Browsing repeats in genomes: Pygram and an application to non-coding region analysis

Background  

A large number of studies on genome sequences have revealed the major role played by repeated sequences in the structure, function, dynamics and evolution of genomes. In-depth repeat analysis requires specialized methods, including visualization techniques, to achieve optimum exploratory power.     

Assembly of seed-associated microbial communities within and across successive plant generations

Background and aims

Seeds are involved in the transmission of microorganisms from one plant generation to another and consequently may act as the initial inoculum source for the plant microbiota. In this work, we assessed the structure and composition of the seed microbiota of radish (Raphanus sativus) across three successive plant generations.

Methods

Structure of seed microbial communities were estimated on individual plants through amplification and sequencing of genes that are markers of taxonomic diversity for bacteria (gyrB) and fungi (ITS1). The relative contribution of dispersal and ecological drift in inter-individual fluctuations were estimated with a neutral community model.

Results

Seed microbial communities of radish display a low heritability across plant generations. Fluctuations in microbial community profiles were related to changes in community membership and composition across plant generations, but also to variation between individual plants. Ecological drift was an important driver of the structure of seed bacterial communities, while dispersal was involved in the assembly of the fungal fraction of the seed microbiota.

Conclusions

These results provide a first glimpse of the governing processes driving the assembly of the seed microbiota.

     

Watershed ‘chemical cocktails’: forming novel elemental combinations in Anthropocene fresh waters

In the Anthropocene, watershed chemical transport is increasingly dominated by novel combinations of elements, which are hydrologically linked together as ‘chemical cocktails.’ Chemical cocktails are novel because human activities greatly enhance elemental concentrations and their probability for biogeochemical interactions and shared transport along hydrologic flowpaths. A new chemical cocktail approach advances our ability to: trace contaminant mixtures in watersheds, develop chemical proxies with high-resolution sensor data, and manage multiple water quality problems. We explore the following questions: (1) Can we classify elemental transport in watersheds as chemical cocktails using a new approach? (2) What is the role of climate and land use in enhancing the formation and transport of chemical cocktails in watersheds? To address these questions, we first analyze trends in concentrations of carbon, nutrients, metals, and salts in fresh waters over 100 years. Next, we explore how climate and land use enhance the probability of formation of chemical cocktails of carbon, nutrients, metals, and salts. Ultimately, we classify transport of chemical cocktails based on solubility, mobility, reactivity, and dominant phases: (1) sieved chemical cocktails (e.g., particulate forms of nutrients, metals and organic matter); (2) filtered chemical cocktails (e.g., dissolved organic matter and associated metal complexes); (3) chromatographic chemical cocktails (e.g., ions eluted from soil exchange sites); and (4) reactive chemical cocktails (e.g., limiting nutrients and redox sensitive elements). Typically, contaminants are regulated and managed one element at a time, even though combinations of elements interact to influence many water quality problems such as toxicity to life, eutrophication, infrastructure corrosion, and water treatment. A chemical cocktail approach significantly expands evaluations of water quality signatures and impacts beyond single elements to mixtures. High-frequency sensor data (pH, specific conductance, turbidity, etc.) can serve as proxies for chemical cocktails and improve real-time analyses of water quality violations, identify regulatory needs, and track water quality recovery following storms and extreme climate events. Ultimately, a watershed chemical cocktail approach is necessary for effectively co-managing groups of contaminants and provides a more holistic approach for studying, monitoring, and managing water quality in the Anthropocene.     

Ecophysiology of frailejones (<Emphasis Type="Italic">Espeletia</Emphasis> spp.), and its contribution to the hydrological functioning of páramo ecosystems

Páramos are high elevation tropical ecosystems in northern Andes, with large water yield and water regulation. One of the main and representative species growing in these páramos is the genus Espeletia, known as frailejones. There is a lack of knowledge of Espeletia ecophysiology, maybe due to its unusual anatomical modifications and the specific climatic conditions of these ecosystems. Therefore, it is important to determine the relationships between the anatomical modifications of Espeletia, its physiological functioning, and its contribution to the ecohydrologic functioning of páramos. Consequently, we studied the physiology of frailejones in two Colombian páramos, focused on the identification of conductive tissues inside the stems, calculated the age, and measured sapflow, using the heat ratio method. Results show that Espeletia spp. have a central pith that increases with height, as the size of secondary xylem decreases. Frailejones respond quickly to the changing conditions of weather factors controlling transpiration such as solar radiation, temperature, and fog presence. However, although environmental factors favor transpiration, the sapflow tends to decrease—a particular behavior of the Espeletia transpiration processes—since this occurs chaotically over time, including sapflow at night. The transformation of sapflow velocity to depth of water in a basin shows that the water lost through their transpiration is very low, which contributes to the high runoff ratio of páramo ecosystems. For the first time, we determine by radiocarbon the real ages of three E. hartwegiana, and their mean growth rates to range between 3.8 and 6.9 cm year^?1.     

