Showing papers on "Nucleolus published in 1974"

Rna synthesis in the mouse oocyte

Abstract: RNA synthesis in the oocyte and granulosa cell nuclei of growing follicles has been studied in the mouse ovary. The RNA precursor [3H]uridine was administered intraperitoneally to adult mice and the amount of label incorporated into ovarian RNA was quantitated autoradiographically using grain-counting procedures. Uridine incorporation into the nucleus is low in oocytes of small, resting follicles but increases during follicle growth and reaches a peak prior to the beginning of antrum formation. Thereafter uptake rapidly declines and is very low in the oocytes of maturing follicles. Uridine incorporation into granulosa cell nuclei, in contrast to that found in the oocyte, increases gradually during most of the period of follicle growth. Qualitative studies of the activity of endogenous, DNA-dependent RNA polymerases have also been made in fixed oocytes isolated from follicles at different stages of growth. Polymerase activity is demonstrable in the nucleolus and nucleoplasm of oocytes from growing follicles, but is absent from maturing oocytes of large follicles.

Nucleolar and perichromosomal RNA synthesis during meiotic prophase in the mouse testis.

Abstract: The transcriptional activity during meiotic prophase in the mouse testis is studied with light microscopy and high-resolution autoradiographic techniques using [3H]uridine as a labeled precursor. In the present study, two types of RNA synthesis are detected during meiotic prophase: an extranucleolar RNA synthesis of perichromosomal localization and a nucleolar RNA synthetic activity. In some of the autosomes and close to the basal knobs, the activity of the nucleolar organizers is evidenced by the incorporation of [3H]uridine into nucleolar masses from zygotene on and at earlier labeling times. The evolution of nucleoli and the formation of a nucleolus attached to the sex pair are described during the different meiotic stages. Perichromosomal labeling, from leptotene on, reaches a maximum during middle pachytene and falls progressively to a low level at longer incorporation times. Sertoli's cell, the most active RNA synthetic cell in the seminiferous epithelium, rises to a maximum of labeling and drops at earlier times compared with the meiotic prophase cells. The condensed sex chromosomes show some scattered silver grains especially at middle pachytene. The axial chromosome cores and synaptonemal complexes are devoid of silver grains during the meiotic prophase. The observations suggest that a control mechanism operates during meiotic prophase to regulate transcriptional activity in the sex chromosomes and to provide differential RNA synthesis in autosomal bivalents at various stages of prophase and within certain segments of the chromosomes.

Transcription sites in spread meiotic prophase chromosomes from mouse spermatocytes.

Abstract: Mouse spermatocytes at pachytene stage have been examined by whole-mount electron microscope techniques complemented with autoradiography as an approach for visualizing their transcriptive activity. Structural elements of meiotic bivalents, such as synaptonemal complexes and chromatin fibers, have been satisfactorily displayed in the total set of autosomal and sexual bivalents in single spermatocytes. Adequate preservation of the entire set of bivalents has provided a basis for recognition of sites where presumptive preribosomal RNA and heterogeneous nuclear RNA species are being transcribed at different segments of autosomal bivalents. Nucleoli attached to the basal knob region where nucleolar organizer cistrons are assumed to be located and ribonucleoprotein fibrils associated with distinct chromatin loops have been recognized. These structural findings have been correlated with display of [3H]uridine incorporation sites in thin-section and whole-mount electron microscopy autoradiographic preparations. A low transcriptive activity of the sexual bivalent contrasted with extensive gene expression in autosomal bivalents. Each sex chromosome shows a double axial core. A short region of pairing with a synaptonemal complex joins the two chromosomes at one end. We conclude that variations in the rate of RNA synthesis throughout meiotic prophase stages in the mouse are expressed as fluctuations in the amount and distribution of distinct RNA species at specific segments of the bivalents.

The nucleolus in primary spermatocytes of Drosophila hydei.

