Variation in DNA-ploidy Levels of Reynoutria Taxa in the Czech Republic

The genus Reynoutria is represented by four taxa in the Czech Republic: Reynoutria japonica var. japonica, R. japonica var. compacta, R. sachalinensis and R. xbohemica. By using flow cytometry, cytological variability within the genus is described based on 257 Reynoutria samples. The varieties of R. japonica are cytologically uniform, var. japonica is exclusively octoploid (2n = 8x = 88) and var. compacta occurs only as a tetraploid (2n = 4x = 44), but R. sachalinensis and R. xbohemica exhibit some variation in chromosome numbers. Reynoutria sachalinensis is predominantly tetraploid (2n = 4 x = 44), but also occurs occasionally as hexaploid and octoploid cytotypes. The most common ploidy level in R. xbohemica is hexaploid (2n = 6x = 66), but tetraploid and octoploid clones were also found. The four taxa occurring in the Czech Republic are described briefly and the possible origins of the cytotypes discussed.     

Extracellular transduction events under pulsed stimulation in moth olfactory sensilla

In natural conditions, pheromones released continuously by female moths are broken in discontinuous clumps and filaments. These discontinuities are perceived by flying male moths as periodic variations in the concentration of the stimulus, which have been shown to be essential for location of females. We study analytically and numerically the evolution in time of the activated pheromone-receptor (signaling) complex in response to periodic pulses of pheromone. The 13-reaction model considered takes into account the transport of pheromone molecules by pheromone binding proteins (PBP), their enzymatic deactivation in the perireceptor space and their interaction with receptors at the dendritic membrane of neurons in Antheraea polyphemus sensitive to the main pheromone component. The time-averaged and periodic properties of the temporal evolution of the signaling complex are presented, in both transient and steady states. The same time-averaged response is shown to result from many different pulse trains and to depend hyperbolically on the time-averaged pheromone concentration in air. The dependency of the amplitude of the oscillations of the signaling complex on pulse characteristics, especially frequency, suggests that the model can account for the ability of the studied type of neuron to resolve repetitive pulses up to 2 Hz, as experimentally observed. Modifications of the model for resolving pulses up to 10 Hz, as found in other neuron types sensitive to the minor pheromone components, are discussed.     

Towards resolving the Knautia arvensis agg. (Dipsacaceae) puzzle: primary and secondary contact zones and ploidy segregation at landscape and microgeographic scales

Background and Aims

Detailed knowledge of variations in ploidy levels and their geographic distributions is one of the key tasks faced in polyploid research in natural systems. Flow cytometry has greatly facilitated the field of cytogeography by allowing characterization of ploidy levels at both the regional and population scale, and at multiple stages of the life cycle. In the present study, flow cytometry was employed to investigate the patterns and dynamics of ploidy variation in the taxonomically challenging complex Knautia arvensis (Dipsacaceae) and some of its allies (K. dipsacifolia, K. slovaca) in Central Europe.

Methods

DNA ploidy levels were estimated by DAPI flow cytometry in 5205 adult plants, 228 seedlings and 400 seeds collected from 292 Knautia populations in seven European countries. The flow cytometric data were supplemented with conventional chromosome counts. A subset of 79 accessions was subjected to estimation of the absolute genome size using propidium iodide flow cytometry.

Key Results and Conclusions

Five different ploidy levels (from 2x to 6x) were found, with triploids of K. arvensis being recorded for the first time. The species also exhibited variation in the monoploid genome size, corresponding to the types of habitats occupied (grassland diploid populations had larger genome sizes than relict and subalpine diploid populations). Disregarding relict populations, the distribution of 2x and 4x cytotypes was largely parapatric, with a diffuse secondary contact zone running along the north-west margin of the Pannonian basin. Spatial segregation of the cytotypes was also observed on regional and microgeographic scales. The newly detected sympatric growth of diploids and tetraploids in isolated relict habitats most likely represents the primary zone of cytotype contact. Ploidy level was found to be a major determinant of the strength of inter-cytotype reproductive barriers. While mixed 2x + 4x populations virtually lacked the intermediate ploidy level at any ontogenetic stage, pentaploid hybrids were common in 4x +6x populations, despite the cytotypes representing different taxonomic entities.Key words: Contact zone, cytogeography, flow cytometry, genome size, hybridization, Knautia arvensis, ploidy mixture, polyploidy, relict, reproductive isolation, serpentine     

