Chemical nature of the egg shell in helminths. II. Mode of stabilization of egg shells of monogenetic trematodes

The chemical nature of the egg shells of the monogenetic trematodes Pseudomicrocotyle sp. and Pricea multae, parasitizing the gills of the marine fish Scomberomorus guttatus was studied with a view to understanding the mode of stabilization of the egg shells.The egg shells of the monogenetic trematodes appeared to contain dimers of tyrosine; a conclusion based on fluorescence properties, affinity to toluidine blue, methylene blue, and a positive test for aromatic amino acids. In addition, they are stabilized by -S-S- linkages as indicated by the positive reaction to tests for disulfides. This is also suggested by the shells being refractile to stain but which can be reversed after treatment with sodium thioglycollate.Absence of phenolic tanning in the egg shells of the monogenetic trematodes is indicated by the negative reaction to tests for quinones, the failure to develop color on incubation in catechol after heat treatment, and by the fluorescence in uv light.In these respects the egg shells of the monogenetic trematodes strongly resemble the egg shell of Fasciola hepatica.     

Polarity of the ascidian egg cortex before fertilization.

The unfertilized ascidian egg displays a visible polar organization along its animal-vegetal axis. In particular, the myoplasm, a mitochondria-rich subcortical domain inherited by the blastomeres that differentiate into muscle cells, is mainly situated in the vegetal hemisphere. We show that, in the unfertilized egg, this vegetal domain is enriched in actin and microfilaments and excludes microtubules. This polar distribution of microfilaments and microtubules persists in isolated cortices prepared by shearing eggs attached to a polylysine-coated surface. The isolated cortex is further characterized by an elaborate network of tubules and sheets of endoplasmic reticulum (ER). This cortical ER network is tethered to the plasma membrane at discrete sites, is covered with ribosomes and contains a calsequestrin-like protein. Interestingly, this ER network is distributed in a polar fashion along the animal-vegetal axis of the egg: regions with a dense network consisting mainly of sheets or tightly knit tubes are present in the vegetal hemisphere only, whereas areas characterized by a sparse tubular ER network are uniquely found in the animal hemisphere region. The stability of the polar organization of the cortex was studied by perturbing the distribution of organelles in the egg and depolymerizing microfilaments and microtubules. The polar organization of the cortical ER network persists after treatment of eggs with nocodazole, but is disrupted by treatment with cytochalasin B. In addition, we show that centrifugal forces that displace the cytoplasmic organelles do not alter the appearance and polar organization of the isolated egg cortex. These findings taken together with our previous work suggest that the intrinsic polar distribution of cortical membranous and cytoskeletal components along the animal-vegetal axis of the egg are important for the spatial organization of calcium-dependent events and their developmental consequences.     

Characterization and functions of the whitefly egg pedicel.

For the silverleaf whitefly, Bemisia argentifolii (Bellows and Perring) (Homoptera: Aleyrodidae), scanning and transmission electron microscopic techniques were used to observe the characteristics of egg oviposition into both plant cells/tissues and artificial membranes, and to document the morphology of mature egg pedicles removed from the ovaries of females. The exterior of the distal portion of the pedicel consisted of a tangled array of fibrous structures (0.2-0.3 microm in diameter) that constituted about 20-25% of the outer diameter of the pedicel. The attachments of the fibers to the core of the pedicle suggested that the pedicel functions as the collector and conduit for water (vapor), and perhaps solute movement into the egg. Silverleaf whitefly eggs on membranes were incubated at various levels of relative humidity and the eggs were scored for egg hatch. At 98-100% rh, the percentage egg hatch was 86-98%. At lower humidity ranges of 0-20, 55-65, and 75-85% rh, none of the eggs hatched. Media (solute) uptake by silverleaf whitefly egg pedicels was determined by exposing the pedicel side of eggs oviposited on membranes to media solutions containing the high molecular weight polysaccharide, [(14)C]-inulin. Solute uptake by the pedicel and movement into developing silverleaf whitefly eggs were demonstrated using [2-(14)C]-acetate, and assaying for radioactivity in hatched nymphs. These studies, using exposure of pedicels to relative humidity and radiolabeled materials, demonstrate that whitefly egg hatch is dependent upon water uptake by the pedicel, and that the pedicel has the ability to transport solutes into the developing egg.     

Ultrastructural studies on the surface membrane of the mouse egg.

Fertilized and unfertilized mouse eggs were examined by scanning and transmission electron microscopy for evidence of mosaicism in the organization and concanavalin A-binding properties of their surface membranes. No obvious quantitative mosaicism in concanavalin A binding was noted. The egg membrane was microvillous over most of its surface, but was smooth in the region overlying the 2nd metaphase spindle of the unfertilized egg and on the polar body of the fertilized egg.     

Mass fractionation of Drosophila egg chambers.

Egg chambers from Drosophila ovaries may now be fractionated in large quantities suitable for biochemical work. A suspension of isolated egg chambers is prepared by subjecting ovaries to digestion with 0.067% collagenase. Egg chambers are separated in size classes (which correspond to different developmental stages) by sieving them through a column of superimposed nylon nets. The method can, in principle, be used for the separation of any particles ranging upward from 10 μm in diameter.     

Magnetic anisotropy of egg lecithin membranes.

Magnetic realignment and rotational diffusion of cylindrical egg lecithin vesicles were measured under a phase contrast microscope. The anisotropy of magnetic susceptibility times membrane thickness was calculated from the data for several thin-walled vesicles. The resulting values were assigned to discrete numbers of bilayers. The difference between the susceptibilities parallel and perpendicular to the long axes of the lecithin molecules is deduced to be X parallel - X perpendicular = -(0.28 +/- 0.02) . 10(-8) cgs at 23 degrees C, if a bilayer thickness of 60 A is assumed.     

Nature of the thiamin-binding protein from chicken egg yolk.

A simple, rapid and efficient procedure for the purification of thiamin-binding protein from chicken egg yolk was developed. The method involved removal, by exclusion, of lipoproteins from DEAE-cellulose and subsequent elution of water-soluble proteins held on the ion-exchanger with 1 M-NaCl, followed by treatment of the eluted protein fraction with an aqueous suspension of dextran/charcoal to generate apoprotein from the holoprotein. The resultant protein fraction was subjected to bioaffinity chromatography on thiamin pyrophosphate--AE (aminoethyl)-Sepharose. The protein eluted specifically with 10 microM-thiamin at pH 7.0, was homogeneous by the criteria of polyacrylamide-gel disc electrophoresis, had a mol.wt. of 38 000 +/- 2000 and was not a glycoprotein. The purified thiamin-binding protein specifically interacted with riboflavin-binding protein with no detectable deleterious affect on its (14C)thiamin-binding capacity. The protein bound [14C]thiamin with a molar ratio of 1.0, with dissociation constant (Kd) 0.41 microM. This protein-ligand interaction was inhibited by thiamin analogues and antagonists. The absorption spectrum of the protein in the presence of thiamin exhibited significant hypochromism at the 278 nm band, indicating the involvement of aromatic amino acid residues of the protein, during its binding to the ligand. The protein cross-reacted with the monospecific antiserum to egg-white thiamin-binding protein, showing thereby that thiamin-binding proteins present in chicken egg yolk and white are the products of the same structural gene.