Abstract: Transcribing ribosomal RNA genes in primary spermatocyte nucleoli of Drosophila hydei have been visualized by electron microscopy using a microspreading technique. The length of the transcribing unit is in agreement with the length of the ribosomal RNA precursor as determined by acrylamide gel electrophoresis (2.6×106 daltons). The length of the non-transcribed spacer is approximately 1.0×106 daltons. The maximum number of active cistrons in wild type (XY) males only approaches one half the estimated number of ribosomal cistrons of the replicated diploid genome (∼300) of D. hydei and is found to vary between 120 and 320 cistrons in different developmental stages of spermatocytes. There is also some variation between different males. In a few cases a variation in the transcriptional activity of different cistrons has been observed. The synthesis of ribosomal RNA therefore seems to be regulated primarily by the activation or inaetivation of varying numbers of ribosomal cistrons. Groups of adjacent cistrons seem to be under coordinated control. Inhibition of RNA synthesis by actinomycin, in contrast, follows a random pattern. The frequent observation of a bipartite nucleolus indicates that the nucleolus organizer regions of the two sex chromosomes are both active in transcription. The number of active ribosomal eistrons found in some X-Y translocation stocks and in XO males deviates considerably from that expected on the basis of DNA/RNA hybridization data. We conclude that, in agreement with the observations of other authors, a mechanism adjusting the number of ribosomal cistrons may be operating in these cases.

The genes for ribosomal RNA in diploid and polytene chromosomes of Drosophila melanogaster

Abstract: The proportion of the Drosophila genome coding for ribosomal RNA was examined in DNA from both diploid and polytene tissues of Drosophila melanogaster by rRNA-DNA hybridization. Measurements were made on larvae with one, two, three and four nucleolus organizer regions per genome. In DNA from diploid tissues the percent rDNA (coding for 28S and 18S ribosomal DNA) was found to be in proportion to the number of nucleolus organizers present. The number of rRNA genes within a nucleolus organizer therefore does not vary in response to changes in the number of nucleolus organizers. On the other hand, in DNA from cells with polytene chromosomes the percent rDNA remained at a level of about 0.1% (two to six times lower than the diploid values), regardless of either the number of nucleolus organizers per genome or whether the nucleolus organizers were carried by the X or Y chromosomes. This independence of polytene rDNA content from the number of nucleolus organizers is presumably due to the autonomous polytenization of this region of the chromosome. When the rDNA content of DNA from whole flies is examined, both the rDNA additivity of the diploid cells and the rDNA independence of polytene cells will affect the results. This is a possible explanation for the relative rDNA increase known to occur in X0 flies, but probably not for the phenomenon of rDNA magnification. — In further studies on DNA from larval diploid tissues, the following findings were made: 1) the Ybb-chromosome carries no rDNA; 2) flies carrying four nucleolus organizers do not tend to lose rDNA, even after eleven generations, and 3) the nucleolus organizer on the wild type Y chromosome may have significantly less rDNA than does that on the corresponding X chromosome.

Effects of 1,3-bis(2-chloroethyl)-1-nitrosourea and related compounds on nuclear rna metabolism.

Abstract: This study examines the effects of 1,3-bis(2-chloroethyl)-1-nitrosourea and certain related compounds on the synthesis and processing of nucleolar and nucleoplasmic RNA in cultured L1210 cells. Treatment with 0.25 mm 1,3-bis(2-chloroethyl)-1-nitrosourea has little effect on either quantity or size distribution of RNA synthesized during the first 10 min after drug addition. By 30 min, however, synthesis of both RNA fractions is markedly inhibited. By contrast, the processing of 45 S nucleolar (ribosomal precursor) RNA is inhibited immediately. The inhibition of nucleolar RNA processing is due to an action at some step after the normal synthesis and methylation of 45 S species. The processing or exit of high-molecular-weight RNA from the nucleoplasmic fraction also is inhibited. Structure-activity studies of some nitrosoureas indicated that inhibition of RNA processing requires a substitution at the N-3 nitrogen of the urea moiety. The isocyanate decomposition products of 1,3-bis(2-chloroethyl)-1-nitrosourea were found to be more effective than the parent compounds, suggesting that it is these isocyanate products that are responsible for the inhibition of RNA processing. In accord with this interpretation, 1-(2-chloroethyl)-1-nitrosourea, which lacks an N-3 nitrogen substituent and therefore cannot form an isocyanate product, was inactive. Since this nitrosourea is known to be highly active against experimental neoplasms, it is concluded that inhibition of RNA processing or other effects of isocyanate products are probably not directly responsible for the antitumor activity. These effects may instead be side effects, potentially related to toxicity.

