bluuebunny/crossref_metadata_2025 huggingface dataset:
Nuclear mutations affecting mitochondrial structure and function in Chlamydomonas.
Authors: Wiseman A , Gillham N W , Boynton J E
Published: May 2004

Wild type cells of the green alga Chlamydomonas reinhardtii can grow in the in the dark by taking up and respiring exogenously supplied acetate. Obligate photoautotrophic (dark dier, dk) mutants of this alga have been selected which grow at near wild type rates in the light, but rapidly die when transferred to darkness because of defects in mitochondrial structure and function. In crosses of the dk mutants to wild type, the majority of the mutants are inherited in a mendelian fashion, although two have been isolated which are inherited in a clearly nonmendelian fashion. Nine mendelian dk mutants have been analyzed in detail, and belong to eight different complementation groups representing eight gene loci. These mutants have been tentatively grouped into three classes on the basis of the pleiotropic nature of their phenotypic defects. Mutants in Class I have gross alterations in the ultrastructure of their mitochondrial inner membranes together with deficiencies in cytochrome oxidase and antimycin/rotenone-sensitive NADH-cytochrome c reductase activities. Mutants in Class II have a variety of less severe alterations in mitochondrial ultrastructure and deficiencies in cytochrome oxidase activity. Mutants in Class III have normal or near normal mitochondrial ultrastructure and reduced cytochrome oxidase activity. Eight of the nine mutants show corresponding reductions in cyanide-sensitive respiration.
Estradiol-induced redistribution of lysosomal proteins in rat preputial gland. Evidence from immunologic probes.
Authors: Szego C M , Nazareno M B , Porter D D
Published: May 2004

The influence of estrogen on the subcellular localization and distribution of lysosomal components of preputial gland was investigated in the ovariectomized rat. Antisera of high titer and specificity toward high-density lysosomal lipoproteins of this organ were raised in rabbits. The immunologic effectiveness of the IgG fraction so obtained was confirmed by microcomplement fixation, immunodiffusion, and immunoelectrophoresis. By both direct and indirect immunofluorescence techniques, cryostat sections of preputial gland from the control animals exhibited pinpoint cytoplasmic fluorescence, of dimensions corresponding to those of lysosomes. In contrast, specific immunoreactive material in corresponding target cells from animals receiving 0.1 microng of estradiol-17 beta/100 g body wt only 2 min earlier was distributed more homogeneously, indicating release of antigen from the membrane-bounded organelles. Moreover, specific immunofluorescence became evident at cell surfaces and in peri- and supranuclear localization, sites essentially negative in the controls. These effects were intensified at 15 min, as well as by maximal physiologic dose (0.5 microng/100 g body wt) of hormone. The relatively less active epimer, estradiol-17 alpha, exhibited only very limited effectiveness by some of these criteria. These observations, taken together with independent biochemical and ultrastructural evidence, lead to the conclusion that structural labilization of lysosomal constituents and their translocation to the nuclear compartment are early correlates of estrogen action.
Spatial relationship of photosystem I, photosystem II, and the light-harvesting complex in chloroplast membranes.
Authors: Armond P A , Staehelin L A , Arntzen C J
Published: May 2004

We have previously demonstrated (Armond, P. A., C. J. Arntzen, J.-M. Briantais, and C. Vernotte. 1976. Arch. Biochem. Biophys. 175:54-63; and Davis, D. J., P. A. Armond, E. L. Gross, and C. J. Arntzen. 1976. Arch. Biochem. Biophys. 175:64-70) that pea seedlings which were exposed to intermittent illumination contained incompletely developed chloroplasts. These plastids were photosynthetically competent, but did not contain grana. We now demonstrate that the incompletely developed plastids have a smaller photosynthetic unit size; this is primarily due to the absence of a major light-harvesting pigment-protein complex which is present in the mature membranes. Upon exposure of intermittent-light seedlings to continuous white light for periods up to 48 h, a ligh-harvesting chlorophyll-protein complex was inserted into the chloroplast membrane with a concomitant appearance of grana stacks and an increase in photosynthetic unit size. Plastid membranes from plants grown under intermediate light were examined by freeze-fracture electron microscopy. The membrane particles on both the outer (PF) and inner (EF) leaflets of the thylakoid membrane were found to be randomly distributed. The particle density of the PF fracture face was approx. four times that of the EF fracture face. While only small changes in particle density were observed during the greening process under continuous light, major changes in particle size were noted, particularly in the EF particles of stacked regions (EFs) of the chloroplast membrane. Both the changes in particle size and an observed aggregation of the EF particles into the newly stacked regions of the membrane were correlated with the insertion of light-harvesting pigment-protein into the membrane. Evidence is presented for identification of the EF particles as the morphological equivalent of a "complete" photosystem II complex, consisting of a phosochemically active "core" complex surrounded by discrete aggregates of the light-harvesting pigment protein. A model demonstrating the spatial relationships of photosystem I, photosystem II, and the light-harvesting complex in the chloroplast membrane is presented.
Ribosome processing in HeLa cells. Studies on structural aspects of precursor and mature ribosomes.
Authors: Johnson T R , Kumar A
Published: May 2004

