Ultrastructural studies of H-1 parvovirus replication. II. Induced changes in the deoxyribonucleoprotein and ribonucleoprotein components of human NB cell nuclei.

Ultrastructural changes in the nuclear DNP and RNP components of human NB cells induced by synchronous infection with H-1 parvovirus were studied using Bernhard's EDTA method of staining. Early events (12 h after infection) occurred in the nucleolus. Chromatin within the nucleolar fibrous centers condensed thereby converting the centers to vacuoles. DNP associated with the granular nucleolonema also contracted markedly, causing a disruption of this skein-like structure; it then migrated peripherally forming a heterochromatic cortex surrounding the granular nucleolar vestige. Subsequently (24–36 h after inoculation), condensation of extranucleolar chromatin took place concurrently with the accumulation of extensive amounts of interchromatin granules in the nucleus and cytoplasm. Conglomerates of perichromatin fibrils and interchromatin granules were frequently juxtapposed to the condensing chromatin. Large clumps of interchromatin granules were also closely associated with fragmenting nucleoli, and the apparent transformation of nucleolar granules into interchromatin granules was observed. Accumulation of H-1 protein on chromatin evidently fostered its condensation resulting in the pathology described.     

Structural and functional relationships between nuclear bodies and the nucleolus-DNA body complex in the oocyte of Amphiporus lactifloreus

The localization, structure and function of two types of nuclear bodies have been investigated by cytological and cytochemical electron microscopy methods in oocytes from the Hoplonemertean, Amphiporus lactifloreus. Type I nuclear bodies differentiate in contact with the nucleolus-DNA body complex, whereas type II nuclear bodies develop close to the diplotenic chromosomal axes. The structure of type I and type II spherical nuclear bodies, 4–5 μ m in width, results from the association of a fibrillar reticulum with some dense included regions. The cytochemical findings following the use of osmium-ammine reaction for DNA and silver reaction for NOR proteins support the hypothesis that type II nuclear bodies, derived from the extranucleolar area, as well as type I nuclear bodies, derived from the nucleolar complex, may be involved in ribosomal biogenesis.     

Cytochemical observations of the coiled bodies in neurons of rat sympathetic ganglia

Coiled bodies were investigated by means of ultrastructural cytochemistry. Preferential staining methods for localization of various proteins (ribonucleoproteins, basic proteins, phosphoproteins and glycoproteins) and DNA were applied. The results of cytochemical tests revealed that coiled bodies have a proteinaceous nature. They are composed of ribonucleoproteins, probably of nucleolar origin. They also contain phosphoproteins and glycoproteins but lack cytochemically detectable DNA. Coiled bodies present ultrastructural and cytochemical characteristics similar to the fibrillar part of the nucleous and to the interchromatin granules. The origin and possible functional role of coiled bodies are briefly discussed.     

Cytochemical observations of the coiled bodies in neurons of rat sympathetic ganglia

Summary Coiled bodies were investigated by means of ultrastructural cytochemistry. Preferential staining methods for localization of various proteins (ribonucleoproteins, basic proteins, phosphoproteins and glycoproteins) and DNA were applied. The results of cytochemical tests revealed that coiled bodies have a proteinaceous nature. They are composed of ribonucleoproteins, probably of nucleolar origin. They also contain phosphoproteins and glycoproteins but lack cytochemically detectable DNA. Coiled bodies present ultrastructural and cytochemical characteristics similar to the fibrillar part of the nucleous and to the interchromatin granules. The origin and possible functional role of coiled bodies are briefly discussed.     

The formation and cytochemical characterization of cortical granules in ovarian oocytes of the golden hamster (Mesocricetus auratus).

The formation and cytochemical characterization of cortical granules in the ovarian oocytes of the golden hamster have been investigated by use of light and electron microscopical techniques. Particular emphasis is given to the changing population of organelles associated with cortical granule formation. Our observations indicate that cortical granules are produced by the participation of both the Golgi complex and the rough endoplasmic reticulum. Ultrastructural cytochemistry reveals that the cortical granules are composed of glycoprotein. The cortical granules are released at fertilization by a merocrine-type of secretory process.     

