Showing papers on "Nucleolus published in 1972"

The interphase distribution of satellite DNA-containing heterochromatin in mouse nuclei.

Abstract: The distribution of sites capable of binding mouse satellite-complementary RNA in the cytological hybridization reaction has been examined in mouse liver and testis interphase nuclei. The approach taken has been to combine hybridization with semi-thin sectioning and autoradiography in order to obtain a clear picture of the relationship of satellite DNA-containing structures to the rest of the interphase nucleus. In liver nuclei, hybridization occurs primarily with blocks of heterochromatin associated with the nuclear envelope. The most prominent of these, in terms both of size and intensity of hybridization, is the nucleolar stalk and the rest of the nucleolus-associated heterochromatin. The nucleolar body itself is not labeled, nor is much of the peripheral condensed chromatin ; in fact, a polarized distribution of satellite DNA is evident. In Sertoli and spematid nuclei, satellite DNA is found in a small number of large heterochromatin blocks with which the nucleolus is associated; some of this material bears a relationship to the nuclear envelope in these cells also.

The nucleolus and its genes in amphibian oogenesis.

Abstract: Summary 1. Amphibian oocyte nucleoli may be solid and round, or they may take the form of beaded rings. Transformation of the nucleoli from the solid form to the ring form and back again may be a regular feature of oogenesis. The variation in circumference of freshly isolated unstretched ring nucleoli is continuous over a range of about fivefold. The longest rings may measure several hundred micra. All oocyte nucleoli have two structural zones, core and cortex. The core consists of finely fibrous material and is analogous to the ‘pars amorpha’ of earlier authors. The cortex has a granular fine structure and is analogous to the ‘nucleolonema’. The beads of ring nucleoli have both core and cortex components. Some solid nucleoli are multicored. 2. When all nucleoli are single cored, the number of nucleoli in an oocyte nucleus remains constant throughout the lampbrush phase of oogenesis. When nucleoli are multicored, the number of nucleoli may vary, but the number of nucleolar cores remains constant. 3. Oocyte nucleoli of between 6 and 12μ diameter have dry masses of between 50 and 200 μμg. 610% of the nucleolar dry mass is attributable to RNA. 4. Each nucleolus has a small DNA component. This component is in the form of a long circular duplex. In solid nucleoli it meanders through the substance of the nucleolar core. It forms the axis of a ring nucleolus and the structural integrity of the ring is maintained by the DNA component. 5. Both round and ring types of nucleoli from some salamanders and frogs have a membrane component attached to them. This takes the form of a membranous tube or a chain of vesicles ending near to or within a depression in the surface of the nucleolar core. Nothing is known at present of the function of the nucleolar membrane. 6. Nucleoli are sites of synthesis of ribosomal RNA. The DNA of the nucleolar core has the base sequence needed for transcription of ribosomal RNA. RNA is transcribed, complexed with protein and eventually distributed between 50S and 30S ribonucleoprotein particles. The 30S particle does not accumulate in the nucleolus. The 50S particle matures in the nucleolus and probably constitutes a substantial portion of the nucleolar cortex. 7. During the pachytene stage in ovaries of young amphibia 20–30 μμg. of extra-chromosomal nucleolar DNA are synthesized. In early diplotene this DNA disperses into thousands of tiny granules which become distributed over the inner surface of the nuclear envelope. A nucleolus then grows in association with each granule. 8. Isolation and purification of nucleolar DNA from pachytene oocytes, and from germinal vesicles, has shown that it contains the sequences for 18 and 28S ribosomal RNA, and it represents the products of an amplification of the genes for ribosomal RNA. 9. In Xenopus the gene complex which is amplified consists of about 450 tandem repeats of a region of double stranded DNA which includes sequences complementary to 28 S and 18 S ribosomal RNA alternating with a spacer sequence. The density of the ribosomal DNA, including the spacer DNA, in somatic cells of X. laevis is 1.7236 g. cm-3, and that of the oocyte amplified rDNA is 1.729. Both DNAs are 67% G + C. The somatic rDNA has 4.5% 5-methyl-deoxycytidylic acid which accounts for its lower density. The spacer sequences are uniform within a species, but differ from one species to another. 10. In oocytes, rDNA is amplified by using one or both of the chromosomal nucleolar organizers as a primary template. Amplification starts in oogonia and proceeds slowly until the pachytene stage. Rapid synthesis of DNA takes place in pachytene and early diplotene stages over a period of about 20 days. It seems likely that DNA synthesis proceeds in a cascade pattern such that each new replica itself serves as a template for further replication. Evidence for synthesis of a primary RNA template upon which the amplified rDNA is made is equivocal. 11. Whatever the size of the nucleolar organizer in an amphibian, and however many ribosomal cistrons it may incorporate, the amplification system in the oocyte is scaled accordingly, so that only a certain number of additional ribosomal cistrons and associated spacer are synthesized. Each of the eight germinal vesicles in an Ascuphus oocyte amplifies its rDNA to only one-eighth of the extent seen in the single oocyte nuclei from other anurans. Mononucleolate Xenopus, whose oocytes have only half the usual complement of chromosomal ribosomal DNA, nevertheless amplify to the level found in the normal wild-type individuals. Triploid Ambystoma, with hexaploid oocytes, appear to amplify their rDNA to three times the extent seen in their diploid relatives. It is clear that the level of gene amplification is related to the number of chromosomal organizers in the oocyte nucleus. It also seems that the level of amplification may be related to the volume of the oocyte nucleus. 12. The lampbrush chromosomes of at least some amphibia carry attached oocyte nucleoli at sites whose positions correspond to those of the somatic nucleolar organizers. The lampbrush nucleolar organizer in the axolotl is often marked by round or ring-like objects attached to the chromosome. 13. These lampbrush nucleolar organizers may carry solid or ring-like nucleoli. The attached nucleoli are thought to be permanently located there. They are not sequentially grown and detached from the chromosome. Attached ring or loop-like nucleoli may form ‘double bridges’, indicating that at least in some cases they are an integral part of the chromosome axis. In Plethodon the lampbrush nucleolar organizer consists of a long stretch of non-chromomeric axis that may appear single or double. There are ring or solid nucleoli attached at intervals along this axis. It is possible that these attached rings are products of amplification of the nucleolar organizer and that they became re-attached to the chromosomal organizer by a process analogous to recombination during or soon after the main phase of amplification.