Mature ribosomes, their subunits, and their ribonucleoprotein precursors were extracted from HeLa cells and prepared for electron microscopy by a method based on adsorption of nucleic acid to a charged grid. Images so obtained revealed the presence of two types of ribonuclease-sensitive fibrils of consistent morphology. One of these types of fibril is present on the 80S and 55S nucleolar ribonucleoprotein precursors and the mature 60S subunit. The other type is restricted to the 80S precursor. The morphology of the fibrils and their distribution in the various preparations suggest identification with known secondary structural features of isolated pre-rRNA.
Glycoprotein transport in the surface mucous cells of the rat stomach.
Authors: Kramer M F , Geuze J J
Published: May 2004

The intracellular transport of glycoproteins pulse-labeled in vitro with tritiated leucine and galactose in the surface mucous lining cells (SMC) of the fundus of the rat stomach was studied by electron microscope autoradiography. The SMC survive for several hours in pieces of the fundus incubated in a bicarbonate-buffered medium. The SMC have a normal ultrastructure for at least 4 h of incubation. Kinetic activity is normal for at least 5 h, as demonstrated by the normal nuclear incorporation of tritiated thymidine; The SMC incorporate labeled leucine and galactose at normal rates up to 4 h and 6 h, respectively. In contrast to the SMC, the cells of the gastric glands show signs of degeneration within 1 h after the start of incubation. In the SMC the secretory protein forms a smaller part of the total protein synthesized than in other secretory cells studied. The intracellular tranpsort of the leucine-labeled moiety of the glycoproteins follows the normal pathway. The RER loses 35% of its transportable labeled protein within 30 min. The Golgi complex is maximally labeled at 40 min and the mucous granules after 120 min. Galactose is attached to the glycoproteins mainly in the Golgi complex. Glycoproteins are not secreted within 2 h after synthesis of their protein moiety.
Quantitative determination of nuclear pore complexes in cycling cells with differing DNA content
Authors: Maul GG , Deaven L
Published: May 2004

The number of pore complexes per nucleus was determined for a wide variety of cultured cells selected for their variable DNA content over a range of 1-5,6000. The pore number was compared to DNA content, nuclear surface area, and nuclear volume. Values for pore frequency (pores/square micrometer) were relatively constant in the species studied. When the pore to DNA ratio was plotted against the DNA content, there was a remarkable correlation which decreased exponentially for the cells of vertebrae origin. Exceptions were the heteroploid mammalian cells which had the same ratio as the diploid mammalian cells despite higher DNA content.

The results are interpreted to mean that neither the nuclear surface, the nuclear volume, nor the DNA content alone determines the pore number of the nucleus, but rather an as yet undetermined combination of different factors. The surface and volume of vertebrate nuclei do not decrease with decreasing DNA content below a given value. The following speculation is suggested to account for the anomalous size changes of the nucleus relative to DNA content in vertebrates. Species with small DNA complements have a relatively large proportion of active chromatin which determines the limits of the physical parameters of the nucleus. The amount of active chromatin maybe the same for at least the vertebrates with low DNA content, At high DNA content, the nuclear parameters may be determined by the relatively high proportion of inactive condensed chromatin which increases the nuclear surface and volume.
Cytasters from sea urchin eggs parthenogenetically activated by procaine.
Authors: Moy G W , Brandriff B , Vacquier V D
Published: May 2004

A method is presented for the isolation of cytasters from unfertilized sea urchin eggs parthenogenetically activated by procaine. These cytasters do not appear to contain centrioles. The microtubules seem to grow out from the condensed chromosomes. The chromosomes have an unusual morphology.
Annulate lamellae in phloem cells of virus-infected Sonchus plants.
Authors: Steinkamp M P , Hoefert L L
Published: May 2004

The occurrence of annulate lamellae (AL) in differentiating phloem of Sonchus oleraceus (Compositae) singly infected with sowthistle yellow vein virus (SYVV) and doubly infected with a combination of SYVV and beet yellow stunt virus is documented by electron microscopy. Cell types in which AL were found were immature sieve elements and phloem parenchyma cells. AL were found only in cells that also contained SYVV particles although a direct association between the virus and AL was not apparent. The substructure of the AL and the relationships between the AL and the nuclear envelope and endoplasmic reticulum are similar to those reported in other descriptions of this organelle in the literature. This report appears to be the first one concerning the association of AL with a plant virus disease.
Light meromyosin paracrystal formation
Authors: Chowrashi PK , Pepe FA
Published: May 2004

Studies of paracrystal formation by column purified light meromyosin (LMM) prepared in a variety of ways led to the following conclusions: (a) different portions of the myosin rod may be coded for different stagger relationships. This was concluded from observations that paracrystals with different axial repeat periodicities could be obtained either with LMM framents of different lengths prepared with the same enzyme, or with LMM fragments of identical lengths but prepared with different enzymes. (b) Paracrystals with a 14-nm axial repeat periodicity are most likely formed by the aggregation of sheets with a 44-nm axial repeat within the sheets which are staggered by 14 nm. All of the axial repeat patterns expected from one sheet or aggregates of more than one sheet, on this basis, were observed in the same electron micrograph. (c) C-protein binding probably occurs preferentially to LMM molecules related in some specific way. This was concluded from the observation that the same axial repeat pattern was obtained in paracrystals formed from different LMM preparations in the presence of C-protein, regardless of differences in the axial repeat obtained in the absence of C-protein. (d) Nucleic acid is responsible for the 43-nm axial repeat patterns observed in paracrystals formed by the ethanol-resistant fraction of LMM. In the absence of nuclei acid, paracrystals with a 14nm axial repeat are obtained. (e) The 43-nm axial repeat pattern observed with the ethanol-resistant fraction of LMM is different for LMM preparations obtained by trypsin and papain digestions.
Changes in cell surface and cortical cytoplasmic organization during early embryogenesis in the preimplantation mouse embryo
Authors: Ducibella T , Ukena T , Karnovsky M , Anderson E
Published: May 2004

