scholarly journals THE PERMEABILITY OF THE AMPHIBIAN OOCYTE NUCLEUS, IN SITU

1972 ◽  
Vol 54 (3) ◽  
pp. 609-625 ◽  
Author(s):  
Samuel B. Horowitz
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Sucrose Concentration ◽  
Short Circuit ◽  
Pure Water ◽  
Oocyte Nucleus ◽  
Bathing Solution ◽  
Cytoplasmic Inclusions ◽  
Amphibian Oocyte ◽  
Water As Solvent

Ultralow temperature radioautography, suitable for the quantitative localization of diffusible solutes, was used to study the permeability of the nuclear envelope in the intact amphibian oocyte Sucrose-3H solutions were injected into mature oocytes, in volumes of 0 016–0 14% of that of the cell, and the subsequent movement of the solute was recorded. The resultant radioautographs show diffusion gradients in the cytoplasm and nucleus, and concentration gradients across the nuclear envelope Analysis of these gradients discloses that the nuclear envelope is as permeable as a comparable structure composed of cytoplasm, and is about 108 times more permeable than the oocyte plasma membrane The diffusion coefficient of sucrose in cytoplasm is 2 x 10-6 cm2/sec, or about one-third its diffusivity in pure water. This reduction can probably be accounted for by an effective lengthening of the diffusional path because of obstruction by cytoplasmic inclusions. The nuclear: cytoplasmic sucrose concentration ratio at diffusional equilibrium is about 3 05, or 1.6 times as great as expected from the water content of the two compartments This asymmetry is attributed to an unavailability of 36% of the cytoplasmic water as solvent Finally, sucrose entry into oocytes from a bathing solution was monitored by whole cell analysis and radioautography. These and the microinjection results are consistent with a model in which sucrose entry into the cell is entirely limited by the permeability of the plasma membrane. The results are inconsistent with cell models that hypothesize a short-circuit transport route from the extracellular compartment to the nucleus, and with models in which cytoplasmic diffusion is viewed as limiting the rate of solute permeation.

scholarly journals SMALL GRANULES IN THE AMPHIBIAN OOCYTE NUCLEUS AND THEIR RELATIONSHIP TO RNA

1956 ◽  
Vol 2 (4) ◽  
pp. 393-396 ◽  
Author(s):  
Joseph G. Gall
Keyword(s):  
Endoplasmic Reticulum ◽  
Salivary Gland ◽  
Nuclear Envelope ◽  
Oocyte Nucleus ◽  
Balbiani Ring ◽  
Lampbrush Chromosomes ◽  
Small Particles ◽  
Amphibian Oocyte

Small particles (100 to 300 A in diameter) are seen in sections of nucleoli, the loops of the amphibian lampbrush chromosomes, and the Balbiani-ring regions of dipteran salivary-gland chromosomes. All of these structures contain cytochemically demonstrable RNA. Furthermore, the annuli seen on the nuclear envelope are composed of small particles which are similar to or identical with those commonly associated with the endoplasmic reticulum. It seems likely that ribonucleoproteins are organized as small particulates in the nucleus as well as in the cytoplasm.

scholarly journals Visualizing Association of the Retroviral Gag Protein with Unspliced Viral RNA in the Nucleus

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Rebecca J. Kaddis Maldonado ◽  
Breanna Rice ◽  
Eunice C. Chen ◽  
Kevin M. Tuffy ◽  
Estelle F. Chiari ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Rous Sarcoma Virus ◽  
Nuclear Export ◽  
Live Cell ◽  
Viral Rna ◽  
Wild Type ◽  
Gag Protein ◽  
Rous Sarcoma ◽  
Sarcoma Virus