Distinct transcription products of ribosomal genes in two different tissues.

Abstract: MOST organisms contain multiple copies of the genes which code for ribosomal RNA (rRNA), the number varying from about 160 to 28,000 per nucleus in different eukaryotic species1; these genes are clustered in the nucleolus. The repeating unit is a DNA sequence containing the structural genes for the 18S and 28S rRNA together with spacer DNA, only a part of which is transcribed2. Ribosomal RNA is transcribed from these genes as a polycistronic precursor molecule (pre-rRNA) which contains the rRNA sequences of the larger and smaller ribosomal subunits together with some additional sequences that are discarded during the maturation to rRNA3. The multiple gene copies are identical in the ribosomal regions within the limits detectable by present methods1, although there is some evidence that regions of non-transcribed spacer DNA vary in length and may therefore not all be identical2. We have suggested that the pre-rRNA may also be heterogeneous in molecular weight4.

Cytological localization of ribosomal cistrons in polytene chromosomes of Phaseolus coccineus

Abstract: Tritiated ribosomal RNA (rRNA) was prepared from hypocotyls of Phaseolus coccineus grown in liquid culture in the dark and in presence of 5-3H-uridine. A mixture of the 18S and 25S 3H-rRNA fractions was used for hybridization with DNA in the polytene chromosome cells of the embryo suspensor of P. coccineus. It was shown that the ribosomal cistrons (rDNA) are located in the nucleolus organizing system (satellite, nucleolar constriction and organizer) of the satellited chromosome pairs I (S1) and V (S2), in the proximal heterochromatic segment of the long arm of chromosomes S1 and in the terminal heterochromatic segment of chromosome pair II. The micronucleoli which are produced by the satellite and nucleolus organizer of the chromosome pair S1 contain rDNA; on the contrary, no rRNA-DNA hybridization is found in the DNA containing granules which are produced by the satellite and nucleolus organizer of chromosome pair S2. The DNA which is amplified during production of DNA puffs at some chromosomal regions apparently does not code for ribosomal RNA (no detectable rRNA-DNA hybridization).