Membrane topography and organization of cortical cytoskeletal elements and organelles during early embryogenesis of the mouse have been studied by transmission and scanning electron microscopy with improved cellular preservation. At the four- and early eight-cell stages, blastomeres are round, and scanning electron microscopy shows a uniform distribution of microvilli over the cell surface. At the onset of morphogenesis, a reorganization of the blastomere surface is observed in which microvilli becomes restricted to an apical region and the basal zone of intercellular contact. As the blastomeres spread on each other during compaction, many microvilli remain in the basal region of imminent cell-cell contacts, but few are present where the cells have completed spreading on each other. Microvilli on the surface of these embryos contain linear arrays of microfilaments with lateral cross bridges.

Microtubules and mitochondria become localized beneath the apposed cell membranes during compaction. Arrays of cortical microtubules are aligned parallel to regions of apposed membranes. During cytokinesis, microtubules become redistributed in the region of the mitotic spindle, and fewer microvilli are present on most of the cell surface. The cell surface and cortical changes initiated during compaction are the first manifestations of cell polarity in embryogenesis. These and previous findings are interpreted as evidence that cell surface changes associated with trophoblast development appear as early as the eight-cell stage. Our observations suggest that morphogenesis involves the activation of a developmental program which coordinately controls cortical cytoplasmic and cell surface organization.
Choline acetyltransferase activity of spinal cord cell cultures increased by co-culture with muscle and by muscle-conditioned medium
Authors: Giller EL , Neale JH , Bullock PN , Schrier BK , Nelson PG
Published: May 2004

Activity of the enzyme choline acetyltransferase (CAT), which mediates the synthesis of the neurotransmitter, acetylcholine, was increased up to 20- fold in spinal cord (SC) cells grown in culture with muscle cells for 2 wk. This increase was directly related to the duration of co-culture as well as to the cell density of both the SC and muscle involved and was not affected by the presence of the acetylcholine receptor blocking agent, α-bungarotoxin. Glutamic acid decarboxylase (GAD) activity was often markedly decreased in SC-muscle cultures while the activities of acetylcholinesterase and several other enzymes were little changed.

Increased CAT activity was also observed when SC cultures were maintained in medium which had been conditioned by muscle cells or by undifferentiated cells from embryonic muscle. Muscle-conditioned medium (CM) did not affect the activities of SC cell GAD or acetylcholinesterase. Dilution or concentration of the CM directly affected its ability to increase SC CAT activity , as did the duration and timing of exposure of the SC cells to the CM. The medium could be conditioned by muscle cells in the presence or absence of serum, and remained effective after dialysis or heating to 58 degrees C. Membrane filtration data were consistent with the conclusion that the active material(s) in CM had a molecular weight in excess of 50,000 daltons. We conclude that large molecular weight material that is released by muscle cells is capable of producing a specific increase in CAT activity of SC cells.
Chromosomal basis of dosage compensation in drosophila: IX. cellular autonomy of the faster replication of the x chromosome in haplo-x cells of drosophila melanogaster and synchronous initiation
Authors: Chatterjee RN , Mukherjee AS
Published: May 2004

[(3)H]Thymidine labeling patterns have been examined in gynandric mosaic salivary glands of drosophila melanogaster. The Ring-X stock, R(1) w(ve)/In(1)dl 49, l (1) J1 y w lz(s), was used for this purpose. 365 labeled XX2A and 40 labeled XO2A nuclei were obtained from a total of 624 nuclei in nine pairs of mosaic salivary glands. It was observed that in all but those nuclei which had DD, 1C, and 2C patterns, the X chromosome of the XO2A nuclei always had fewer sites labeled than the X chromosomes of the XX2A nuclei, for a given pattern of the autosomes in either sex. Such asynchronous labeling of the X chromosome in the XO2A (male) nuclei was observed regardless of the proportion of the XO2A cells (2.0-73.7 percent), in the mosaic glands. Moreover, while the frequency of [(3)H]thymidine labeling for all of the 39 replicating units except the two late replicating sites (3C and 11A) in the X chromosome of the XO2A nuclei, was consistently lower than in the X chromosome of the XX2A nuclei, the mean number of grains on the X chromosome was relatively (to autosomes) similar in both XX2A and XO2A cells. The results, therefore, suggest that, as in XY2A larval glands, the X chromosome in the XO2A cells also completes the replication earlier than autosomes and that the XO2A nuclei show cellular autonomy with respect to the early replication of the X chromosome, like its counterpart, RNA transcription. Absence of the asynchrony during the initial phase (DD-2C) further completes the replication earlier but that the rate of replication of its DNA is possibly faster, and (b) that there might be a common regulation with respect to the initiation of replication of different chromosomes in a genome.
Immunological and ultrastructural studies of neurofilaments isolated from rat peripheral nerve
Authors: Schlaepfer WW
Published: May 2004