ABSTRACT Packaging of genomic RNA (gRNA) by retroviruses is essential for infectivity, yet the subcellular site of the initial interaction between the Gag polyprotein and gRNA remains poorly defined. Because retroviral particles are released from the plasma membrane, it was previously thought that Gag proteins initially bound to gRNA in the cytoplasm or at the plasma membrane. However, the Gag protein of the avian retrovirus Rous sarcoma virus (RSV) undergoes active nuclear trafficking, which is required for efficient gRNA encapsidation (L. Z. Scheifele, R. A. Garbitt, J. D. Rhoads, and L. J. Parent, Proc Natl Acad Sci U S A 99:3944–3949, 2002, https://doi.org/10.1073/pnas.062652199; R. Garbitt-Hirst, S. P. Kenney, and L. J. Parent, J Virol 83:6790–6797, 2009, https://doi.org/10.1128/JVI.00101-09). These results raise the intriguing possibility that the primary contact between Gag and gRNA might occur in the nucleus. To examine this possibility, we created a RSV proviral construct that includes 24 tandem repeats of MS2 RNA stem-loops, making it possible to track RSV viral RNA (vRNA) in live cells in which a fluorophore-conjugated MS2 coat protein is coexpressed. Using confocal microscopy, we observed that both wild-type Gag and a nuclear export mutant (Gag.L219A) colocalized with vRNA in the nucleus. In live-cell time-lapse images, the wild-type Gag protein trafficked together with vRNA as a single ribonucleoprotein (RNP) complex in the nucleoplasm near the nuclear periphery, appearing to traverse the nuclear envelope into the cytoplasm. Furthermore, biophysical imaging methods suggest that Gag and the unspliced vRNA physically interact in the nucleus. Taken together, these data suggest that RSV Gag binds unspliced vRNA to export it from the nucleus, possibly for packaging into virions as the viral genome. IMPORTANCE Retroviruses cause severe diseases in animals and humans, including cancer and acquired immunodeficiency syndromes. To propagate infection, retroviruses assemble new virus particles that contain viral proteins and unspliced vRNA to use as gRNA. Despite the critical requirement for gRNA packaging, the molecular mechanisms governing the identification and selection of gRNA by the Gag protein remain poorly understood. In this report, we demonstrate that the Rous sarcoma virus (RSV) Gag protein colocalizes with unspliced vRNA in the nucleus in the interchromatin space. Using live-cell confocal imaging, RSV Gag and unspliced vRNA were observed to move together from inside the nucleus across the nuclear envelope, suggesting that the Gag-gRNA complex initially forms in the nucleus and undergoes nuclear export into the cytoplasm as a viral ribonucleoprotein (vRNP) complex.

scholarly journals The polymerization of actin: II. how nonfilamentous actin becomes nonrandomly distributed in sperm: evidence for the association of this actin with membranes

1976 ◽  
Vol 69 (1) ◽  
pp. 51-72 ◽  
Author(s):  
LG Tilney
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Thin Section ◽  
Actin Filaments ◽  
Early Stage ◽  
Mature Sperm ◽  
Vacuole Membrane ◽  
Other Substances ◽  
Associated Proteins ◽  
Basal Half

At an early stage in spermiogenesis the acrosomal vacuole and other organelles including ribosomes are located at the basal end of the cell. From here actin must be transported to its future location at the anterior end of the cell. At no stage in the accumulation of actin in the periacrosomal region is the actin sequestered in a membrane-bounded compartment such as a vacuole or vesicle. Since filaments are not present in the periacrosomal region during the accumulation of the actin even though the fixation of these cells is sufficiently good to distinguish actin filaments in thin section, the actin must accumulate in the nonfilamentous state. The membranes in the periacrosomal region, specifically a portion of the nuclear envelope and the basal half of the acrosomal vacuole membrane, become specialized morphologically in advance of the accumulation of actin in this region. My working hypothesis is that the actin in combination with other substances binds to these specialized membranes and to itself and thus can accumulate in the periacrosmoal region by being trapped on these specialized membranes. Diffusion would then be sufficient to move these substances to this region. In support of this hypothesis are experiments in which I treated mature sperm with detergents, glycols, and hypotonic media, which solubilize or lift away the plasma membrane. The actin and its associated proteins remain attached to these specialized membranes. Thus actin can be nonrandomly distributed in cells in a nonfilamentous state presumably by its association with specialized membranes.