Nucleolar behaviour in Triticum

Abstract: The maximum number of major nucleoli (macronucleoli) per nucleus of hexaploid, tetraploid and diploid wheat, Aegilops speltoides and Ae. squarrosa corresponded to the number of satellited chromosomes of each species. Smaller nucleoli (micronucleoli) were rare or absent in all of these species except the hexaploid, in which they were predominantly organized on chromosome arm 5Ds. — Fewer than the maximum number of macronucleoli in a mitotic interphase nucleus resulted from fusion of developing nucleoli. Enforced proximity of nucleolus-organizing regions resulted in more frequent fusion of nucleoli. — Analyses of related interphase nuclei showed that nucleoli, and hence probably chromosomes, undergo limited movement during mitotic interphase. These observations also indicate that specific chromosomes do not occupy specific sites in the interphase nucleus.

Evolution of the Nucleoli During Oogenesis in Xenopus Laevis Studied by Electron Microscopy

Abstract: Xenopus laevis tadpoles and toads were killed at several ages. The structure of the nuclei of the germinal cells has been observed by light and electron microscopes. We distinguish 11 successive stages in nucleolar structure: (1) a single, essentially granular nucleolus in the oogonium (10 µm diameter), (2) a reticulated nucleolus in the leptotene oocyte, (3) fragmentation of this nucleolus into a few smaller nucleoli, (4) multiple tiny nucleoli appearing in the cap of the pachytene oocyte, (5) enrichment in the fibrillar constituent of these intra-cap nucleoli, (6) grouped spherical nucleoli, with well segregated granular and fibrillar constituents, as the disintegration of the cap is going on (diplotene A oocyte, 30 µm diameter), (7) dispersion of those nucleoli in the nuclear sap (diplotene B oocyte, 50 µm diameter), (8) formation of long, ribboned nucleoli with multiple DNA-rich spots (diplotene C oocyte, 100 µm diameter), (9) fragmentation of the nucleolar ribbons into multiple spherical nucleoli with excentric fibrillar core and granular cortex (diplotene D oocyte, 150 µm diameter), (10) multiple purely fibrillar nucleoli (diplotene E oocyte, between 150 and 400 µm diameter), and (11) multiple classical nucleoli with concentric fibrillar core and granular cortex (diplotene F oocyte, between 400 and 1000 µm diameter). The multiplication of the nucleoli in Xenopus laevis may occur successively ( a ) by the fragmentation of the single oogonium nucleolus at the leptotene stage, ( b ) by de novo formation of nucleolar bodies inside the cap at the pachytene stage, and ( c ) by the growth of those nucleoli lying free in the nucleolar sap at the early diplotene stage. They evolve into nucleolar ribbons which later on fragment into spherical bodies. Four successive phases during the growth of an oocyte can be distinguished with respect to the ribosomal system. (I) The first phase is characterized by the nucleolar DNA amplification. (II) During the second phase, the multiplication of the nucleoli is going on. Ribosomes are present in the cytoplasm and the rate of cellular growth is very high. (III) During the third phase, the synthesis of rRNAs seem to be repressed while the synthesis of heterogenous small RNAs is going on. Ribosomes are no longer visible in the cytoplasm. The nucleoli are purely fibrillar. The rate of cell growth is lower than in the preceding phase. (IV) During the fourth ( = Duryee lampbrush stages 3-6), or vitellogenic phase, rRNAs are actively synthesized and numerous ribosomes appear in the cytoplasm. The nucleoli have the classical structure and the rate of growth is about the same as during phase III.

Functional organization of the nucleus in the oogenesis of chrysopa perla L. (insecta, neuroptera).

Abstract: On the basis of light, autoradiographic (uridine-3H incorporation) and electron microscopic investigation changes of nuclear structures were examined during the oogenesis in Chrysopa perla L. — In early meiotic prophase the oocyte nuclei were found to contain a large body of extrachromosomal DNA. In certain cases the latter splits up into several DNA clumps giving rise to a few (4–7) primary nucleoli, 3–5 μ in diameter. The primary nucleoli consist of densely packed fibrils 50–100 A thick. They contain no granular component and are inactive in RNA synthesis. — At the beginning of large growth the extrachromosomal DNA bodies disappear and numerous electron-dense clumps, 0,5–1 μ in diameter, appear in the nucleus. Instead of the primary nucleoli, the nucleus now contains a great number of ring nucleoli about 0,5–1 μ in diameter with a granular component (granules are 150 A). The space between them is filled up with nucleolar strands running from the surface of the ring nucleoli. — At the stage ring nucleoli of uridine−3 H incorporation into the oocyte nucleus begins. — During later previtellogenesis and at the beginning of vitellogenesis the ring nucleoli disappear and the nucleus is filled with the network of nucleolar strands. Among them there are specific complexes. These consist of electron dense masses, of granular clusters (granules 500 A in diameter) and large fibrillar electron light bodies. At this stage the nucleus takes the most active part in RNA synthesis. — The process of karyosphere capsule formation was studied by electron microscopy. The capsule was found to be of fibrillar nature; its structure is very peculiar and unlike any known membrane components of the cell. On the basis of cytochemical evidences the characteristics of the capsule are given. — The development of a powerful nucleolar apparatus based on the extrachromosomal DNA and a possible role of the synaptonemal complex and extrachromosomal DNA in formation of the karyosphere capsule is discussed.