Neurofilaments were isolated from desheathed and minced segments of rat peripheral nerve by osmotic shock into 0.01 M Tris-HCI buffer, pH 7.2. Freshly isolated neurofilaments were observed to undergo disassembly by progressive fragmentation upon exposure of dilute tissue extracts to this buffer. Low- and high-speed centrifugations of these tissue extracts separated membranous and particulate constituents and produced a progressive enrichment of 68,000-dalton polypeptide band in successive supernates, as determined by analyses of soluble proteins by SDS-polyacrylamide electrophoresis. The final high-speed supernatant fractions (S3) of nerve extracts, which were predominantly composed of 68,000-dalton polypeptide, were used to raise a specific experimental antisera in rabbits.

Utilizing techniques of immune electron microscopy, experimental rabbit antisear was shown to contain antibodies against neurofilaments. Intact neurofilaments isolated from rat nerves and attached to carbon-coated grids became decorated when exposed to experimental rabbit antisera or purified gamma globulin (IgG) derivatives. The decoration of neurofilaments closely resembled the IgG coating seen in immune electron microscopy. Antibody absorption techniques were used to identify the biochemical constituency of neurofilamentous antigenic determinants. The decoration of neurofilament by experimental IgG was not altered by additions of tubulin or bovine serum albumin, but was prevented by additions of S3 fractions as well as the 68,000-dalton polypeptide of this fraction which was eluted and recovered from polyacrylamide gels.

These findings are indicative that a 68,000-dalton polypeptide is a constituent subunit of rat peripheral nerve neurofilaments.
Immunofluorescence studies of neurofilaments in the rat and human peripheral and central nervous system
Authors: Schlaepfer WW , Lynch RG
Published: May 2004

Localization of antisera to neurofilament antigens derived from rat peripheral nerve was carried out in tissues of rat and human peripheral and central nervous systems by indirect immunofluorescence. Unfixed and chloroform-methanol-fixed frozen sections of tissues were incubated in purified IgG of the experimental rabbit antisera and subsequently exposed to goat anti-rabbit IgG conjugated with fluorescein isothiocyanate. Control studies were conducted on identical tissue preparations incubated in the same concentrations of nonspecific rabbit IgG or in experimental rabbit IgG absorbed with extracts of rat peripheral nerve containing neurofilament antigen. Extensive immunofluorescence was observed in rat and human peripheral and central nervous systems. The distribution and configuration of immunofluorescence corresponded to neurofilament-rich structural components of these tissues. Prominent immunofluorescence was also noted in neuronal cell bodies of spinal sensory ganglia, especially in perikarya of the large neuronal type. Immunofluorescence of the central nervous system was located predominantly in myelinated axons of the white matter in cerebrum, cerebellum, brain stem, and spinal cord. Less intense immunofluorescence was also seen in neuronal perikarya and in short thin linear processes of grey matter.
The effect of myosin antibody on the division of starfish blastomeres
Authors: Mabuchi I , Okuno M
Published: May 2004

Antiserum against starfish egg myosin was produced in rabbits. Antibody specificity to myosin was demonstrated by Ouchterlony's immunodiffusion test and by immunoelectrophoresis in the presence of sodium dodecylsulfate (SDS). The latter technique showed that the antibody binds to both heavy and light chains of egg myosin. Furthermore, the antibody reacted with starfish sperm mysosin and starfish adult muscle myosin at both the heavy and light chains. It did not react with bovine platelet mysosin or rabbit skeletal muscle myosin in Ouchterlony's test; however, a weak reaction was observed in the presence of SDS between the antibody and these myosin heavy chains. Ca- and Mg-ATPase activities of egg myosin were not affected by the antibody, but it did inhibit actin-activated ATPase activity of egg myosin. Microinjection of the antibody into blastomeres of starfish eggs at the two-cell stage was carried out. Anti-egg myosin γ-globulin inhibited the subsequent cleavages at an amount of more than 0.3 ng when injected at interphase. The inhibition was reduced when the injection was carried out near the initiation of cleavage. At the onset of the second cleavage the antibody was not inhibitory; however, an appropriate amount inhibited the third cleavage. Although the disappearance of the nuclear membrane was observed in the presence of the antibody, the formation of the mitotic apparatus was more or less disturbed. However the formation of daughter nuclei seemed to be scarcely affected by the antibody except that the distance between the nuclei was significantly smaller than normal.
Identification of concanavalin A receptors and galactose-binding proteins in purified plasma membranes of Dictyostelium discoideum.
Authors: West C M , McMahon D
Published: May 2004