THE ACTION OF α-AMANITIN ON RNA SYNTHESIS IN CHINESE HAMSTER OVARY CELLS: Ultrastructural and Biochemical Studies

Abstract: α-Amanitin acts in vitro as a selective inhibitor of the nucleoplasmic form B RNA polymerases. Treatment of Chinese hamster ovary (CHO) cells with this drug leads principally to a severe fragmentation of the nucleoli. While the ultrastructural lesions induced by α-amanitin in CHO cells and in rat or mouse liver are quite similar, the results diverge concerning the effect on RNA synthesis. It has been shown that in rat or mouse liver α-amanitin blocks both extranucleolar and nucleolar RNA synthesis. Our autoradiographic and biochemical evidence indicates that in CHO cells high molecular weight extranucleolar RNA synthesis (HnRNA) is blocked by the α-amanitin treatment, whereas nucleolar RNA (preribosomal RNA) synthesis remains unaffected even several hours after the inhibition of extranucleolar RNA synthesis. Furthermore, the processing of this RNA as well as its transport to the cytoplasm seem only slightly affected by the treatment. Finally, under these conditions, the synthesis of the low molecular RNA species (4–5S) still occurs, though less actively. The results are interpreted as evidence for a selective impairment of HnRNA synthesis by α-amanitin in CHO cells.

The action of α-amanitin in vivo on the synthesis and maturation of mouse liver ribonucleic acids

Abstract: α-Amanitin acts in vitro and in vivo as a selective inhibitor of nucleoplasmic RNA polymerases. Treatment of mice with low doses of α-amanitin causes the following changes in the synthesis, maturation and nucleocytoplasmic transfer of liver RNA species. 1. The synthesis of the nuclear precursor of mRNA is strongly inhibited and all electrophoretic components are randomly affected. The labelling of cytoplasmic mRNA is blocked. These effects may be correlated with the rapid and lasting inhibition of nucleoplasmic RNA polymerase. 2. The synthesis and maturation of the nuclear precursor of rRNA is inhibited within 30min. (a) The initial effect is a strong (about 80%) inhibition of the early steps of 45S precursor rRNA maturation. (b) The synthesis of 45S precursor rRNA is also inhibited and the effect increases from about 30% at 30min to more than 70% at 150min. (c) The labelling of nuclear and cytoplasmic 28S and 18S rRNA is almost completely blocked. The labelling of nuclear 5S rRNA is inhibited by about 50%, but that of cytoplasmic 5S rRNA is blocked. (d) The action of α-amanitin on the synthesis of precursor rRNA cannot be correlated with the slight gradual decrease of nucleolar RNA polymerase activity (only 10–20% inhibition at 150min). (e) The inhibition of precursor rRNA maturation and synthesis precedes the ultrastructural lesions of the nucleolus detected by standard electron microscopy. 3. The synthesis of nuclear 4.6S precursor of tRNA is not affected by α-amanitin. However, the labelling of nuclear and cytoplasmic tRNA is decreased by about 50%, which indicates an inhibition of precursor tRNA maturation. The results of this study suggest that the synthesis and maturation of the precursor of rRNA and the maturation of the precursor of tRNA are under the control of nucleoplasmic gene products. The regulator molecules may be either RNA or proteins with exceedingly fast turnover.

Localization of the 5S RNA genes in Zea mays by RNA-DNA hybridization in situ

Abstract: 5S RNA was extracted from Zea mays tissue and iodinated in vitro with 125I to a high specific activity. Acrylamide gel electrophoresis of the 125I-5S RNA, 11/2 weeks after iodination demonstrated that most of the 5S RNA molecules were degraded to half-size or smaller. In situ hybridization with this iodinated RNA to pachytene microsporocyte chromosomes showed that the 5S RNA cistrons are located near the end of the long arm of chromosome 2. No obvious association of the 5S locus with the nucleolus was seen during pachytene or later stages.