Ultrastructure of the nucleus of Noctiluca (free living dinoflagellate) during sporulation

Abstract: The trophozoit of Noctiluca miliaris has a large nucleus (30 μ) with several nucleoli of considerable size that contain DNA fibrillae lying in the interspaces. — Before and during the first sporogenetic divisions, the nucleoli disintegrate, releasing towards the cytoplasma numerous groups of ribonucleic granules passing through the nuclear ampullae. At the end of the sporulation, there are no nucleoli visible in the nuclei and no ampullae. — The nucleoplasm diminishes, as the DNA filaments are built up, to form the meshes of a network which limit the masses of chromatic material that take the shape of chromosomes characterized by regular fibrillar arches, at the 8–16 nuclei stage. In their centre, there is an axial structure which remains intact during the chromosomal segregation; its function during mitosis seems to be important: supplementary layers of arches appear at this level. — The progressive condensation of the chromosomes is correlated to the sporogenetic evolution of the nuclei, not to the different phases of the mitotic cycle. — The karyokinesis is brought about, during early stages, by mere splitting of the chromatic mass and of its envelope, and later one by separation into two lots of chromosomes. The segregation of these chromosomes is effected by partial intervention and growth of the envelope of the nucleus; there is no centromeric structure visible. At the end of divisions, the nucleus is almost entirely formed by its chromosomes. — The nucleolar structure, the karyokinesis, the structure of the nuclear envelope and the chromosomal cycle show the particularly high evolution of Noctiluca, within the Dinoflagellata.

Further Experiments on the Role of the Nucleolus in the Expression of Structural Genes

Abstract: When a chick erythrocyte nucleus is introduced into the cytoplasm of a mouse cell it undergoes reactivation and eventually determines the synthesis of chick-specific proteins in the cytoplasm of the heterokaryon. It has previously been shown that there is a close temporal correlation between the onset of synthesis of the chick-specific proteins and the appearance of nucleoli in the erythrocyte nuclei. We now show that the synthesis of chick-specific proteins in the heterokaryon decays if the nucleolus in the reactivated erythrocyte nucleus is irradiated with an ultraviolet microbeam. The synthesis of chick-specific proteins does not, however, decay if only one of two nucleoli in the erythrocyte nucleus or if extranucleolar areas of the nucleus are irradiated. These observations confirm that some function located at or near the nucleolus is essential for the full expression of structural genes.

EFFECTS OF α-AMANITIN ON IN VITRO LABELING OF RNA FROM DEFINED NUCLEAR COMPONENTS IN SALIVARY GLAND CELLS FROM CHIRONOMUS TENTANS

Abstract: The effect of α-amanitin on nucleoside labeling of RNA in nucleoli, chromosomes, nuclear sap, and cytoplasm from Chironomus tentans salivary gland cells was investigated by radioautography and gel electrophoresis. Preribosomal RNA formation and processing in the nucleolus was not measurably influenced by the drug, and both 28 S and 18 S ribosomal RNA were transferred to the cytoplasm. In the chromosomes the heterogeneous RNA labeling was completely inhibited for the large size range (above 45–50 S) and partially for the low range. The labeling of 4–5 S chromosomal RNA was only moderately reduced. Most of the chromosomes showed radioautographically a disappearance of the normal band pattern, but some retained a pattern of weakly labeled bands. The electrophoretic results for the nuclear sap paralleled those for the chromosomes. The effect of α-amanitin on RNA labeling in these cells is similar but not identical to that of the substituted benzimidazole 5,6-dichloro-1(β-D-ribofuranosyl) benzimidazole (DRB).