Bathing Solution Tonicity and Potassium Transport by the Midgut of the Tobacco Hornworm Manduca Sexta

1979 ◽  
Vol 78 (1) ◽  
pp. 213-223
Author(s):  
DAVID F. MOFFETT
Keyword(s):  
Manduca Sexta ◽  
Short Circuit ◽  
Weight Ratio ◽  
Short Circuit Current ◽  
Dry Weight ◽  
Potassium Transport ◽  
Bathing Solution ◽  
Histological Studies ◽  
Wet Weight ◽  
Small Solute

Potassium transport by the isolated midgut of Manduca larvae, as measured by the short circuit current, is inhibited by substitution of small organic solutes (M.W. < 340) for the sucrose normally included in bathing solution formulated for this tissue. Other solutes of molecular weight equal to or greater than sucrose are essentially as effective as sucrose in promoting the short circuit current. Equilibration of midgut in solutions containing the small solute mannitol results in a decrease in the dry weight/wet weight ratio of the tissue, suggesting that the small solutes can penetrate into areas of the tissue which are not accessible to sucrose. Histological studies suggest that sites of swelling in the presence of mannitol include both cytoplasm and goblet cell lumen. The inhibition of the short circuit current is rapidly reversible on return to bathing solution containing sucrose or another large solute. The effect of small solutes probably does not involve compromise of the energy source for potassium transport since oxygen uptake is unchanged in the presence of a small solute.

Effect of pH on chloride absorption in the flounder intestine

1988 ◽  
Vol 255 (2) ◽  
pp. G247-G252 ◽  
Author(s):  
A. N. Charney ◽  
J. I. Scheide ◽  
P. M. Ingrassia ◽  
J. A. Zadunaisky
Keyword(s):  
Small Intestine ◽  
Short Circuit ◽  
Ph Effect ◽  
Short Circuit Current ◽  
Bathing Solution ◽  
Solution Ph ◽  
Transport Pathway ◽  
Ussing Chambers ◽  
Chloride Absorption ◽  
In Series

Chloride absorption in the small intestine of the winter flounder, Pseudopleuronectes americanus, is reported to be sensitive to ambient pH. We studied this sensitivity in isolated stripped intestinal mucosa mounted in modified Ussing chambers. Unidirectional 36Cl fluxes (JClm----s, JCls----m) were measured under short-circuited conditions in bathing solutions containing various combinations of HCO3- (0-20 mM), partial pressure of CO2 (0-36 mmHg), and pH (6.77-7.85). We found that JClm----s, net 36Cl flux (JClnet), and short-circuit current (Isc) increased and JCls----m decreased predominately in response to increases in bathing solution pH. There was a linear relationship between pH and both JClnet (r = 0.92, P less than 0.01) and Isc (r = 0.96, P less than 0.005) between pH 6.77 and 7.74. The pH effect was completely reversible, did not require either CO2 or HCO3-, and was not affected by the presence of mucosal barium at 1 mM. Mucosal bumetanide (0.1 mM) completely inhibited the pH effect. These data suggest that the process by which Cl- is absorbed in the flounder intestine is sensitive to pH. The data do not indicate whether pH affects Na+-K+-2Cl- cotransport or a Cl- transport pathway in series with this process. The direction of Cl- absorption in response to pH contrasts with inverse relation of pH and Cl- absorption in mammalian small intestine.

Effects of histamine and histamine receptor antagonists on ion transport in rabbit descending colon

1984 ◽  
Vol 247 (4) ◽  
pp. G411-G418 ◽  
Author(s):  
R. D. McCabe ◽  
P. L. Smith
Keyword(s):  
Ion Transport ◽  
Receptor Antagonist ◽  
Prostaglandin E1 ◽  
Short Circuit ◽  
Bathing Solution ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
H2 Receptor Antagonist ◽  
Maximal Inhibition ◽  
Descending Colon