Microsatellite mapping of the genes for brittle rachis on homoeologous group 3 chromosomes in tetraploid and hexaploid wheats

The brittle rachis character, which causes spontaneous shattering of spikelets, has an adaptive value in wild grass species. The loci Br1 and Br2 in durum wheat (Triticum durum Desf.) and Br3 in hexaploid wheat (T. aestivum L.) determine disarticulation of rachides above the junction of the rachilla with the rachis such that a fragment of rachis is attached below each spikelet. Using microsatellite markers, the loci Br1, Br2 and Br3 were mapped on the homoeologous group 3 chromosomes. The Br2 locus was located on the short arm of chromosome 3A and linked with the centromeric marker, Xgwm32, at a distance of 13.3 cM. The Br3 locus was located on the short arm of chromosome 3B and linked with the centromeric marker, Xgwm72 (at a distance of 14.2 cM). The Br1 locus was located on the short arm of chromosome 3D. The distance of Br1 from the centromeric marker Xgdm72 was 25.3 cM. Mapping the Br1, Br2 and Br3 loci of the brittle rachis suggests the homoeologous origin of these 3 loci for brittle rachides. Since the genes for brittle rachis have been retained in the gene pool of durum wheat, the more closely linked markers with the brittle rachis locus are required to select against brittle rachis genotypes and then to avoid yield loss in improved cultivars.     

Disruption of the mouse mTOR gene leads to early postimplantation lethality and prohibits embryonic stem cell development

The mammalian target of rapamycin (mTOR) is a key component of a signaling pathway which integrates inputs from nutrients and growth factors to regulate cell growth. Recent studies demonstrated that mice harboring an ethylnitrosourea-induced mutation in the gene encoding mTOR die at embryonic day 12.5 (E12.5). However, others have shown that the treatment of E4.5 blastocysts with rapamycin blocks trophoblast outgrowth, suggesting that the absence of mTOR should lead to embryonic lethality at an earlier stage. To resolve this discrepancy, we set out to disrupt the mTOR gene and analyze the outcome in both heterozygous and homozygous settings. Heterozygous mTOR (mTOR(+/-)) mice do not display any overt phenotype, although mouse embryonic fibroblasts derived from these mice show a 50% reduction in mTOR protein levels and phosphorylation of S6 kinase 1 T389, a site whose phosphorylation is directly mediated by mTOR. However, S6 phosphorylation, raptor levels, cell size, and cell cycle transit times are not diminished in these cells. In contrast to the situation in mTOR(+/-) mice, embryonic development of homozygous mTOR(-/-) mice appears to be arrested at E5.5; such embryos are severely runted and display an aberrant developmental phenotype. The ability of these embryos to implant corresponds to a limited level of trophoblast outgrowth in vitro, reflecting a maternal mRNA contribution, which has been shown to persist during preimplantation development. Moreover, mTOR(-/-) embryos display a lesion in inner cell mass proliferation, consistent with the inability to establish embryonic stem cells from mTOR(-/-) embryos.     

Cytochemical localization of GalNAc and GalNAcβ1,4Galβ1,4 disaccharide in mouse zona pellucida