In situ demonstration of dna hybridizing with chromosomal and nuclear sap rna in chironomus tentans

Abstract: Cytological hybridization combined with microdissection of Chironomus tentans salivary gland cells was used to locate DNA complementary to newly synthesized RNA from chromosomes and nuclear sap and from a single chromosomal puff, the Balbiani ring 2 (BR 2). Salivary glands were incubated with tritiated nucleosides. The labeled RNA was extracted from microdissected nuclei and hybridized to denatured squash preparations of salivary gland cells under conditions which primarily allow repeated sequences to interact. The bound RNA, resistant to ribonuclease treatment, was detected radioautographically. It was found that BR 2 RNA hybridizes specifically with the BR 2 region of chromosome IV. Nuclear sap RNA was fractionated into high and low molecular-weight RNA; the former hybridizes with the BR 2 region of chromosome IV, the latter in a diffuse distribution over the whole chromosome set. RNA from chromosome I hybridizes diffusely with all chromosomes. Nucleolar RNA hybridizes specifically with the nucleolar organizers, contained in chromosomes II and III. It is concluded that the BR 2 region of chromosome IV contains repeated DNA sequences and that nuclear sap contains BR 2 RNA.

The Effect of α-Amanitin and (NH4)2SO4 on RNA Synthesis in Nuclei and Nucleoli Isolated from Physarum polycephalum at Different Times during the Cell Cycle

Abstract: Nuclei isolated from various points in the mitotic cycle were examined for RNA synthesis in the presence of Mg2+ or Mn2++ (NH4)2SO4. A peak of activity 2.5—3 h after mitosis was only slightly increased by Mn2++ (NH4)2SO4 over that in the presence of Mg2+, and inhibited some 50% by low levels of α-amanitin in the presence of either Mg2+ or Mn2++ (NH4)2SO4. A G2 peak of activity 2.5—3 h before mitosis was 100% increased by Mn2++ (NH4)2SO4 over that observed in the presence of Mg2+, the increase being completely sensitive to low levels of α-amanitin, whereas the Mg2+ activity remained insensitive. Nucleoli isolated some 2.5—3 h before mitosis exhibited RNA synthesis which was unstimulated by Mn2++ (NH4)2SO4 and insensitive to α-amanitin. Similarities in RNA synthesis in isolated nucleoli and isolated G2 nuclei in the presence of Mg2+ with respect to pH optima and actinomycin-D sensitivity were observed. It is concluded that there is a high basal level of α-amanitin-insensitive RNA synthesis activity which is nucleolar and therefore probably ribosomal, rising to a peak some 2.5—3 h before mitosis, and a major peak of α-amanitin-sensitive nucleoplasmic RNA synthesis activity some 2.5—3 h after mitosis, thereafter dropping to a low level.

Ultrastructural analysis of antibody synthesis in cells from lymph and lymph nodes.

Abstract: A variety of cells containing antibody were found in lymph from sheep responding to secondary challenges with horseradish peroxidase. Antibody was present in blast cells in lymph within the perinuclear space, the endoplasmic reticulum, the Golgi apparatus and on polyribosomes. Some lymphocytes in lymph also contained antibody but in these cells it was located principally in the perinuclear space. No cells were found containing antibody distributed throughout their cytoplasm nor were any lymphocytes found with a Golgi apparatus positive for antibody. After the immune response in the lymph had died away, a population of cells containing antibody was still present in the regional lymph node. These were all plasma cells in which the antibody was present for the most part in a highly organized endoplasmic reticulum. Cells of this type were never found in the lymph. The cells containing the smallest amounts of antibody had a few discrete focal points in the perinuclear space and a few positive groups of ribosomes in their cytoplasm. The endoplasmic reticulum in some cells was filled completely with antibody, while in others positive segments were found adjacent to negative ones. The antibody in the cytoplasmic endoplasmic reticulum was in continuity with the antibody in the perinuclear space. The Golgi apparatus contained antibody in only a small proportion of the cells but when it was positive it was strongly so, suggesting that antibody was concentrated in this organelle. In some cells a positive reaction to horseradish peroxidase antibody appeared in the nucleus over the nucleoli. The significance of this finding is not known.

Effects of deoxyribonucleic acid-reactive drugs on ribonucleic acid synthesis in leukemia L1210 cells.

