Isolation and characterization of plasma membrane-associated cortical granules from sea urchin eggs

Cortical granules, which are specialized secretory organelles found in ova of many organisms, have been isolated from the eggs of the sea urchins Arbacia punctulata and Strongylocentrtus pupuratus by a simple, rapid procedure. Electron micropscope examination of cortical granules prepared by this procedure reveals that they are tightly attached to large segments of the plasma membrane and its associated vitelline layer. Further evidence that he cortical granules were associated with these cell surface layers was obtained by (125)I-labeling techniques. The cortical granule preparations were found to be rich in proteoesterase, which was purified 32-fold over that detected in a crude homogenate. Similarly, the specific radioactivity of a (125)I-labeled, surface glycoprotein was increased 40-fold. These facts, coupled with electron microscope observations, indicate the isolation procedure yields a preparation in which both the cortical granules and the plasma membrane-vitelline layer are purified to the same extent. Gel electrophoresis of the membrane-associated cortical granule preparation reveals the presence of at least eight polypeptides. The major polypeptide, which is a glycotprotein of apparent mol wt of 100,000, contains most of the radioactivity introduced by (125)I-labeling of the intact eggs. Lysis of the cortical granules is observed under hypotonic conditions, or under isotonic conditions if Ca(2+) ion is present. When lysis is under isotonic conditions is induced by addition of Ca(2+) ion, the electron-dense contents of the granules remain insoluble. In contrast, hypotonic lysis results in release of the contents of the granule in a soluble form. However, in both cases the (125)I-labeled glycoprotein remains insoluble, presumably because it is a component of either the plasma membrane or the vitelline layer. All these findings indicate that, using this purified preparation, it should be possible to carry out in vitro studies to better define some of the initial, surface-related events observed in vivo upon fertilization.     

A protease-activatable luminescent biosensor and reporter cell line for authentic SARS-CoV-2 infection

Efforts to define serological correlates of protection against COVID-19 have been hampered by the lack of a simple, scalable, standardised assay for SARS-CoV-2 infection and antibody neutralisation. Plaque assays remain the gold standard, but are impractical for high-throughput screening. In this study, we show that expression of viral proteases may be used to quantitate infected cells. Our assays exploit the cleavage of specific oligopeptide linkers, leading to the activation of cell-based optical biosensors. First, we characterise these biosensors using recombinant SARS-CoV-2 proteases. Next, we confirm their ability to detect viral protease expression during replication of authentic virus. Finally, we generate reporter cells stably expressing an optimised luciferase-based biosensor, enabling viral infection to be measured within 24 h in a 96- or 384-well plate format, including variants of concern. We have therefore developed a luminescent SARS-CoV-2 reporter cell line, and demonstrated its utility for the relative quantitation of infectious virus and titration of neutralising antibodies.     

Highly syntenic regions in the genomes of soybean, <Emphasis Type="Italic">Medicago truncatula</Emphasis>, and <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Background  

Recent genome sequencing enables mega-base scale comparisons between related genomes. Comparisons between animals, plants, fungi, and bacteria demonstrate extensive synteny tempered by rearrangements. Within the legume plant family, glimpses of synteny have also been observed. Characterizing syntenic relationships in legumes is important in transferring knowledge from model legumes to crops that are important sources of protein, fixed nitrogen, and health-promoting compounds.     

Characterization and isolation of DNA microsatellite primers in Raja clavata L. (thornback ray,Rajidae)

The thornback ray, Raja clavata, is an elasmobranch (cartilaginous fish). Since the 1950s, its stock has severely declined. In order to investigate the genetic population structure of the species, we developed microsatellite loci. The five loci reported here have eight to 48 alleles per locus and display an observed heterozygosity ranging from 0.32 to 0.98 with no deviation from the Hardy–Weinberg equilibrium. In the test panel of 122 individuals from three populations, no null alleles, band‐stuttering, large allele dropouts or linkage disequilibrium was detected.     

Nocturne for an unknown pollinator: first description of a night‐flowering orchid (Bulbophyllum nocturnum)

Bulbophyllum nocturnum , a species of section Epicrianthes from New Britain, is described and illustrated. It is the first known example of an orchid species in which the flowers open after dark and close in the morning. The poorly understood pollination biology of section Epicrianthes, a clade with highly unusual flowers, is discussed. Attention is drawn to the close resemblance between the petal appendages of some species and the fruiting bodies of certain Myxogastria. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 167 , 344–350.     

Sucrose Breakdown in Relation to Fruit Growth of Acid Lime (Citrus aurantifolia)

The physiological capacity for sucrose breakdown in developingjuice sac cells of acid limes was estimated by assaying theactivity of the three enzymes of sucrose catabolism in additionto vacuolar acid hydrolysis. The maximum potential rates ofsucrose breakdown were compared with the observed rates of carbonutilization. Highest potential rates of sucrose breakdown (28.621mmol cm^–3 per hydrated active space d^–1) occurredat the initial stages of fruit development where carbon utilizationwas highest. As the fruit developed, the potential rates ofsucrose breakdown and carbon utilization declined to very lowlevels. At 80% of development, vacuolar acid hydrolysis becamethe only physiological mechanism for sucrose breakdown. Therelatively low amounts of sucrose hydrolysed by acid hydrolysisat this time were just sufficient to account for the measuredcarbon demands. The results suggest that carbon supplied bythis distinct sucrose catabolizing system is able to provideadequate levels of carbon skeletons for the observed levelsof respiration and dry weight deposition early in development,but becomes a limiting factor for growth in the later stages. Key words: Vacuolar acid hydrolysis, Citrus aurantifolia     

Positioning the nodule,the hormone dictum

The formation of a nitrogen-fixing nodule involves two diverse developmental processes in the legume root: infection thread initiation in epidermal cells and nodule primordia formation in the cortex. Several plant hormones have been reported to positively or negatively regulate nodulation. These hormones function at different stages in the nodulation process and may facilitate the coordinated development of the epidermal and cortical developmental programs that are necessary to allow bacterial infection into the developing nodule. In this paper, we review and discuss how the tissue specific nature of hormonal action dictates where, when and how a nodule is formed.Key words: nodulation, hormone regulation, epidermis, cortex