Induction of Nucleolar and Mitochondrial DNA Replication in Tetrahymena pyriformis

Abstract: Selective induction of replication of nucleolar and mitochondrial DNA has been demonstrated in starved-refed cultures of Tetrahymena pyriformis by different techniques. Labeling of starved cells with [(3)H]thymidine during a nutritional shift-up and analysis of the DNA in isopycnic CsCl gradients shows that the two initially labeled species of DNA are two species banding on the heavy and light side of the bulk macronuclear DNA. In isolated macronuclei the radioactivity is found only in the high density fraction, which has been shown to be of nucleolar origin. In sucrose gradients the newly replicated mitochondrial and nucleolar DNAs sediment considerably slower than the bulk DNA, as one discrete band corresponding to a molecular weight of about 3 to 4 x 10(7). Electron microscope autoradiography of cells labeled with [(3)H]thymidine as above shows that the peripheral nucleoli of the macronucleus as well as the mitochondria are labeled before any radioactivity is found in the chromatin granules of the macronucleus. The results clearly indicate that nucleolar and mitochondrial DNA replication are under a control independent of that for the replication of bulk DNA.

A Correlated Light- and Electron-Microscope Investigation of the Structural Evolution of the Nucleolus during the Cell Cycle in Plant Meristematic Cells (Allium Porrum)

Abstract: The interphase nucleolus consists of particulate zones and of a number of skein-like regions each of which contains a complex filamentous structure or nucleolonema. DNA is known to be present within the lacunar portions of these nucleolar regions. In the course of prophase the nucleolonemata become less tortuous and take the form of elongate structures extending across the nucleolar mass. Their continuity with the nucleolar organizing chromosomes can then be much more easily established than during interphase. When the peripheral granular portion of the late prophase nucleolus begins to disperse, the nucleolonemal lacunae appear smaller and less conspicuous. Examination of serial sections discloses that these convoluted filaments progressively contract during disorganization of the nucleolus but always remain less compact than the remaining portions of the nucleolar chromosomes. At very late prophase these chromosome loops have become quite small and have evolved into rather dense, homogeneous structures made up mostly of fibrillar material. By the time the nuclear envelope breaks down, these structures appear less compact under phase-contrast optics and, judging from their progressive contraction and their change in ultrastructural characteristics, it is evident that they have undergone extensive reorganization during the nucleolar dissolution process. This striking morphological evolution of the nucleolonemata is best interpreted by assuming that they lose a great deal of their constituent material during the later part of prophase and that the remaining portion, part of which is organizer DNA, retracts back to the chromosome axis and gives rise to nucleolar secondary constrictions. From metaphase to early telophase these constrictions are observed to consist of fine fibrillar material which stains much less intensely with heavy metals than other chromosome segments. When first detected as structured bodies, the late telophase nucleoli are roundish in shape and, under phase-contrast optics, their density closely matches that of the late prophase contracted nucleolonemata. At higher magnification incipient nucleoli are found to consist of a tight glomerulus predominantly fibrillar in texture. During the early G 1 period, growth of the nucleolus first results from expansion of the nucleolonemal skein and concurrent formation of numerous lacunar areas. Electron microscopy reveals that these lighter nucleolar zones contain diffuse fibrillar material similar to that observed in the lacunae of mature interphase nucleoli. Favourable preparations also show that this lacunar, fibrillar material is continuous with immediately adjacent segments of the nucleolar chromosomes. Concomitant with this expansion of the central skein region and enlargement of its lacunae, the growing nucleolus also gradually acquires an irregular layer of particulate material at its periphery. Since the formation of chromatin-containing lacunar regions and the appearance of peripheral RNP granules are found to coincide with a resumption in the RNA-synthesizing activity of the early interphase nucleous (authors9 unpublished observations), growth of this organelle appears to be partly dependent on derepression of its nucleolar loops. Mid- G 1 nucleoli exhibit all the morphological characterized of mature interphase organelles.

Rabbit Antibodies to Nucleoli of Novikoff Hepatoma and Normal Liver of the Rat

Abstract: Summary Antinucleolar antisera were produced in rabbits immunized with whole isolated nucleoli from normal rat liver and rat Novikoff hepatoma ascites cells. These antisera produced positive nucleolar fluorescence of varying degrees in nucleoli and nuclei of Novikoff tumor, rat liver, kidney, and Walker tumor cells. Novikoff hepatoma and liver antinucleolar antisera fixed complement when combined with 0.15 m NaCl-soluble proteins extracted from the hepatoma and liver nucleoli. Nucleolar specificity of the antibodies was demonstrated by inhibition of fluorescence or complement-fixation following pretreatment of the immune sera with whole nucleoli or nucleolar 0.15 m NaCl-soluble protein fractions.