Cortical granule-specific components are present within oocytes and accessory cells during sea urchin oogenesis

The coordinate expression of cortical granule-specific components in sea urchin oogenesis was studied using antibody probes. The components used to generate the organelle-specific antibodies included the whole cortical granule exudate, fertilization envelopes, hyalin, beta, 1-3,glucanase, and Ig8. Using immunolocalization techniques at both the light and electron microscopic levels, these molecules were found to be specific to cortical granules in three distinct cell types: developing oocytes, eggs, and accessory cells. In early oocytes, each of the cortical granule components are coordinately accumulated in the developing cortical granules dispersed throughout the cytoplasm. No other organelle within the developing oocytes or eggs contained detectable levels of any of these epitopes. In the somatic interstitial tissue of the ovary, cortical granule components also were accumulated specifically within cortical granule structures. Found only in select accessory cells, these cortical granules were indistinguishable in morphology and epitope composition from eggs and were contained within cytoplasmic aggregates termed mosaic globules. The mechanism of cortical granule concentration into mosaic globules is unknown, but this demonstration of common organelle constituents between oocytes and accessory cells may provide insight for such an understanding.     

Freeze-fracture analysis of structural reorganization during meiotic maturation in oocytes of Xenopus laevis

Summary During meiotic maturation, the cortex of oocytes of Xenopus laevis undergoes structural reorganization, visualized in this study by freeze-fracture electron microscopy. In the full-grown but immature oocyte, annulate lamellae are dispersed throughout the subcortex of the egg, 5 to 20 m from the plasma membrane. The annulate lamellae consist of well-organized stacks of membrane with visible pores. Stimulation of meiotic maturation by progesterone leads to disruption of the annulate lamellae and formation of an elaborate cortical endoplasmic reticulum which surrounds the cortical granules and intertwines throughout the cortex of the mature egg. Pore-like structures similar to those previously observed in the subcortical annulate lamellae are observed in the mature cortical endoplasmic reticulum. The cortical endoplasmic reticulum is often in close apposition with the plasma membrane and with membranes of cortical granules, but no junctions are visualized. This study provides further evidence that the cortical endoplasmic reticulum develops during progesterone-stimulated meiotic maturation in vitro, and that the annulate lamellae are precursors to the cortical endoplasmic reticulum.     

Biogenesis of small nucleolar ribonucleoproteins

Eukaryotic cells contain a very complex population of small nucleolar RNAs. They function, as small nucleolar ribonucleoproteins, in pre-ribosomal RNA processing reactions, and also guide methylation and pseudouridylation of ribosomal RNA, spliceosomal small nuclear RNAs, and possibly other cellular RNAs. Synthesis of small nucleolar RNAs frequently follows unusual strategies. Some newly discovered brain-specific small nucleolar RNAs of unknown function are encoded in introns of tandemly repeated units, expression of which is paternally imprinted. Recent studies of the protein components and factors participating in small nucleolar ribonucleoprotein assembly have revealed interesting connections with other classes of cellular ribonucleoproteins such as spliceosomal small nuclear ribonucleoproteins and telomerase. Cajal bodies emerge as nuclear structures important for the biogenesis and function of small nucleolar ribonucleoproteins.     

Starfish sperm-oocyte jelly binding triggers functional changes in cortical granules. A study using acid phosphatase and ruthenium red ultrastructural histochemistry

Starfish oocytes were examined before fertilization, immediately after insemination, and during the cortical reaction by means of acid phosphatase and ruthenium red ultrastructural histochemistry. Oocyte cortical granules are composed of a lamellar body and a surrounding matrix which is subdivided into dense and light portions. In unfertilized oocytes cortical granules are not stained by ruthenium red but show a weak acid phosphatase activity in the light portion of the granule matrix. Immediately after the adhesion of the spermatozoon to the oocyte jelly coat, the light matrix portion of cortical granules appears stained by ruthenium red and shows a strong acid phosphatase activity. During the cortical reaction, cortical granules are released into the perivitelline space and the lamellar body, surrounded by the stained matrix, fuses with the fertilization envelope. Our data suggest that membrane permeability changes and enzyme activation occur in the egg when the spermatozoon binds to the oocyte jelly coat.     

