SLR1 function is dispensable for both self-incompatible rejection and self-compatible pollination processes inBrassica

TheSLR1 gene inBrassica is related both in DNA sequence and in pattern of expression to theS-locus glycoprotein (SLG) gene involved in the self-incompatibility mechanism which recognises and arrests the germination of self pollen. However,SLR1 shows minimal allelic variation and is expressed in both self-incompatible and compatibleBrassica lines and in related, self-compatible cruciferous plants. The function of the SLR1 protein is unknown. TheSLR1 gene was specifically ablated in self-incompatible and self-compatibleBrassica plants byAgrobacterium-mediated transformation with an antisense construct. Primary transformants and homozygous T2 progeny of both self-incompatibleB. oleracea and self-compatibleB. napus recipients were found to exhibit normal pollination responses despite having no detectable SLR1 glycoprotein. This shows that the high, wild-type level of SLR1 protein is not required to sustain the self-incompatibility reaction, nor is it necessary for successful intra-specific cross-pollination between compatible lines.     

Extracellular organic compounds from the ichthyotoxic red tide alga Heterosigma akashiwo elevate cytosolic calcium and induce apoptosis in Sf9 cells

Toxin(s) from the ichthyotoxic red tide alga Heterosigma akashiwo have been responsible for the destruction of millions of dollars of finfish aquaculture around the globe. Mechanisms of toxicity may include the production of reactive oxygen species (ROS) or organic toxins. The purpose of this study was to investigate the bioactivity of extracellular organic compounds from cultures of H. akashiwo. Cytosolic free calcium levels ([Ca²⁺]_i) in Spodoptera frugiperda (Sf9) insect cells infected with baculoviruses encoding the M1 muscarinic receptor were monitored.Exposure of cells to Heterosigma organics increased [Ca²⁺]_i up to 120 nM above basal levels (two-fold increase). Within minutes following exposure of the cells to the organics, the increase in [Ca²⁺]_i peaked and was followed by a slightly reduced, yet sustained plateau. This plateau was maintained for the duration of an experiment (>15 min) and was inhibitable by lanthanum. Furthermore, stimulation of Ca²⁺ release from intracellular stores by carbachol (muscarinic agonist) or thapsigargin (sarco-endoplasmic reticulum Ca²⁺-ATPases, SERCA inhibitor) potentiated the [Ca²⁺]_i response induced by the organics resulting in a maximal increase of >250 nM above basal levels (three-fold increase). However, the [Ca²⁺]_i response to Heterosigma organics was strictly dependent on the presence of extracellular calcium. Flow cytometric analyses revealed that these organics induced apoptosis of these same cells. Collectively, our data indicate that extracellular organics from cultures of H. akashiwo acutely increase [Ca²⁺]_i in cells by inhibiting the plasma membrane Ca²⁺-ATPase transporter and ultimately induce apoptotic cell death. These organics may play a significant role in the ichthyotoxic and allelopathic behaviour of this alga.     

Delineation of metabolic gene clusters in plant genomes by chromatin signatures

Plants are a tremendous source of diverse chemicals, including many natural product-derived drugs. It has recently become apparent that the genes for the biosynthesis of numerous different types of plant natural products are organized as metabolic gene clusters, thereby unveiling a highly unusual form of plant genome architecture and offering novel avenues for discovery and exploitation of plant specialized metabolism. Here we show that these clustered pathways are characterized by distinct chromatin signatures of histone 3 lysine trimethylation (H3K27me3) and histone 2 variant H2A.Z, associated with cluster repression and activation, respectively, and represent discrete windows of co-regulation in the genome. We further demonstrate that knowledge of these chromatin signatures along with chromatin mutants can be used to mine genomes for cluster discovery. The roles of H3K27me3 and H2A.Z in repression and activation of single genes in plants are well known. However, our discovery of highly localized operon-like co-regulated regions of chromatin modification is unprecedented in plants. Our findings raise intriguing parallels with groups of physically linked multi-gene complexes in animals and with clustered pathways for specialized metabolism in filamentous fungi.     

PHYSIOLOGICAL PROFILES OF SYNECHOCOCCUS (CYANOPHYCEAE) IN IRON-LIMITING CONTINUOUS CULTURES1

We examined the physiology and biochemistry associated with the iron-limited continuous culture of the halotolerant cyanobacterium Synechococcus PCC 7002. Biomass production, photosynthetic pigment levels, photosynthetic efficiency, and the production of hydroxamate- and catechol-type siderophores are reported for cells grown over a range of available iron concentrations. The relationship between the yield of Synechococcus PCC 7002 in iron-limited chemostats and the concentration of available iron was not linear. Synechococcus PCC 7002 expressed an inducible physiological response that led to alterations either in the cellular iron quotient or, more likely, in levels of available iron due to induced iron-scavenging processes. During iron limitation these cyanobacteria produced components consistent with the activation of a high-affinity iron transport system; both hydroxamate- and catechol-type siderophores were detected. Iron-limited Synechococcus PCC 7002 also reduced CO₂ fixation rates from luxury levels to a rate that matched the cellular growth rate, presenting interesting implications for oceanic carbon flux models.     

