Fate of the sea urchin egg receptor for sperm following fertilization

The sea urchin egg receptor for sperm is a 350 kDa glycoprotein containing a large extracellular domain that contains the sperm binding site, a transmembrane domain and a short COOH- terminal intracellular domain. During oogenesis, the receptor protein is first detected in Golgi-associated vesicles and cortical granules. Not until the egg is mature does the receptor appear on the cell surface; at this stage the intact receptor is found in approximately equal quantities on the egg cell surface and in cortical granules. As a potentially unique type of receptor, we were interested in its fate following fertilization. Several techniques have revealed that, following sperm binding, the amount of receptor markedly decreases. Using western blot analysis as well as direct measurement of the receptor protein, it was found that the membrane-bound form of the receptor rapidly disappeared following sperm binding to the egg, with only 3% of the receptor remaining after 30 s. Analysis by immupoelectron microscopy revealed that 30 s after sperm binding, 30% of the initial level of receptor was present. This remaining 30% was found mostly within the perivitelline space formed by the raised fertilization envelope. The disparity between these two sets of results (i.e. 3 vs 30%) is most likely accounted for by the exocytosis of receptor molecules from cortical granules; this fraction of the receptor would have been lost during isolation of the membrane-bound form of the receptor. Thus, unlike other cell surface receptors, the sea urchin egg receptor for sperm is not endocytosed and recycled following ligand binding. Rather, it disappears, presumably as a result of proteolysis. Transiently, the cortical granule form of the receptor is found released into the perivitelline space where it may bind to sperm and thereby prevent polyspermy. Despite the apparent secretion of this form of the receptor, experiments with antibodies to the extracellular and intracellular domains indicate that the receptors in cortical granules and in the plasmic membrane are similar, if not identical.     

Ethylene biosynthesis in Regnellidium diphyllum and Marsilea quadrifolia

The pathway of ethylene biosynthesis was examined in two lower plants, the semi-aquatic ferns Regnellidium diphyllum Lindm. and Marsilea quadrifolia L. As a positive control for the ethylene-biosynthetic pathway of higher plants, leaves of Arabidopsis thaliana (L.) Heynh. were included in each experiment. Ethylene production by Regnellidium and Marsilea was not increased by treatment of leaflets with 1-aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene in higher plants. Similarly, ethylene production was not inhibited by application of aminoethoxyvinylglycine and -aminoisobutyric acid, inhibitors of the ethylene biosynthetic enzymes ACC synthase and ACC oxidase, respectively. However, ACC was present in both ferns, as was ACC synthase. Compared to leaves of Arabidopsis, leaflets of Regnellidium and Marsilea incorporated little [¹⁴C]ACC and [¹⁴C]methionine into [¹⁴C]ethylene. From these data, it appears that the formation of ethylene in both ferns occurs mainly, if not only, via an ACC-independent route, even though the capacity to synthesize ACC is present in these lower plants.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AdoMet S-adenosyl-l-methionine - AIB -aminoisobutyric acid - AVG aminoethoxyvinylglycine This research was supported by the U.S. Department of Energy through grant No. DE-FG02-91ER20021 and, in part, by a fellowship of the National Engineering and Research Council of Canada to Jacqueline Chernys.     

The observation of a hedamycin-d(CACGTG)2 covalent adduct by electrospray mass spectrometry

Covalent binding of the antitumour antibiotic hedamycin to the self-complementary hexadeoxyribonucleotide 5′-CACGTG-3′ has been investigated by electrospray ionization mass spectrometry (ESI-MS). Ions due to double-stranded forms of the free 5′-CACGTG-3′ and the hedamycin-5′-CACGTG-3′ adduct have been observed in ESI mass spectra and their identity has been confirmed by resolution of individual charge states in ESI-MS spectra. Clear evidence that specific base-paired associations are being observed in ESI-MS is provided by the results of a titration experiment involving alkylated and non-alkylated complementary strands. This work demonstrates the potential of this powerful new tool for studying ligand-DNA binding.     