Two techniques have been modified to provide simple means for the identification of molecules which bind concanavalin A (Con A). Crossed immunoelectrophoresis was altered by replacing antibody with Con A, and receptors were identified by the precipitin arcs which they produced. Con A, tagged with fluorescein isothiocyanate, was also diffused into prefixed sodium dodecyl sulfate (SDS)-polyacrylamide gels, and additional receptors identified by fluorescence. More than 35 molecules in the plasma membranes of the cellular slime mold Dictyostelium discoideum which bind Con A were identified with these techniques. At least 12 of these diminish and 12 increase in importance as receptors during differentiation of the cells from the vegetative to the preculmination stage of development. In the course of these experiments, it was possible to confirm the presence of the galactose-binding protein discoidin, in the plasma membrane, by electrophoresing membrane proteins into an agarose gel. This lectin regains its sugar-binding activity after denaturation and electrophoresis in SDS.
Assembly of the sarcoplasmic reticulum proteins in differentiating rat skeletal muscle cell cultures: localization by immunofluorescence of sarcoplasmic reticulum proteins in differentiating rat skeletal muscle cell cultures
Authors: Jorgensen AO , Kalnins VI , Zubrzyca E , MacLennan DH
Published: May 2004

Immunofluorescent staining techniques were used to study the distribution of the Ca(2) + Mg(2+)-dependent ATPase and calsequestrin in primary cultures of differentiating rat skeletal muscle cells, grown for different periods of time under various culture conditions.

In mononucleated myoblasts calsequestrin was detected after 45 h in culture whereas the ATPase was not detected until 60 h. After cell fusion began, both proteins could be identified in all multinucleated cells. Myoblasts grown for longer than 60 h in low Ca(2+) medium contained calsequestrin and the ATPase, even though they were unable to fuse. These studies at the cellular level confirm biochemical findings on the biosynthesis of calsequestrin and the ATPase.

Immunofluorescent staining of myoblasts showed that calsequestrin first appears in a well-defined region of the cell near one end of the nucleus. At later times, the staining occupied progressively larger regions adjacent to the nucleus and took on a fibrous appearance. This suggests that calsequestrin first accumulates in the Golgi region and then gradually spreads throughout the cell. In contrast, the ATPase appeared to be concentrated in many small patches or foci throughout the cytoplasm and was never confined to one particular region, although some parts of the cell often stained more intensely than others. In multinucleated cells, alternating dark and fluorescent strands parallel to the longitudinal axis of the cells were evident.
Hepatocyte innervation in primates.
Authors: Forssmann W G , Ito S
Published: May 2004

The efferent innervation and some characteristics of nerve fibers of the liver lobule in the tree shrew, a primate, are described. Nerve endings on hepatocytes were encountered regularly and were determined to be efferent adrenergic nerves. Transmission electron microscopy revealed nerve endings and varicosities in close apposition to the hepatocytes adjacent to the connective tissue of the triads as well as within the liver lobule in the space of Disse. Fluorescence microscopy indicated the existence of adrenergic nerves with a similar distribution. Autoradiography of the avid uptake of exogenous [3H]norepinephrine indicated that all intralobular nerves are potentially norepinephrinergic (adrenergic). Chemical sympathectomy with 6-OH-dopamine resulted in the degeneration of all intralobular liver nerve fibers as revealed by fluorescence microscopy and electron microscopy. Substantial regeneration occurred after 60-90 days but was not completed by that time. Some nerves were also observed in close association with von Kupffer cells and endothelial cells. The functional significance of the efferent liver innervation is discussed.
Morphometry and ultrastructure of prismatic cristae in mitochondria of a crayfish muscle. A hypothesis of the structural principle.
Authors: Murdock L L , Cahill M A , Reith A
Published: May 2004

Approximately 40% of the mitochondria in the sphincter muscle of the crayfish vas deferens have prismatic-type cristae. In cross section, the angular cristae have either triangular or rhomboid profiles which are surrounded by a hexagonal array of electron-dense dots. In longitudinal section, these mitochondria exhibit both thick and thin parallel lines, which represent cristae and filaments, respectively. It is postulated that the matrix of the prismatic-type mitochondria is packed with rodlets composed of an electron-dense core and a less dense shell. Close packing of these rodlets results in the regular hexagonal dot array. Deletion of fascicles of 3 or 4 rodlets results in spaces with triangular or rectangular cross sections. Lining of these spaces with membranes results in cristae with triangular or rhomboid cross sections.
Studies on the mechanism for entry of vesicular stomatitis virus glycoprotein g mRNA into membrane-bound polyribosome complexes
Authors: Grubman MJ , Weinstein JA , Shafritz DA
Published: May 2004

Glycoprotein mRNA (G mRNA) of vesicular stomatitis virus is synthesized in the cytosol fraction of infected HeLa cells. Shortly after synthesis, this mRNA associates with 40S ribosomal subunits and subsequently forms 80S monosomes in the cytosol fraction. The bulk of labeled G mRNA is then found in polysomes associated with the membrane, without first appearing in the subunit or monomer pool of the membrane-bound fraction. Inhibition of the initiation of protein synthesis by pactamycin or muconomycin A blocks entry of newly synthesized G m RNA into membrane-bound polysomes. Under these circumstances, labeled G mRNA accumulates into the cytosol. Inhibition of the elongation of protein synthesis by cucloheximide, however, allows entry of 60 percent of newly synthesized G mRNA into membrane-bound polysomes. Furthermore, prelabeled G mRNA associated with membrane-bound polysomes is released from the membrane fraction in vivo by pactamycin or mucomycon A and in vitro by 1mM puromycin - 0.5 M KCI. This release is not due to nonspecific effects of the drugs. These results demonstrate that association of G mRNA with membrane-bound polysomes is dependent upon polysome formation and initiation of protein synthesis. Therefore, direct association of the 3' end of G mRNA with the membrane does not appear to be the initial event in the formation of membrane-bound polysomes.
Temporal expression of membrane antigens during mouse spermatogenesis
Authors: Millete CF , Bellve AR
Published: May 2004