Abstract: Drugs having various modes of interaction with DNA were examined for their effects on RNA synthesis in L1210 cell cultures. Two aspects of RNA synthesis were emphasized: effects on chain lengths of RNA molecules synthesized in the presence of drug, and selective inhibition of nucleolar 45 S RNA synthesis. The drugs studied appeared to fall into three groups. Group I (actinomycin, daunomycin, and cordycepin) caused moderate reduction in chain length of nucleoplasmic RNA and marked selectivity in inhibition of nucleolar 45 S RNA synthesis. Group II (anthramycin, nitrogen mustard, and camptothecin) caused marked reduction in RNA chain length in both nucleoplasm and nucleolus, and did not selectively inhibit nucleolar 45 S RNA synthesis. Camptothecin, however, differed from the first two in that RNA chains eventually approached normal lengths if sufficient time was allowed. Group III (proflavin, ethidium, and ellipticine) did not produce RNA chain shortening and had moderate selectivity in inhibition of nucleolar 45 S RNA synthesis. Some of these findings could be interpreted in terms of known modes of interaction with DNA. In particular, it is proposed that the degree of reversibility of the DNA-inhibitor complex is an important, although not exclusive, determinant of the type of RNA synthesis effects produced.

Effects of d-Glucosamine, d-Mannosamine, and 2-Deoxy-d-glucose on the Ultrastructure of Ascites Tumor Cells in Vitro

Abstract: Ehrlich ascites carcinoma and Sarcoma 180 ascites tumor cells were treated in vitro with amino sugars, and the resulting changes were studied by light and electron microscopy. Addition of d-glucosamine or d-mannosamine to the incubation medium provoked striking cytoplasmic and nuclear changes. The earliest changes, seen after incubation for 15 min, included vacuolization of the cytoplasm and separation of the electron-lucent filamentous parts of the nucleolonema and nucleolar vacuole at the periphery of the nucleolus. As the time of incubation increased, vacuolization of the cytoplasm increased gradually, accompanied by retraction of the cytoplasm around the nucleus. After complete extrusion of its electron-lucent components, the nucleolus became condensed. At the end of 3 hr of incubation, 95% of the tumor cells had pycnotic, polymorphic, or disintegrating nuclei, and the nucleoli in nearly all the cells examined consisted almost exclusively of the compacted, granular, electron-dense nucleolonema. After 4 hr, most tumor cells exhibited various degrees of disintegration. Incubation of these tumor cell lines for 4 hr with 2-deoxyglucose resulted in no significant structural alteration in the ascites tumor cells.

Studies on Nucleoli and Cytoplasmic Fibrillar Bodies of Human Hepatocellular Carcinomas

Abstract: The ultrastructure of neoplastic cells of hepatocellular carcinomas in 3 untreated male patients of similar ages was studied and compared with liver cells of 2 persons with normal histology of the liver. As in rapidly growing experimental hepatomas and the Walker tumor of rats, nucleoli in a rapidly growing human hepatocellular carcinoma were characterized by a decrease of the relative percentages of total nucleolar areas containing fibrillar components when compared with slowly growing tumors or normal liver. The nucleolus-associated chromatin of the rapidly growing human hepatocellular carcinoma contained some “giant” microspherules with diameters up to 1400 A. The cytoplasm of these hepatocellular carcinoma cells contained fibrillar inclusion bodies and another type of fibrillar body.

Deoxyribonucleic acid metabolism in mouse trichinosis.

Abstract: Trichinosed mouse diaphragms showed an increase in total RNA, total DNA, and in the incorporation rates of thymidine over that of control muscle. Larval RNA, protein, /Lg RNA per mg protein, and incorporation rates of thymidine were determined. The incorporation of thymidine by infected diaphragms was adjusted for the larval component. Alterations in DNA metabolism are discussed in terms of the morphological changes and changes in RNA metabolism and rates of incorporation of amino acids by infected muscle. Fasske and Themann (1961), and RibasMujal and Rivera-Pomar (1968) have shown that during the early stages of infection with Trichinella spiralis host muscle fibers undergo numerous morphological changes. Read (1970) has reviewed several studies which demonstrate concomitant alterations in the enzymology of infected fibers. These chemical and morphological changes in infected host muscle are accompanied by marked alterations in the total ribonucleic acid (RNA) and rates of synthesis of RNA (Stewart and Read, 1972a). Immediately following a period of rapid RNA turnover there are increases in the rates of incorporation of amino acids by trichinosed muscle (Stewart and Read, 1972b). An increase in the size of nuclei and nucleoli, as well as a proliferation of mitochondria, in infected muscle fibers has been demonstrated by Fasske and Themann (1961) and Ribas-Mujal and Rivera-Pomar (1968). The present study deals with the chronology and magnitude of change in the total deoxyribonucleic acid (DNA) and rates of synthesis of DNA in infected and uninfected mouse