Subcellular localization of secretogranin II and synaptophysin by immunoelectron microscopy in differentiated hypothalamic neurons in culture

Secretogranin II (SgII), a tyrosine-sulfated secretory protein, is a widespread component of endocrine and neuronal cells. In the present study we used mouse hypothalamic neurons differentiated in culture and studied the subcellular localization of SgII by two methods, i.e., by the use of immunoperoxidase or immunogold electron microscopy. By immunoperoxidase labeling, SgII was mainly detected in the matrix of large dense-core vesicles (LDCVs). In addition, usually in nerve terminals containing LDCVs, peroxidase reaction product was also found in association with the membrane of small synaptic vesicles (SSVs). By immunogold labeling, SgII was detected only in the matrix of LDCVs. We also compared the localization of SgII and synaptophysin (SY), an integral membrane protein of SSVs, by double labeling, using a combination of pre-embedding immunogold and -peroxidase techniques for SgII and SY, respectively. In perikarya, SgII-positive LDCVs were observed in the vicinity of the Golgi complex and scattered in the cytoplasm. In contrast, SY labeling was restricted to electron-translucent vesicles and tubular membranes in the Golgi area. Moreover, membrane structures positive for both SgII and SY were not found either in the Golgi zone or in other regions of the cytoplasm. In synaptic boutons, immunolabeling of LDCVs and SSVs with anti-SgII and anti-SY, respectively, was mutually exclusive. In summary, within the limitation of the methods used, our data are consistent with the notion that SgII and SY are segregated from each other on exit from the trans-Golgi network, than follow two distinct membrane traffic pathways, and that the presence of SgII on the membrane of some SSVs is due to endocytosis.     

Study of the potential spermatogonial stem cell compartment in dogfish testis, <Emphasis Type="Italic">Scyliorhinus canicula</Emphasis> L.

In the lesser-spotted dogfish (Scyliorhinus canicula), spermatogenesis takes place within spermatocysts made up of Sertoli cells associated with stage-synchronized germ cells. As shown in testicular cross sections, cysts radiate in maturational order from the germinative area, where they are formed, to the opposite margin of the testis, where spermiation occurs. In the germinative zone, which is located in a specific area between the tunica albuginea of the testis and the dorsal testicular vessel, individual large spermatogonia are surrounded by elongated somatic cells. The aim of this study has been to define whether these spermatogonia share characteristics with spermatogonial stem cells described in vertebrate and non-vertebrate species. We have studied their ultrastructure and their mitotic activity by 5′-bromo-2′-deoxyuridine (BrdU) incorporation and proliferating cell nuclear antigen (PCNA) immunodetection. Additionally, immunodetection of c-Kit receptor, a marker of differentiating spermatogonia in rodents, and of α- and β-spectrins, as constituents of the spectrosome and the fusome, has been performed. Ultrastructurally, nuclei of stage I spermatogonia present the same mottled aspect in dogfish as undifferentiated spermatogonia nuclei in rodents. Moreover, intercellular bridges are not observed in dogfish spermatogonia, although they are present in stage II spermatogonia. BrdU and PCNA immunodetection underlines their low mitotic activity. The presence of a spectrosome-like structure, a cytological marker of the germline stem cells in Drosophila, has been observed. Our results constitute the first step in the study of spermatogonial stem cells and their niche in the dogfish. G.L. is supported by a CIFRE grant (ANRT and C.RIS Pharma).     

A brain-penetrant triazolopyrimidine enhances microtubule-stability,reduces axonal dysfunction and decreases tau pathology in a mouse tauopathy model

Background

Alzheimer’s disease (AD) and related tauopathies are neurodegenerative diseases that are characterized by the presence of insoluble inclusions of the protein tau within brain neurons and often glia. Tau is normally found associated with axonal microtubules (MTs) in the brain, and in tauopathies this MT binding is diminished due to tau hyperphosphorylation. As MTs play a critical role in the movement of cellular constituents within neurons via axonal transport, it is likely that the dissociation of tau from MTs alters MT structure and axonal transport, and there is evidence of this in tauopathy mouse models as well as in AD brain. We previously demonstrated that different natural products which stabilize MTs by interacting with β-tubulin at the taxane binding site provide significant benefit in transgenic mouse models of tauopathy. More recently, we have reported on a series of MT-stabilizing triazolopyrimidines (TPDs), which interact with β-tubulin at the vinblastine binding site, that exhibit favorable properties including brain penetration and oral bioavailability. Here, we have examined a prototype TPD example, CNDR-51657, in a secondary prevention study utilizing aged tau transgenic mice.