The temporal expression of cell surface antigens during mammalian spermatogenesis has been investigated using isolated populations of mouse germ cells. Spermatogenic cells at advanced stages of differentiation, including pachytene primary spermatocytes, round spermatids, and residual bodies of Regaud and mature spermatozoa, contain common antigenic membrane components which are not detected before the pachytene stage of the first meiotic prophase. These surface constituents are not detected on isolated populations of primitive type A spermatogonia, type A spermatogonia, type B spermatogonia, preleptotene primary spermatocytes, or leptotene and zygotene primary spermatocytes.

These results have been demonstrated by immunofluorescence microscopy, by complement-mediated cytotoxicity, and by quantitative measurements of immunoglobulin (Ig) receptors on the plasma membrane of all cell populations examined. The cell surface antigens detected on germ cells are not found on mouse thymocytes, erythrocytes, or peripheral blood lymphocytes as determined by immunofluorescence and by cytotoxicity assays. Furthermore, absorption of antisera with kidney and liver tissue does not reduce the reactivity of the antibody preparations with spermatogenic cells, indicating that these antigenic determinants are specific to germ cells.

This represents the first direct evidence for the ordered temporal appearance of plasma membrane antigens specific to particular classes of mouse spermatogenic cells. It appears that at late meiotic prophase, coincident with the production of pachytene primary spermatocytes, a variety of new components are inserted into the surface membranes of developing germ cells. The further identification and biochemical characterization of these constituents should facilitate an understanding of mammalian spermatogenesis at the molecular level.
Organization of cell junctions in the peritoneal mesothelium
Authors: Simionescu M , Simionescu N
Published: May 2004

Intercellular junctions in the mesothelium of the visceral (mesentery and omentum), and parietal (diaphragm, pre-aortic, and iliac region) peritoneum were examined in rats and mice by using freeze-cleaved preparations. In addition to usual intercellular junctions (cell body junctions), special junctions are found between cell processes and the surface of the neighboring cell (cell process junctions). Cell body junctions are provided with tight junctions and communicating (gap) junctions. The former consist of one to two junctional strands which show a characteristic staggered arrangement, and focal discontinuities. In cell process junctions, the strands form loops or appear as short, free-ending elements; their polymorphism suggests considerable lability, probably in connection with their assembly and disassembly. The existence of free-ending strands indicates that such structures can be used as attachment devices without being concomitantly involved in the formation of occluding zonules. In both types of junctions, the strands can be resolved into bars, approximately 80- 100nm long, frequently provided with terminal enlargements and intercalated particles which occur singly or in small clusters. These particles are morphologically similar to those present in communicating (gap) junctions. The mesothelium is also provided with isolate composite macular junctions. Throughout the mesothelium, the cleavage plane follows the outer contour of junctional strands and particles, suggesting that strand-to-strand interactions in the apposed membranes are weaker than interactions between each strand and underlying cytoplasmic structures. In their general geometry and cleavage characteristics, the mesothelial junctions resemble the junctions found in the venular endothelium.
Binding of viral glycoprotein mRNA to endoplasmic reticulum membranes is disrupted by puromycin.
Authors: Lodish H F , Froshauer S
Published: May 2004

Previous studies showed that the glycoprotein (G) of vesicular stomatitis virus is synthesized in association with the endoplasmic reticulum (ER) membrane and that all G mRNA co-fractionates with ER membrane. Here, we show that treatment of infected cells with puromycin results in dissociation of G mRNA, and presumably the associated ribosomes, from the ER membrane. Even it extracts from treated cells are kept at low ionic strength (0.01 M KCl), over 80% of G mRNA is found unattached to membranes. There is no evidence for direct interaction of GmRNA with membranes; rather, the linkage apparently is mediated by the nascent G polypeptide.
Role of the tubulin-microtubule system in lymphocyte activation
Authors: Sherline P , Mundy GR
Published: May 2004

The role of the tubulin-microtubule system was examined in human peripheral blood leukocytes after activation with phytohemagglutinin (PHA). Soluble tubulin and microtubules were measured with a [(3)H]colchicine-binding assay. It was found that the tubulin content of PHA-activated lymphocytes was consistently increased relative to total protein content after 36 h of culture. There was no increase in the proportion of total tubulin synthesis which was present as microtubules at 36 h. Nevertheless, as a result of increased tubulin synthesis, there was a two-to three-fold increase in total microtubular mass. Colchicine, which disrupts microtubles, was used to assess the role of microtubule assembly in the sequence of events which follow lymphocyte activation, namely lymphokine release, protein synthesis, RNA synthesis, and DNA synthesis. Colchicine consistently inhibited DNA synthesis but did not inhibit release of the lymphokine, osteoclast activating factor (OAF). Protein and RNA syntheses were inhibited much less than DNA synthesis. The fact that some effects of PHA on lymphocytes appear to require intact microtubules and at least one does not suggest that the microtubule dependent step in PHA-stimulated lymphocyte activation occurs at a stage after propagation of the signal from the membrane to the cell interior.
On the mechanism of anaphase spindle elongation in Diatoma vulgare.
Authors: McDonald K , Pickett-Heaps J D , McIntosh J R , Tippit D H
Published: May 2004