The effects of histamine on colonic ion transport were examined in in vitro preparations of rabbit descending colon. Serosal addition of histamine (10(-5) M) produced a transient increase in short-circuit current (Isc) and transepithelial conductance. The Isc response to histamine could be blocked by removing Cl from both bathing solutions, adding furosemide (10(-3) M) to the serosal bathing solution, adding indomethacin to the serosal and mucosal bathing solutions (10(-5) M), or removing Ca from the serosal bathing solution. In addition, the histamine-induced increase in Isc was inhibited in a dose-dependent manner by the H1-receptor antagonist diphenhydramine, with a maximal inhibition at 10(-4) M and a half-maximal inhibition at 3 X 10(-7) M. The H2-receptor antagonist cimetidine (10(-3) M) was without effect on the histamine response. Measurement of unidirectional Na, K, and Cl fluxes revealed that serosal addition of diphenhydramine (10(-3) M) reduced basal Isc due to a decrease in mucosal-to-serosal Na flux. Serosal addition of diphenhydramine (10(-3) M) also inhibited the increase in Isc produced by serosal addition of prostaglandin E1, 8-bromo-cAMP, cholera toxin, or the ionophore A23187. Measurement of unidirectional K and Cl fluxes revealed that prostaglandin E1 alone increased serosal-to-mucosal K and Cl fluxes and reduced the mucosal-to-serosal K flux, thereby increasing net K and Cl secretion. Serosal diphenhydramine (10(-3) M) abolished the changes in Cl fluxes produced by prostaglandin E1 and reduced the magnitude of the changes in K fluxes.(ABSTRACT TRUNCATED AT 250 WORDS)

The preparation and ultrastructure of avian erythrocyte nuclear envelope enclosed by the plasma membrane

1978 ◽  
Vol 34 (1) ◽  
pp. 81-90
Author(s):  
J.R. Harris
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Ammonium Molybdate ◽  
Negative Staining ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Thin Sectioning ◽  
Avian Erythrocyte ◽  
Pore Complexes ◽  
Low Ionic Strength

A procedure is described for the preparation of avian erythrocyte nuclear envelope ghosts which remain enclosed by the ellipsoid plasma membrane. Haemoglobin-free nucleated chicken erythrocyte ghosts are treated in a low ionic strength buffer plus heparin which brings about decondensation of the chromatin. This is followed by solubilization of the chromatin by digestion with pancreatic deoxyribonuclease-1. When studied by light microscopy using either phase-contrast or Nomarski interference optics, the ellipsoid plasma membrane is clearly seen to remain with the collapsed nuclear envelope trapped inside. This interpretation is supported by negative-staining electron microscopy using ammonium molybdate, which in addition reveals the presence of the nuclear pore complexes. The suggestion is advanced that structural protection is provided for the fragile nuclear envelope system by the surrounding plasma membrane, which might account for the final nuclear envelope being in the form of relatively intact ghosts with well defined nuclear pore complexes. The nuclear envelope is highly fragmented when the plasma membrane is absent, the nuclear pore complexes showing appreciable breakdown. Thin sectioning supports the results of negative staining and in addition shows the nuclear envelope retained within the plasma membrane to be composed of both inner and outer nuclear membranes, but the nuclear pore complexes are not clearly defined.

Pattern of Incorporation of [3H]Uridine into RNA of Amphibian Oocyte Nucleoli

1967 ◽  
Vol 2 (2) ◽  
pp. 145-150
Author(s):  
H. C. MACGREGOR
Keyword(s):  
Nuclear Envelope ◽  
Incubation Time ◽  
Core Material ◽  
Distinct Component ◽  
Thin Sections ◽  
The Core ◽  
Amphibian Oocyte ◽  
Nucleolar Rna ◽  
Silver Grains