Multiple gene sites for 5s and 18 + 28s rna on chromosomes of glyptotendipes barbipes (staeger)

Abstract: Ribosomal RNAs (28 + 18S and 5S) and 4S RNA extracted from the chironomid Glyptotendipes barbipes were iodinated in vitro with 125I and hybridized to the salivary gland chromosomes of G. barbipes and Drosophila melanogaster. Iodinated 18 + 28 S RNA labeled three puffed sites with associated nucleoli on chromosomes IR, IIL, and IIIL of G. barbipes and the nucleolar organizer of Drosophila. Labeled 5S RNA hybridized to three sites on chromosome IIIR, two sites on chromosome IIR and one site in a Balbiani ring on chromosome IV of Glyptotendipes. Most of the label produced by this RNA was localized seven bands away from the centromere on the right arm of chromosome III, and we consider this to be the main site complementary to 5S RNA in the chironomid. This same RNA preparation specifically labeled the 56 EF region of chromosome IIR of Drosophila which has been shown previously to be the only site labeled when hybridized with homologous 5S RNA. Hybridization of G. barbipes chromosomes with iodinated 4S RNA produced no clearly localized labeled sites over the exposure periods studied.

Neuronal perikarya with dispersed, single ribosomes in the visual cortex of Macaca mulatta.

Abstract: Numerous small and medium-sized neuronal perikarya in layers III and IV of the visual cortex display an unusual pattern of ribosomal distribution. Instead of being aggregated in clusters, spirals, rows, and other regular polysomal configurations, the ribosomes, whether free or attached to the endoplasmic reticulum, are randomly dispersed, with no discernible pattern. The endoplasmic reticulum in such cells is reduced to a few (perhaps only one) meandering, broad cisternae, which delimit broad fields of cytoplasmic matrix occupied almost solely by scattered, single ribosomes. The Golgi apparatus is elaborate. Mitochondria are either small and numerous or large and infrequent. The other organelles, including the nucleus and nucleolus, are not remarkable. Axonal terminals synapse in the normal fashion on the surfaces of these cells and their dendrites. Associated with these cells are more numerous intermediate cells in which a few to many polysomal clusters can be found. It is proposed that the neurons with dispersed, single ribosomes are inactive in protein synthesis and that the suspension of such an important metabolic activity is probably temporary. Thus, these cells are considered to be part of a population undergoing cyclic fluctuations in the intensity of protein synthesis that should be correlated with their specific neural behavior.

RNA Synthetic Activities of Sertoli Cells in the Mouse Testis

Abstract: Electron microscope autoradiographic studies of RNA synthesis in Sertoli cells of the adult mouse, using [‘H]uridine as a precursor, show a random distribution of grains in the nucleoplasmic area and a localized product on the nucleolar fibrillar component at early labeling times (15 mm). The condensed chromatin bodies associated with the nucleolus seem to have a nucleolar organizing function. At later labeling times (3 h) certain interchromatmn granules occurring in clusters in the nucleoplasm become partially labeled. After 7-8 days, the disappearance of label from Sertoli cells, contrasting with the longer persistence in spermatocytes, suggests a rapid turnover of RNA in Sertoli cells. Differential labeling patterns and structural changes in Sertoli cells cytoplasm are related to variations in functional activities presumably required for normal spermatogenesis and/or spermiogenesis. Sertoli cells are found in the seminiferous epithelium of several mammalian species. Though various functions have been attributed to these cells, it is generally accepted that Sertoli cells provide mechanical support for germinal cells. Moreover, they participate in the release of sperm into the lumen of the seminiferous tubules (VitaleCalpe and Burgos, 1970), dispose of cell debris by a phagocytic mechanism (Clegg and Macmillan, 1965; Kierszenbaum, 1970), and play an important role in the transfer of nutrients and metabolites between the intertubular space and the lumen of the tubules, acting as “bridge” cells (Vilar et al., 1962). It is also known that Sertoli cells constitute a unique stable cell population in the seminiferous epithelium. Whereas spermatogonia, spermatocytes, and sper

Inhibition of Rat-Liver RNA Polymerase in vitro by Aflatoxin B1 in the Presence of a Microsomal Fraction

