus. At the N terminus, only three amino acids differentiate a noncrystallizing from a crystallizing construct. This study addresses the very common issues associated with difficult to crystallize proteins, those of solubility and stability, and the crucial importance of particular residues in the formation of crystal contacts. A methodology is suggested that includes biophysical measurements to efficiently identify and produce construct variants of isolated protein domains which exhibit higher crystallization propensity.     

SSF production of lactic acid from cellulosic biosludges

The use of cellulosic biosludges generated in a Kraft pulp mill was investigated as substrate for lactic acid production by simultaneous saccharification and fermentation (SSF). The effect of the operation mode (batch or fedbatch), the initial liquid to solid ratio (12 or 30 g/g) and the nutrient supplementation (MRS components or none) on several parameters including lactic acid concentration, volumetric productivity and product yields, were evaluated. When the operation was carried out in fedbatch mode with nutrient supplementation and using a LSR(0)=12 g/g, a broth containing 42 g/L was obtained after 48 h with a volumetric productivity of 0.87 g/L h and a product yield of 37.8 g lactic acid/100 g biosludges. In a similar experiment carried out without nutrient supplementation, a lactic acid concentration of 39.4 g/L was obtained after 48 h with a volumetric productivity of 0.82 g/L h and a product yield of 35.5 g L-lactic acid/100 g biosludges.     

Microbial communities associated with phosphogenic sediments and phosphoclast‐associated DNA of the Benguela upwelling system

The processes that lead to the precipitation of authigenic calcium phosphate minerals in certain marine pore waters remain poorly understood. Phosphogenesis occurs in sediments beneath some oceanic upwelling zones that harbor polyphosphate‐accumulating bacteria. These bacteria are believed to concentrate phosphate in sediment pore waters, creating supersaturated conditions with respect to apatite precursors. However, the relationship between microbes and phosphorite formation is not fully resolved. To further study this association, we examined microbial community data generated from two sources: sediment cores recovered from the shelf of the Benguela upwelling region where phosphorites are currently forming, and DNA preserved within phosphoclasts recovered from a phosphorite deposit along the Benguela shelf. iTag and clone library sequencing of the 16S rRNA gene showed that many of our sediment‐hosted communities shared large numbers of phylotypes with one another, and that the same metabolic guilds were represented at localities across the shelf. Sulfate‐reducing bacteria and sulfur‐oxidizing bacteria were particularly abundant in our datasets, as were phylotypes that are known to carry out nitrification and the anaerobic oxidation of ammonium. The DNA extracted from phosphoclasts contained the signature of a distinct microbial community from those observed in the modern sediments. While some aspects of the modern and phosphoclast communities were similar, we observed both an enrichment of certain common microbial classes found in the modern phosphogenic sediments and a relative depletion of others. The phosphoclast‐associated DNA could represent a relict signature of one or more microbial assemblages that were present when the apatite or its precursors precipitated. While these taxa may or may not have contributed to the precipitation of the apatite that now hosts their genetic remains, several groups represented in the phosphoclast extract dataset have the genetic potential to metabolize polyphosphate, and perhaps modulate phosphate concentrations in pore waters where carbonate fluorapatite (or its precursors) are known to be precipitating.