Methods

9-Month old female PS19 mice with a low amount of existing tau pathology received twice-weekly administration of vehicle, or 3 or 10 mg/kg of CNDR-51657, for 3 months. Mice were examined in the Barnes maze at the end of the dosing period, and brain tissue and optic nerves were examined immunohistochemically or biochemically for changes in MT density, axonal dystrophy, and tau pathology. Mice were also assessed for changes in organ weights and blood cell numbers.

Results

CNDR-51657 caused a significant amelioration of the MT deficit and axonal dystrophy observed in vehicle-treated aged PS19 mice. Moreover, PS19 mice receiving CNDR-51657 had significantly lower tau pathology, with a trend toward improved Barnes maze performance. Importantly, no adverse effects were observed in the compound-treated mice, including no change in white blood cell counts as is often observed in cancer patients receiving high doses of MT-stabilizing drugs.

Conclusions

A brain-penetrant MT-stabilizing TPD can safely correct MT and axonal deficits in an established mouse model of tauopathy, resulting in reduced tau pathology.

     

Characterisation of the radish introgression carrying the Rfo restorer gene for the Ogu-INRA cytoplasmic male sterility in rapeseed (Brassica napus L.)

 Bulked segregant analysis and comparative mapping were applied to identify molecular markers linked to the Rfo restorer gene used for the Ogu-INRA cytoplasmic male-sterility system in rapeseed. These markers were then used to localise the radish introgression on the B. napus genetic map constructed from the cross ‘Darmor.bzh’ x ’Yudal’. The introgression mapped on the DY15 linkage group. From the comparison of this latter group to the linkage group constructed on a F₂ progeny segregating for the radish introgression, it was concluded that the introgression had occurred through homoeologous recombination, that it was not distal and that it had replaced a B. napus region of around 50 cM. A QTL involved in aliphatic seed glucosinolate content was located on the DY15 linkage group at a position corresponding to one end of the introgression. The DNA markers identified in this study are being used in map-based cloning of the Rfo gene and in marker-assisted selection. Received: 3 December 1997 / Accepted: 17 December 1997     

Biochemical characterization of the uptake and release of [3H]dopamine by dopaminergic hypothalamic neurons: a developmental study using serum-free medium cultures

The development of the biochemical properties of mouse hypothalamic dopaminergic neurons has been analyzed in vivo and in cultures of cell taken on the 16th day of gestation and grown in serum-free medium for up to 3 weeks. In the course of in vivo development, the dopamine (DA) content remains low during fetal life (10% of the adult value), beginning to increase on the 19th fetal day. In contrast, the specific accumulation of [3H]DA increased markedly during the last days of gestation from 20% of the adult value on the 16th fetal day to 70-80% of the adult value on Postnatal Day 3. Hypothalamic DA neurons in culture accumulate endogenous DA although at a lower level than in vivo. They take up [3H]DA by an active transport system which is specific for DA, and which shows time, temperature, and sodium dependency (Km = 1 microM). HPLC analysis showed that the newly taken up [3H]DA was not metabolized in the short run under the conditions used. It was stored in a form that could be released when neurons were depolarized in a high K+ (60 mM) medium. The K+-evoked [3H]DA release was found to be strictly dependent on extracellular Ca2+. Moreover the release of [3H]DA was also stimulated by veratridine in a Ca2+-dependent manner. Similar data have been obtained with the release of endogenous dopamine. No specific uptake and no K+-evoked dopamine release occurred in 2-day-old cultures. The specific [3H]DA uptake and the K+-evoked release appeared in 5-day-old cultures and increased with time in culture at least until Day 15. We examined the effects on [3H]DA release of polyunsaturated fatty acid, triiodothyronine, and corticosterone, all of which have been shown to play an important role in synaptogenesis in culture. These components, either separately or together, did not modify the percentage of the basal or the stimulated [3H]DA release. These results showed that hypothalamic DA neurons grown in serum-free medium progressively acquired the functional properties of adult DA neurons as concerns DA synthesis, DA uptake, and release. From a development point of view, this study suggests that the capacity to specifically take up [3H]DA and to respond to high K+ concentration is not expressed at early stages of neuronal development.(ABSTRACT TRUNCATED AT 400 WORDS)