Abstract: Aflatoxin B1 inhibits rat liver nucleoplasmic RNA polymerase B (40–50%) and cytoplasmic RNA polymerase C activities (25–35%) if applied in vivo. Nucleolar RNA polymerase A activity is not inhibited under the same conditions. Aflatoxin B1 has no effect on the activities of purified RNA polymerase enzymes A, B and C or on [3H]UTP incorporation of isolated rat liver nuclei in vitro. Aflatoxin B1, upon preincubation with a rat liver microsomal fraction, however, is apparently converted to a compound which then inhibits the activities of purified nucleoplasmic RNA polymerase (20–40%), cytoplasmic RNA polymerase (10–20%) and the incorporation of [3H]-UTP into isolated nuclei (38%). Nucleolar RNA polymerase activity is not affected under these conditions. The microsomal fraction, which most effectively converts aflatoxin B1 to an inhibitor of RNA polymerase is also the most effective cellular fraction catalysing the metabolism of aflatoxin B1.

Study of nucleolar vacuolation and RNA synthesis in embryonic root cells of Zea mays.

Abstract: The process of nucleolar vacuolation has been quantitatively studied in root cells of the embryo of Zea mays . During germination, we have estimated the percentage of vacuolated nucleoli, the number of vacuoles per nucleolus and the volume density of the vacuoles, the latter by application of the stereological principles of morphometry. In the dormant embryo, no vacuoles can be detected in the nucleolus. When germination occurs at 22 or 16 °C, there is a rapid increase in the percentage of vacuolated nucleoli and in the volume density (V A ), with a maximum after 4 and 8 h. At 6 °C, a temperature which does not permit emergence of the root, the percentage of vacuolated nucleoli increases regularly but never reaches the level observed at 16 and 22 °C. In parallel with the appearance of nucleolar vacuoles, the size of the nucleolus decreases statistically and shows a minimum when vacuolation is maximum. This suggests that the appearance of vacuoles within the nucleolus is the result of loss of material from this organelle. Indeed, electron micrographs show that granular components are lost from the nucleolus during the first 8 h of germination at 16 °C. According to the literature, these granular components are probably ribosomal subunits. An autoradiographic study of tritiated uridine incorporation shows that the nucleolus does not synthesize rRNA at the early beginning of germination. Hence the nucleolus is unable for several hours to rebuild the lost granular elements. Application of actinomycin D, 5-fluoro-uracil and 2-thio-uracil to the seeds, during the first 48 h of soaking, inhibits rRNA synthesis, the vacuolation process, the loss of granular components and the reduction in size of the nucleolus. These results allow us to determine some of the physiological conditions characterizing the appearance of the nucleolar vacuoles.

Are ‘N bands’ selective staining of specific heterochromatin?

Abstract: BY in situ DNA–RNA-hybridisation and the association patterns in meiotic and mitotic chromosomes, the nucleolus organiser regions (NOR) have been localised on specific chromosomal segments of some mammalian species1–4. Matsui and Sasaki5 described a staining technique (‘N bands’) which was assumed to stain the NOR of mammalian chromosomes with high specificity.

Ultrastructure of early development of mouse parthenogenones.

Abstract: Mouse eggs were parthenogenetically stimulated with hyaluronidase and osmotic shock and the resulting two-, four- and eight-cell stage embryos examined with the electron microscope. The ultrastructure of parthenogenones during cleavage showed numerous differences when compared with normal development. Cortical reaction was absent and there were no microvilli on the cell surface of developing blastomeres. The cytoplasm contained numerous bundles of fibrous material, vacuoles and lipid droplets, multivesicular bodies, and agglomerates of vesicles sometimes connected with cortical granules. Cortical granules and vacuolated mitochondria were numerous and usually situated near the cell membrane. The number of free ribosomes was very small and agglomerates of ribosomes or rough endoplasmic reticulum were not observed. Nucleoli in almost all cells were so-called primary nucleoli consisting of a dense, osmiophilic core and narrow ring of granular material. In only a few cells was reticulation of nucleoli and formation of fibrillar and granular parts noticed. Nuclear fragments and nuclei with broken membranes were present in some cells. Primary nucleoli persisted during mitosis and were in close contact with chromosomes. Cytokinesis was irregular and small and large fragments of cytoplasm surrounded by membrane were often observed between dividing cells. Our results suggested that failure of parthenogenones to develop normally is probably caused by numerous disturbances during early cleavage. Incomplete cortical reaction, persistence of primary nucleoli, a probable disturbance in RNA and protein synthesis, and irregularities during nuclear and cellular division were observed in all embryos.