Stream periphyton response to grazing and changes in phosphorus concentration

Grazing by the large caddisfly larva, Dicosmoecus gilvipes (Trichoptera; Limnephilidae), drastically reduced periphyton biomass in laboratory channels at a current velocity of 20 cm s^–1. Reduction in biomass as chl a and AFDW ranged from 88 to 93% and 82 to 85%, respectively. On average, grazing rate increased with in-channel SRP (soluble reactive phosphorus) content from 6 to 10 µg 1^–1. Grazing rates averaged 25.9–29.3 µg chl a m^–2 d^–1 and 10.8–12.2 µg chl a mg^–1 d^–1 based on area and grazer biomass, respectively, with most variability among treatments being due to the grazing effect. Grazing tended to shift the algal community increasingly to filamentous blue-green algae regardless of enrichment. After three weeks, Phormidium comprised over 61% of the community in grazed treatments but only 35% in ungrazed treatments. The stalked diatom Gomphonema comprised only 4% of the grazed community, but 11% in the three ungrazed channels with similar values for Scenedesmus. A model that includes grazing was calibrated to the data and produced a reasonable expectation of periphyton biomass over a range in SRP concentrations. While the model with constant grazer abundance predicts a gradually increasing grazed biomass as SRP increases, grazer production in natural streams may actually increase to accommodate the increased food production.     

Mutations of the Microsomal Triglyceride-Transfer–Protein Gene in Abetalipoproteinemia

Elevated plasma levels of apolipoprotein B (apoB)–containing lipoproteins constitute a major risk factor for the development of coronary heart disease. In the rare recessively inherited disorder abetalipoproteinemia (ABL) the production of apoB-containing lipoproteins is abolished, despite no abnormality of the apoB gene. In the current study we have characterized the gene encoding a microsomal triglyceride-transfer protein (MTP), localized to chromosome 4q22-24, and have identified a mutation of the MTP gene in both alleles of all individuals in a cohort of eight patients with classical ABL. Each mutant allele is predicted to encode a truncated form of MTP with a variable number of aberrant amino acids at its C-terminal end. Expression of genetically engineered forms of MTP in Cos-1 cells indicates that the C-terminal portion of MTP is necessary for triglyceride-transfer activity. Deletion of 20 amino acids from the carboxyl terminus of the 894-amino-acid protein and a missense mutation of cysteine 878 to serine both abolished activity. These results establish that defects of the MTP gene are the predominant, if not sole, cause of hereditary ABL and that an intact carboxyl terminus is necessary for activity.     

The biotransformation of methyl ent-15-oxokaur-16-en-19-oate by Rhizopus stolonifer and Mucor plumbeus

Incubation of methyl ent-15-oxokaur-16-en-19-oate with Rhizopus stolonifer and Mucor plumbeus gave methyl ent-7β,11-dihydroxy-15-oxokauran-19-oate and methyl ent-7β,16β-dihydroxy-15-oxokauran-19-oate, respectively.     

Statistical design of the effect of inoculum conditions on growth of hairy root cultures of Catharanthus roseus

Summary The factors in the inoculum conditions that influence the growth rate of hairy roots of Catharanthus roseus in liquid culture were investigated in a randomized two-level, three-factor statistical design. The three variables were the number of root tips, the length of root tips, and the initial volume of media. The experiments and analysis demonstrated the effect of inoculum conditions on hairy root growth, with the length of hairy root tips being the dominant variable without any clonal variability. The best growth rates (doubling times) were obtained with an inoculum of 5 hairy root tips, each 35–40 mm long in 50 mL media: 3.7 and 3.1 days for C. roseus hairy root clones LBE-6-1 and LBE-4-2, respectively.     

Infant development in two aquarium bottlenose dolphins

The behavioral development of two aquarium bottlenose dolphin calves, one male (born to a primiparous mother) and one female (born to a multiparous mother), was examined. From birth to 9 weeks of age, a total of 558.7 h of nursing and behavioral data were collected using focal sampling methods. Data from both calves revealed a pattern of increasing independence as they aged, but there were significant differences between the two calves. Suckling rates, mean suckles per bout, suckling bout rates, and bout durations for the male calf were higher than those for the female calf throughout the study. These frequencies declined significantly over time in the male's case but not in the female's case. Her suckling durations (lock-on time) were longer than his. The male spent more time in infant position (under the mother) than the female calf and less time swimming in echelon position (alongside the mother). Over time, the male spent less time in infant position, and the female calf spent more time socializing and less time echelon swimming. Both calves decreased the amount of time spent close to their mothers over the 9 week period, and the mothers decreased their role in proximity maintenance over time. We suggest that factors related to maternal primiparity caused the male calf to develop an effective compensatory response of frequent nursing and spending more time in infant position. Both calves survived and were healthy at the end of the observation period. © 1995 Wiley-Liss, Inc.