Amphibian oocytes were incubated in vitro in the presence of [3H]uridine, and autoradiographs were made of nucleoli isolated from these oocytes and of sections of oocytes. After incubations of 2 h or less the nucleoli of oocytes larger than 0.6 mm diameter are asymmetrically labelled. With longer incubations nucleoli from oocytes of 0.6 to 1.1 mm diameter become more uniformly labelled. Those of oocytes larger than 1.2 mm diameter remain asymmetrically labelled whatever the incubation time. Autoradiographs of 1-µ sections through oocytes larger than 0.6 mm diameter show, after short incubations, asymmetrically labelled nucleoli. In these autoradiographs silver grains are concentrated over a distinct component of each nucleolus which is eccentrically placed towards the nuclear envelope. Thin sections of oocytes show nucleoli consisting of core and cortex. The core material is always concentrated into the half of the nucleolus which lies nearer the nuclear envelope. Autoradiographs of separated nucleolar cores and cortices from oocytes larger than 0.6 mm diameter show, after short incubations, silver grains over cores but not over cortices. Similar autoradiographs prepared from oocytes of 0.6 to 1.1 mm diameter, after longer incubations, show grains over cores and cortices. These results appear to indicate that nucleolar RNA is synthesized in the nucleolar core, in association with the nucleolar DNA, and is thence transferred to the cortex where it is built into ribonucleoprotein particles. Initial asymmetrical labelling is a consequence of the eccentric location of the nucleolar core. The nucleoli of oocytes smaller than 0.6 mm diameter always label symmetrically; such nucleoli consist entirely of core material. It is suggested that the nucleoli of oocytes larger than 1.2 mm diameter always label asymmetrically because transfer of RNA from core to cortex proceeds more slowly than in smaller oocytes.

Recovery of Transmembrane Potentials in Plants Resistant to Aryloxyphenoxypropanoate Herbicides: A Phenomenon Awaiting Explanation

Weed Science ◽  
1994 ◽  
Vol 42 (2) ◽  
pp. 293-301 ◽  
Author(s):  
Joseph A. M. Holtum ◽  
Rainer E. Häusler ◽  
Malcolm D. Devine ◽  
Stephen B. Powles
Keyword(s):  
Plasma Membrane ◽  
Single Gene ◽  
Membrane Vesicles ◽  
Bathing Solution ◽  
Root Tips ◽  
Plasma Membrane Vesicles ◽  
Bathing Medium ◽  
Transmembrane Potentials ◽  
Wild Oats ◽  
Rigid Ryegrass

Aryloxyphenoxypropanoate (APP) herbicides, such as diclofop, depolarize membranes in parenchyma cells of coleoptiles and root tips, and isolated tonoplast or plasma membrane vesicles from a variety of plant species. Some APP-resistant biotypes of rigid ryegrass and wild oat repolarize membranes after removal of herbicide from a bathing medium. The repolarization ability does not require presence of either APP-insensitive acetyl coenzyme A carboxylase or an increased capacity for herbicide detoxification. The kinetics of depolarization and repolarization depend upon the herbicide, the herbicide concentration, the biotype, and the pH of the bathing solution. For rigid ryegrass, depolarization in the presence of diclofop acid is more rapid than in the presence of diclofop-methyl, and 50% depolarization required about 4 μM diclofop acid. Both the nonherbicidal S(–) and the herbicidal R(+) enantiomers of diclofop acid depolarized membranes in susceptible and resistant ryegrass. Susceptible biotypes regenerated transmembrane potentials following removal of the S(–) but not the R(+) enantiomer, whereas resistant biotypes repolarized following exposure to either enantiomer or a mixture of the two. The herbicide 2,4-D affected, in a complex manner, the ability of both susceptible and resistant ryegrass biotypes to depolarize and repolarize. It is postulated that the intracellular concentration of diclofop acid in susceptible and resistant plants is not the same due to differences in the partitioning of diclofop acid between the extracellular spaces and the cytoplasm. The mechanism producing the postulated difference is unknown, but observations on the proton extrusion capacity of both ryegrass and wild oats, the responses of ryegrass to [K+] and PCMBS, and the single-gene inheritance pattern of resistance in wild oats indicate that changes in the diclofop sensitivity of a plasma membrane protein involved in the generation of proton or ion gradients may be involved.

Sign in / Sign up

Export Citation Format

Share Document