Combining SNP discovery from next-generation sequencing data with bulked segregant analysis (BSA) to fine-map genes in polyploid wheat

Background  

Next generation sequencing (NGS) technologies are providing new ways to accelerate fine-mapping and gene isolation in many species. To date, the majority of these efforts have focused on diploid organisms with readily available whole genome sequence information. In this study, as a proof of concept, we tested the use of NGS for SNP discovery in tetraploid wheat lines differing for the previously cloned grain protein content (GPC) gene GPC-B1. Bulked segregant analysis (BSA) was used to define a subset of putative SNPs within the candidate gene region, which were then used to fine-map GPC-B1.     

Extracellular organics from specific cultures of Heterosigma akashiwo (Raphidophyceae) irreversibly alter respiratory activity in mammalian cells

Raphidophyte blooms have been well documented in several coastal areas around the world. Centring raphidophyte-bloom research has been a focus evolving around issues of ichthyotoxicity, allelopathy and anti-predatory activity. However, the details of these phenomena such as the identity of the compounds and the mechanisms underlying these processes are poorly understood. One such raphidophyte, Heterosigma akashiwo (Hada) Hara et Chihara, has historically received much attention with regard to its ichthyotoxic and allelopathic properties. In this study, we collected extracellular organic compounds from cultures of nine H. akashiwo isolates and tested those exudates on two mammalian cell lines: rat osteoblastic sarcoma (UMR-106) and human embryonic kidney (HEK-293). A tetrazolium colourimetric assay was used to determine the activity of mitochondrial dehydrogenases. Exposure of the mammalian cell lines to exudates collected from cultures of H. akashiwo (strain 764) significantly increased activity in a concentration- and time-dependent manner. Exudate concentrations of as little as 0.3 mg ml⁻¹ elicited a stimulatory response in the mammalian cells. This is comparable to the range of exudate concentrations that were originally in the algal cultures (>0.1 mg ml⁻¹). Significant increases in activity were observed 12–24 h following continuous or 1 h (transient) exposure to the exudate. Production of the stimulatory bioactive exudate was not altered by nutrient-stressed H. akashiwo cultures (reduced iron, phosphate or nitrate). Collectively, these bioactive compound(s) consistently increased cellular activity 3–15-fold. Interestingly, of the nine isolates tested, four of them produced the stimulatory exudate, whereas four others did not produce the stimulatory compound(s) and isolate 560R produced a compound(s) that was inhibitory in nature. Thus, we have shown that cultures of H. akashiwo produce organic compounds that can alter the metabolic activity of mammalian cells. Future isolation and characterization of these bioactive compounds may determine them to have ecological relevance, potentially involved in the ichthyotoxic, allelopathic and/or anti-predatory behaviour of this alga.     

Fine structure of the phylogenetically important marine algaRhodogorgon carriebowensis (Rhodophyta,Batrachospermales?)

Summary The fine structure of the recently described red algaRhodogorgon carriebowensis J. Norris et Bucher was studied by light microscopy and scanning and transmission electron microscopy to aid in the ordinal placement of this unusual alga. Most significant in this context were findings that pit plugs had two-layered plug caps, the outer layer of which formed a large dome and was composed of glycoprotein. Cap membranes appeared to be absent. Medullary cells were remarkable in having extremely thick, layered cell walls, whose inward deposition left little room for cytoplasm. Medullary filaments branched little except near the base of the cortex. The assimilatory filaments that made up most of the cortex contained almost all the pigmentation and starch reserves of the thallus. These filaments terminated in either slender apical cells with smooth cell walls or bulbous apical cells bearing spinulose projections. Two types of elongated cells were found in the cortex, those with calcified cell walls and those surrounded by multiple, spreading layers of wall material. Neither cell type was pigmented. The latter type sometimes supported normal assimilatory cells.The hypothesis is proposed thatRhodogorgon is a descendant of the marine progenitors of the Batrachospermales and thus is a member of this order.Abbreviations DIC differential interference contrast - PAS periodic acid Schiff - PTA-CA phosphotungstic acid-chromic acid     

Characterization of filament forming Bacillus strains isolated from bulking sludge

Summary Filament-forming bacilli were isolated from bulking sludge. They were physiologically very similar. However, they developed into rhizoid or non-rhizoid colonies. According to their morphological, physiological, and genetical properties the isolates were identified as Bacillus mycoides and Bacillus cereus.