Carbohydrate residues contained in the zona pellucida play a key role in the process of sperm-egg interaction. In vitro fertilization experiments have shown that a specific monoclonal antibody against GalNAcş,4Galş,4 disaccharide inhibits fertilization in mice. In the present study, the ultrastructural cytochemical localization of GalNAc residues and the GalNAcş,4Galş,4 disaccharide was carried out in ovarian and postovulatory oocytes by using lectin-gold cytochemistry and immunocytochemistry. Plant lectins SBA and DBA showed an affinity for the entire zona pellucida matrix of ovarian oocytes throughout the follicular maturation; however, immunoreactivity for GalNAcş,4Galş,4 disaccharide was not detected in ovarian oocytes at the earliest stages of follicular development but was found to be associated with the inner region of the zona matrix at the trilaminar primary follicle stage. The Golgi apparatus, vesicular aggregates, and cortical granules of the oocyte were intensely labeled by SBA and DBA throughout follicular development. Immunoreactivity to GalNAcş,4Galş,4 disaccharide was first observed in the Golgi apparatus and vesicular aggregates in trilaminar primary follicles. No immunoreactivity was observed in the cortical granules. In postovulatory oocytes, results were similar to those observed in ovarian oocytes. Our results thus suggest that (1) GalNAcş,4Galş,4 disaccharide residues are present only in the inner region of the zona pellucida and, therefore, might be involved in sperm penetration through the zona pellucida, (2) the inner and outer regions of the zona pellucida contain different oligosaccharide chains, (3) the vesicular aggregates detected in the oocyte could represent an intermediate step in the secretory pathway of zona pellucida glycoproteins and might be involved in the formation of cortical granules.     

Effect of oxygen on ubiquinone-10 production by Paracoccus denitrificans

Summary Ubiquinone-10 (Q₁₀) production was measured in batch cultures of Paracoccus denitrificans grown for 8 h at increasing oxygen concentrations (0–21 % O₂ in the sparging gas). Whereas the cellular level of Q₁₀ decreased monotonically from 1.2 to 0.5 mol/g d.w., the total yield of Q₁₀ was maximal at 2.5 % O₂ and amounted to 350 nmol (0.3 mg) per L of culture.     

A High-throughput-compatible FRET-based Platform for Identification and Characterization of Botulinum Neurotoxin Light Chain Modulators

Botulinum neurotoxin (BoNT) is a potent and potentially lethal bacterial toxin that binds to host motor neurons, is internalized into the cell, and cleaves intracellular proteins that are essential for neurotransmitter release. BoNT is comprised of a heavy chain (HC), which mediates host cell binding and internalization, and a light chain (LC), which cleaves intracellular host proteins essential for acetylcholine release. While therapies that inhibit toxin binding/internalization have a small time window of administration, compounds that target intracellular LC activity have a much larger time window of administrations, particularly relevant given the extremely long half-life of the toxin. In recent years, small molecules have been heavily analyzed as potential LC inhibitors based on their increased cellular permeability relative to larger therapeutics (peptides, aptamers, etc.). Lead identification often involves high-throughput screening (HTS), where large libraries of small molecules are screened based on their ability to modulate therapeutic target function. Here we describe a FRET-based assay with a commercial BoNT/A LC substrate and recombinant LC that can be automated for HTS of potential BoNT inhibitors. Moreover, we describe a manual technique that can be used for follow-up secondary screening, or for comparing the potency of several candidate compounds.     

Lasting the distance: The survival of alien birds shipped to New Zealand in the 19th century

Invasive alien species are a major threat to biodiversity and human activities, providing a strong incentive to understand the processes by which alien invasion occurs. While it is important to understand the determinants of success at each of several invasion stages—transport, introduction, establishment, and spread—few studies have explored the first of these stages. Here, we quantify and analyze variation in the success of individual animals in surviving the transport stage, based on shipping records of European passerines destined for New Zealand. We mined the original documents of Acclimatisation Societies, established in New Zealand for the purpose of introducing supposedly beneficial alien species, in combination with recently digitized newspaper archives, to produce a unique dataset of 122 ships that carried passerines from Europe to New Zealand between 1850 and 1885. For 37 of these shipments, data on the survival of individual species were available. Using generalized linear mixed models, we explored how survival was related to characteristics of the shipments and the species. We show that species differed greatly in their survival, but none of the tested traits accounted for these differences. Yet, survival increased over time, which mirrors the switch from early haphazard shipments to larger organized shipments. Our results imply that it was the quality of care received by the birds that most affected success at this stage of the invasion process.