Cytotoxic Effects of d-Glucosamine on the Ultrastructures of Normal and Neoplastic Tissues in Vivo

Abstract: SUMMARY The effects of D-glucosamine on the fine structures of Walker carcinoma and of the liver and kidney in rats were studied in vivo. Continuous administration of D-glucosamine for 18 hr resulted in a slight dilation of the cisternae of the endoplasmic reticulum and Golgi sacs of the tumor cells and liver parenchymal cells. The epithelial cells of the renal tubules contained lamellar structures within numerous autophagic vacuoles. Infusion of D-glucosamine for 40 hr resulted in conspicuous degenerative changes in the nuclei and nucleoli of the Walker tumor cells. The nuclear matrix showed a much-decreased electron density and contained clumped interchromatin granules. The nucleoli had rounded up, and the strands of the nucleolonema had coalesced. Conspicuous small aggregates of high contrast, probably representing chromatin, had formed predominantly in the electron-lucent areas of the nucleolus. Cytoplasmic changes in tumor cells were relatively slight. The parenchymal cells of the liver showed almost complete fragmentation of the long profiles of the rough endoplasmic reticulum into small vesicles. Only single rows of the usual staggered array of cisternae remained surrounding mitochondria. Autophagic vacuoles were frequently seen. The lining epithelium of the proximal convoluted tubules of the kidney showed striking vesiculation of the cytoplasm. At no time was necrosis of the renal or hepatic cells observed. The most significant finding was the complete necrosis of tumor cells in rats sacrificed 5 days after 40 hr of infusion with D-glucosamine. This was accompanied by a remarkable recovery of the fine structure of the renal and hepatic parenchymal cells.

Nucleoli and Chromosomes: Their Relationships during the Meiotic Prophase of the Human Fetal Oocyte

Abstract: The number of nucleoli and the relations between them and chromosomes in the human fetal oocyte have been investigated in this study. The differences existing between first oocytes and spermatocytes have been emphasized. These differences concern essentially the number of nucleoli, the stage during which they appear and the quantity of heterochromatin associated with the nucleoli. The latter appear very early in the oocyte: they are already present at the beginning of the preleptotene stage. This stage is characterized by the processes of spiralization and despiralization, heretofore not described. The first prophase is characterized by the presence of abundant nucleolar material, especially in the diplotene stage. This abundance is certainly in relation with the active protein synthesis which characterizes the growth of the oocyte. As in the spermatocyte of the pachytene stage, the majority of nucleolar chromosomes, in the oocyte, are acrocentric. But in the latter, the quantity of heterochromatin associated with the nucleolus greatly exceeds the quantity present in the nucleolar organizers of acrocentric chromosomes, particularly during the leptotene stage. This supports the opinion that there are multiple sites for the synthesis of the various nucleolar componnents. Also discussed are the roles of heterochromatin and nucleolar organizers in nucleogenesis.

Studies on the metabolism of ribosomal structural proteins of regenerating rat liver. Sites of biosynthesis of structural proteins of large subunit and of their assembly with RNA moiety.