Central spindles from five dividing cells (one metaphase, three anaphase, and one telophase) of Diatoma vulgare were reconstructed from serial sections. Each spindle is made up of two half-spindles that are composed almost entirely of polar microtubules. A small percentage of continuous microtubules and free microtubules were present in every stage except telophase. The half-spindles interdigitate at the midregion of the central spindle, forming a zone of overlap where the microtubules from one pole intermingle with those of the other. At metaphase the overlap zone is fairly extensive, but as elongation proceeds, the spindle poles move apart and the length of the overlap decreases because fewer microtubules are sufficiently long to reach from the pole to the zone of interdigitation. At telophase, only a few tubules are long enough to overlap at the midregion. Concurrent with the decrease in the length of the overlap zone is an increase in the staining density of the intermicrotubule matrix at the same region. These changes in morphology can most easily be explained by assuming zone mechanochemical interaction between microtubules in the overlap zone which results in a sliding apart of the two half-spindles.
Relationship between the golgi apparatus, gerl, and secretory granules in acinar cells of the rat exorbital lacrimal gland
Authors: Hand AR , Oliver C
Published: May 2004

The method of secretory granuleformation in the acinar cells of the rat exorbital lacrimal gland was studied by electron microscope morphological and cytochemical techniques. Immature secretory granules at the inner face of the Golgi apparatus were frequently attached to a narrow cisternal structure similar to GERL as described in neurons by Novikoff et al. (Novikoff, P. M., A. B. Novikoff, N. Quintana, and J.-J. Hauw. 1971. J. Cell Bio. 50:859-886). In the lacrimal gland. GERL was located adjacent to the inner Golgi saccule, or separated from it by a variable distance. Portions of GERL were often closely paralleled by modified cisternae of rough endoplasmic reticulum (RER), which lacked ribosomes on the surface adjacent to GERL. Diaminobenzidine reaction product of the secretory enzyme peroxidase was localized in the cisternae of the nuclear envelope, RER, peripheral Golgi vesicles, Golgi saccules, and immature and mature secretory granules. GERL was usually free of peroxidase reaction product or contained only a small amount. Thiamine pyrophosphatase reaction product was present in two to four inner Golgi saccules; occasionally, the innermost saccule was dilated and fenestrated, and contained less reaction product than the next adjacent saccule. Acid phosphatase (AcPase) reaction product was present in GERL, immature granules, and, rarely, in the innermost saccule, but not in the rest of the Golgi saccules. Thick sections of AcPase preparations viewed at 100 kV revealed that GERL consisted of cisternal, and fenestrated or tublular portions. The immature granules were attached to GERL by multiple connections to the tublular portions. These results suggest that, in the rat exorbital lacrimal gland, the Golgi saccules participate in the transport of secretory proteins, and that GERL is involved in the formation of secretory granules.
The sites of synthesis of the principal thylakoid membrane polypeptides in Chlamydomonas reinhardtii.
Authors: Chua N H , Gillham N W
Published: May 2004

The sites of synthesis of the major thylakoid membrane polypeptides have been studied in the green alga Chlamydomonas reinhardtii by pulse labeling of cells with [14C]acetate in the presence of inhibitors specific for chloroplast and cytoplasmic protein synthesis. The labeled membrane polypeptides were separated by an improved method of sodium dodecyl sulfate (SDS) gradient gel electrophoresis, and autoradiographs were made of the dried gels. The results demonstrate that of the 33 polypeptides resolved in the gels, at least nine are made on chloroplast ribosomes. Two of these (polypeptides 2 and 6) are associated with the reaction centers of photosystems I and II. Another polypeptide (polypeptide 5) appears from genetic data to be coded by chloroplast DNA. Experiments with a mutant whose chloroplast ribosomes are resistant to spectinomycyn (spr-u-1-6-2) show that polypeptides whose synthesis takes place on chloroplast ribosomes are made in the presence of spectinomycin in the mutant although their synthesis is blocked by this antibiotic in wild type cells.
Changes in the distribution of a major fibroblast protein, fibronectin, during mitosis and interphase
Authors: Stenman S , Wartiovaara J , Vaheri A
Published: May 2004

The distribution of a major fibroblast protein, fibronectin, was studied by immunofluorescence and immunoscanning electron microscopy in cultures of human and chicken fibroblasts during different phases of the cell cycle. The main findings were: (a) In interphase cells, the intensity of surface-associated fibronectin fluorescence correlated with that of intracellular fibronectin fluorescence. (b) The intensity of the fluorescence of both surface-associated and intracellular fibronectins was not changed in cells that were synthesizing DNA. (c) Mitotic cells had reduced amounts of surface-associated but not of intracellular fibronectin. The surface fibronectin that remained on meta-, ana-, or telophase cells had a distinct punctate distribution and was also localized to strands attaching the cells to the substratum. Fibronectin strands first reappeared on the surface of flattening cytoplasmic parts of telophase cells. (d) Fibronectin was also detected in extracellular fibrillar material on the growth substratum, particularly around dividing cells.