A morphological and histochemical study of sertoli cells in normal and XX sex‐reversed mice

Abstract: Morphological and histochemical studies on the testes of mice with XX sex-reversal demonstrates several profound differences in the ultracytology and oxidative enzyme activity of Sertoli cells as compared to the normal. Membranous structures, cytoplasmic inclusions, and certain dehydrogenases in Sertoli cells of normal mice are described in relation to a possible role in steroid synthesis. In XX sex-reversed mice, two types of Sertoli cells are observed in the seminiferous tubules which are devoid of germ cells. One type is the randomly dispersed cells, which reveal the following ultrastructural alterations when compared with the normal cells: large spherical nuclei with prominent nucleoli, reduced smooth endoplasmic reticulum, pleomorphic mitochondria with no apparent cristae, and decreased lipid droplets. The other type is clustered cells which appear similar to immature Sertoli cells. These cells have small nuclei with poorly developed nucleoli, small profiles of rough endoplasmic reticulum, pleomorphic mitochondria, decreased lipid droplets, and an absence of specialized inter-Sertoli cell junctions. The results of this study suggest that functional and ultrastructural alterations of Sertoli cells in XX sex-reversed mice may result from degeneration or absence of germinal cells, and that steroidogenesis by Sertoli cells in mice may be associated with the development of germinal cells. It is also suggested that Sertoli cells may secrete a steroid which controls follicle-stimulating hormone secretion.

Variation in rDNA redundancy level and nucleolar organizer length in normal and variant lines of the Mexican axolotl.

Abstract: The level of redundancy of ribosomal genes, and the relationship of this level to nucleolar formation at different stages of embryonic development, have been examined in the Mexican axolotl, Ambystoma mexicanum . Individuals from 4 inbred stocks were examined, as well as descendants from 2 nucleolar variants which, in the heterozygous condition, are distinguished by exceptionally small nucleoli. Ribosomal RNA-DNA hybridization assays show that one of the 4 wild type lines has only about one-third as much ribosomal DNA (rDNA) as the other three. One of the nucleolar variants has the same level of rDNA as the larger wild-type level; the other variant has the same amount as the smaller ribosomal genome line. Both original nucleolar variants arose as F 1 progeny of crosses between a large rDNA genome line and the small genome line. Cytological examination of pregastrula stage embryos from wild type and nucleolar variant lines show that the lengths of the nucleolar organizer regions (NOR) and the sizes of nucleoli formed, are directly correlated with the amount of rDNA present at the nucleolar locus. During gastrulation of the nucleolar variants, however, a transition appears to take place and the amount of rDNA ceases to be the determining factor in nucleolar size. After late gastrula, heterozygous progeny resulting from crosses of either large rDNA genome or small rDNA genome wild type individuals with either nucleolar variant line, have a small and a large nucleolus. The factor or factors associated with this apparent lack of competitive ability of the variant NOR, when opposed to a normal NOR, are unknown. It might be suggested that since the chromosomal alterations which produced the nucleolar variants in both cases eliminated the gene determining the dark colour pattern, they could at the same time have eliminated other genetic material.

Synthesis of RNA in oocytes of Xenopus laevis during culture in vitro

Abstract: RNA synthesis can be maintained in large oocytes of Xenopus laevis during periods of in vitro culture of at least 10 days. A simple salt medium, modified Barth's solution, is found to be as effective a culture medium for these oocytes as several other complex media. The newly synthesized RNA is characterized electrophoretically and shown to consist predominantly of ribosomal RNA precursor, 28S and 18S ribosomal RNA, and 4S RNA. The distribution of this RNA within the oocyte is detected autoradiographically, where it is found to be greatly concentrated over the nucleoli. No qualitative alterations in either of these parameters are found during culture, within the limits of sensitivity of the assay procedures.

Modification of Sporulation in Yeast Strains with Two-Spored Asci (Saccharomyces, Ascomycetes)

Abstract: Electron-microscope observations indicate that the single nuclear division of sporulating yeasts which form 2-spored asci resembles, in several aspects, the nuclear behaviour during sporulation of the more common 4-spored yeasts. Specifically the parental nucleolus remains behind in the ascus cytoplasm, a characteristic nucleolus-associated spherical structure is formed, and the nucleus divides by budding rather than by elongation and medial constriction.