Ultrastructure of in vivo fertilization in the goat

In vivo fertilization of goat eggs has been studied by electron microscopy. Eggs were recovered from superovulated or natural cyclic goats, 32 to 52 hours after the onset of oestrus; only eggs recovered between 46 and 52 hours were fertilized. Spermatozoa penetrated the zona pellucida tangentially leaving vesiculated products of the acrosome reaction at the zona surface. As sperm penetrated into the ooplasm, the second meiotic division completed and cortical granule exocytosis occurred. However a few unreacted cortical granules usually remained in the cortex of the two fertilized eggs, adjacent to the plasma membrane. After swelling the two pronuclei presented similar ultrastructural morphology: they contained small, compact, agranular nucleoli and unevenly distributed chromatin. The cytoplasm in close vicinity to the apposed pronuclei contained large stacks of annulate lamellae, smooth endoplasmic reticulum, prominent Golgi complexes, as well as dense areas of unidentified material. The abundance of cytoplasmic organelles near the pronuclei might be the expression of intensive metabolic activity. Conversely, in the cortex of fertilized ova several large organelles-free cytoplasmic areas were randomly distributed.     

Starfish sperm-oocyte jelly binding triggers functional changes in cortical granules

Summary Starfish oocytes were examined before fertilization, immediately after insemination, and during the cortical reaction by means of acid phosphatase and ruthenium red ultrastructural histochemistry. Oocyte cortical granules are composed of a lamellar body and a surrounding matrix which is subdivided into dense and light portions. In unfertilized oocytes cortical granules are not stained by ruthenium red but show a weak acid phosphatase activity in the light portion of the granule matrix. Immediately after the adhesion of the spermatozoon to the oocyte jelly coat, the light matrix portion of cortical granules appears stained by ruthenium red and shows a strong acid phosphatase activity. During the cortical reaction, cortical granules are released into the perivitelline space and the lamellar body, surrounded by the stained matrix, fuses with the fertilization envelope. Our data suggest that membrane permeability changes and enzyme activation occur in the egg when the spermatozoon binds to the oocyte jelly coat.     

Perivitelline space of marsupial oocytes: Extracellular matrix of the unfertilized oocyte and formation of a cortical granule envelope following the cortical reaction

The purpose of this study was to characterize the structure of the vestments surrounding unfertilized and cortical granule-reacted oocytes from a marsupial, the grey short-tailed opossum Monodelphis domestica and to determine if a cortical granule envelope (CGE) forms in the perivitelline space (PVS) following the cortical reaction. Unfertilized oocytes collected from mature ovarian follicles and oviducal oocytes that had undergone a cortical reaction were fixed for electron microscopy in the presence of ruthenium red which stabilizes extracellular matrices (ECM) and facilitates demonstration of a CGE. Unfertilized oocytes were surrounded by a zona pellucida and had a PVS which contained a thick ECM comprised of granules and filaments. This matrix appeared to attach to the oolemma and was structurally similar to matrices reported previously in the PVS of unfertilized oocytes from eutherian mammals and two other marsupials, the Virginia opossum and the fat-tailed dunnart. The cortex of unfertilized oocytes contained cortical granules which were absent in oocytes recovered from the oviducts of mated females. Oviducal oocytes which lacked cortical granules exhibited a new coat within the PVS between the zona pellucida and the tips of the oocyte microvilli. This coat, the CGE, appeared structurally similar to CGEs described previously around fertilized eutherian oocytes. The CGE of the grey short-tailed opossum is approximately 1 μm thick and is made up of numerous small dense granules. The coats of the opossum oocyte are compared to those present around other marsupial and eutherian oocytes. © 1995 Wiley-Liss, Inc.     

Electron-microscopic study of silver staining of nucleoli in growing oocytes of rat ovaries

The nucleoli of dictyate-stage growing oocytes in rat ovaries were examined both with routine electron microscopy and electron microscopy after silver nitrate and ammoniacal silver nitrate (Ag-AS) staining. The nucleoli of the unilaminar follicular oocytes consist of twisted strands of dense fibrillar components, aggregates of granular components, and small fibrillar centers. After Ag-AS staining, silver grains are numerous on the dense fibrillar strands, fewer on the fibrillar centers, and very sporadic on the granular aggregates. The same stainability of three nucleolar components with the Ag-AS method was also confirmed in the nucleoli segregated by actinomycin D. During the transition of growing oocytes from bilaminar to plurilaminar follicle stage, the nucleolar dense fibrillar strands gradually conglomerate and are transformed into large and compact spherules. The stainability of dense fibrillar components with the Ag-AS method was lost along with this nucleolar transformation. These results may provide some new clues on the functional significance of Ag-AS-positive proteins in the nucleoli.