Thus, surface-associated fibrillar fibronectin was present during G(1), S, and G(2) but in cells undergoing mitosis the distribution was altered and the amount appeared to be reduced. The observations on the distribution of surface-associated fibronectin suggest that rather than being involved in growth control this fibronectin plays a structural role in interactions of cells with the environment.
Studies on proinsulin and proglucagon biosynthesis and conversion at the subcellular level: II. Distribution of radioactive peptide hormones and hormone precursors in subcellular fractions after pulse and pulse- chase incubation of islet tissue
Authors: Noe BD , Baste CA , Bauer GE
Published: May 2004

Anglerfish proinsulin and insulin were selectively labeled with [(14)C]isoleucine, while proglucagon, conversion intermediate(s), and glucagon were selectively labeled with[(3)H]tryptophan. After various periods of continuous or pulse-chase incubation, islet tissue was subjected to subcellular fractionation. Fraction extracts were analyzed by gel filtration for their content of precursor, conversion intermediate(s), and product peptides. Of the seven subcellular fractions prepared after each incubation, only the microsome and secretory granule fractions yielded significant amounts of labeled insulin-related and glucagon-related peptides. After short-pulse incubations, levels of both [(14)C]proinsulin and [(3)H]proglucagon (mol wt approximately 12,000) were highest in the microsome fraction. This fraction is therefore identified as the site of synthesis. With increasing duration of continuous incubation or during chase incubation in the absence of isotopes, proinsulin, proglucagon, and conversion intermediate(s) are transported to secretory granules. Conversion of proinsulin to insulin and proglucagon to a approximately 4,900 mol wt conversion intermediate and 3,500 mol wt glucagon occurs in the secretory granules. Converting activity also was observed in the microsome fraction. The recovery of most of the incorporated radioactivity in microsome and secretory granule fractions indicates that the newly synthesized islet peptides are relegated to a membrane-bound state soon after synthesis at the RER is completed. This finding supports the concept of intracisternal sequestration and intragranular maintenance of peptides synthesized for export from the cell of origin.
The proliferative response of hepatic peroxidomes of neonatal rats to treatment with SU-13 437 (nafenopin).
Authors: Stäubli W , Schweizer W , Suter J , Weibel E R
Published: May 2004

The repeated administration of the hypolipidemic agent Su-13 437 (nafenopin) to neonatal rats roughly doubled the number of peroxisomes in the liver tissue and caused a sixfold volumetric expansion of the peroxisomal compartment. During the proliferative response, the size-distribution of the peroxisomes was reversibly altered, enlarged particles appearing in numbers varying according to the dose given. By means of a new method for quantitative autoradiography, it was shown that (a) the concentration of silver grains over peroxisomes was comparable to that found over the endoplasmic reticulum; (b) the peak incorporation of [3H]arginine into the peroxisomes was dealyed in comparison with that into the endoplasmic reticulum; (c) the label, once incorporated into the expanding peroxisomal compartment, displayed the same shift to large particles as did the whole population. These results are compatible with the biosynthetic pathway for peroxisomal catalase proposed earlier (cf. reference 12), and with the notion that the drug-induced size-shift might have resulted from progressive growth of a particular class of peroxisomes formed in the presence of the agent. Evidence is presented to show that during the recovery period the larger peroxisomes are removed preferentially.
RNA polymerase activity in bovine spermatozoa.
Authors: Fuster C D , Farrell D , Stern F A , Hecht N B
Published: May 2004

Washed mature spermatozoa from bulls incorporate ribonucleoside triphosphates into RNA using an endogenous template. Maximum incorporation was observed at 31 degrees C in the presence of MgCl2, all four ribonucleoside triphosphates, beta-mercaptoethanol, and glycine sodium hydroxide buffer at pH 9.0. The amount of synthesis was linearly dependent upon the concentration of spermatozoa and continued for at least 4 h. Digestion studies revealed the RNA to be present in a protected (intracellular?) location in the spermatozoa. The RNA synthesis was inhibited by ethidium bromide, rifampicin, acriflavine, actinomycin D, and caffeine, but not by alpha-amanitine or rifamycin SV. Fractionation of the spermatozoa by sonication and separation of the heads and tails by centrifugation through a discontinuous gradient revealed that more than half of the total RNA polymerase activity was associated with the tail fraction.
Heat-induced reversible hexagonal packing of spindle microtubules.
Authors: Rieder C , Bajer A S
Published: May 2004

Epithelial cells cultured from the lung of the Northwest rough-skinned newt (Taricha granulosa granulosa) were subjected to brief (10-15 min) elevated temperature shocks of 33 degrees-36 degrees C during metaphase. Electron microscope studies on these cells reveal that the spindle microtubules (Mts) are differentially stable to heat treatment. The great majority of nonkinetochore Mts are destroyed within the first few minutes of the shock while kinetochore and adjacent Mts rearrange to form hexagonal closely packed structures before disassembling, the latter occurring only after prolonged heat treatment. The significance and theoretical implications of the formation of hexagonal closely packed Mt structures and of the differential stability of spindle Mts to heating are discussed. The data suggest the existence of one or more heat-sensitive structural component(s) which maintain the individual minimum spacing seen between spindle Mts. To our knowledge, this is the first reported instance of the experimental rearrangement of kinetochore Mts into reversible, hexagonal closely packed bundles.