Abstract: In order to elucidate the site of the synthesis of ribosomal proteins and of their assembly with ribosomal RNA in liver cells, labeling experiments were performed in vivo, employing regenerating rat liver and the following results were obtained. 1 The relative synthetic rates of 28-S ribosomal RNA and ribosomal proteins at various time after partial hepatectomy, which were expressed as the ratios to those of the normal rat liver, ran parallel with each other and had maximal values, more than 10-times higher than those of normal rats, at 12 h after partial hepatectomy. 2 Disc electrophoresis of the acid-soluble proteins extracted from nucleolar detergent extract or 60-S particles by the procedures of Narayan and Birnstiel [1] revealed that the protein fraction contained most species of structural proteins of cytoplasmic ribosomes. Furthermore, the profiles of radioactivity on disc electrophoresis of the mixture of acid-soluble proteins of nucleolar 60-S particles and those of the large subunit of cytoplasmic ribosomes which were labeled with 3H and 14C-labeled amino acids, respectively, showed that most of the peaks of 3H and 14C radioactivities coincided with each other. The results may indicate the similarities between the protein moieties of these two particles. 3 The time courses of the incorporation of [3H]leucine into acid-soluble proteins of the nucleolar detergent extract and into the basic structural proteins of the cytoplasmic ribosomes suggested the precursor nature of proteins of the nucleolar extracts to structural proteins of the cytoplasmic ribosomes. Similarly the time-course study on the incorporation of [14C]orotic acid into RNA moieties and that of [3H]leucine into protein moieties of both nucleolar 60-S particles and the large subunit of the cytoplasmic ribosomes, indicated that 60-S nucleolar particles may be the precursor to the large subunit of cytoplasmic ribosomes. 4 The nascent ribosomal structural protein fraction, which was purified from the nascent protein fraction of regenerating-liver ribosomes labeled with [3H]leucine was subjected to disc electrophoresis. The radioactive proteins were distributed in the region of ribosomal structural proteins in addition to some amounts of radioactivity in more fast-moving regions. The time course of the incorporation of [3H]leucine into this nascent ribosomal proteins showed a sharp increase after the injection, reached a maximal value after 5 min and thereafter decreased rapidly, while that into proteins of nucleolar 60-S particles took the maximal value after 20 min. 5 These results may indicate that most of structural proteins of liver ribosomes are synthesized on cytoplasmic ribosomes, then transferred to the nucleolus where their assembly with ribosomal precursor RNA occurs to form 60-S nucleolar particles. The nucleolar 60-S particles may be the precursor to the large subunit of rat liver ribosomes.

Chromosomal localization of repetitive DNA in the newt, Triturus.

Abstract: The repetitive DNA sequences of the newt, Triturus viridescens, have been studied by nucleic acid hybridization procedures. Complementary RNA was synthesized enzymatically from unfractionated newt DNA. This RNA hybridized strongly to the centromeric regions of both somatic and lampbrush chromosomes It also bound to other loci scattered along the lengths of the chromosomes The amplified ribosomal DNA in the multiple oocyte nucleoli was demonstrated by in situ hybridization

Ultrastructural Changes Inducible with the Plant Alkaloid Camptothecin

Abstract: The fine structural effects of camptothecin, a plant alkaloid with known antitumor activity, have been examined in ME-180 tissue culture cells. The drug has recently been shown to inhibit ribosomal RNA synthesis; nucleolar changes were therefore expected to occur. As shown here, camptothecin induces nucleoli to segregate but, in addition, it produces microspherules and perichromatin-type granules that are associated with the fibrillar nucleolar elements. Following removal of the latter structures by digestion, fibrils 100 to 160 A thick are observed at their place that, because of a close association with condensed chromatin, are suspected to be a dispersed form of chromatin. The fibrils, microsperules, and perichromatin-type granules accumulate around fibrillar centers where the fibrillar nucleolonemal strands are normally located. It appears that the fibrillar strands fail to develop, which may explain why transcription of the 45 S ribosomal RNA cannot take place. Camptothecin cannot induce the microspherules and the perichromatin-type granules in the presence of actinomycin D. Since template inactivation by actinomycin D prevents their formation, it is postulated that templates, under the action of camptothecin, are in an activated state, permitting these structures to develop, but precluding transcription of ribosomal RNA.

Studies on nucleolar rna synthesis in drosophila melanogaster i. the relationship between number of nucleolar organizers and rate of synthesis

Abstract: SUMMARY Nucleolar RNA synthesis in salivary glands from XX, XY, and XO larvae having different numbers of nucleolus organizers was examined autoradiographically following incubation of the tissues in [ 3 H]uridine. In addition, the presence or absence of secondary constrictions was monitored in neuroblast preparations using acridine orange staining. It was observed that: (1) The rate of nucleolar [ 3 H]uridine incorporation is independent of the number of nucleolus organizers in the cell; (2) nucleolar incorporation in XO tissues is elevated relative to that observed in XX or XY material; and (3) where the number of organizers is in excess of the wild type (2), secondary constrictions form at only 2 of the nucleolus organizer sites. From these and related observations we suggest that there are 2 forms of transcriptional control for nucleolar RNA synthesis. One acts as an on-off control, influencing the formation of secondary constrictions at the nucleolus organizers. The other form modulates rates of transcription at organizers where constrictions have been formed.