Pollen development in orchids

1991 ◽  
Vol 99 (2) ◽  
pp. 273-281 ◽  
Author(s):  
R. C. BROWN ◽  
B. E. LEMMON
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Pollen Development ◽  
Nuclear Migration ◽  
Pollen Mitosis ◽  
Polar Structure ◽  
Microtubular Cytoskeleton ◽  
Distal Surface

Cytoplasmic preparation for the unequal first mitosis in non-vacuolate pollen of moth orchids (Phalaenopsis) includes reorganization of the microtubular cytoskeleton and nuclear migration. Following meiotic cytokinesis, both microtubules and F-actin are unpolarized in microspores of persistent tetrads. Microtubules radiate from the centrally located nucleus and F-actin forms a reticulate pattern in the cytoplasm. Polarization of the microspores is marked by a dramatic reorganization of microtubules while the pattern of F-actin remains unchanged. We describe a novel system of microtubules at the generative pole (GPMS), which forms a polar structure structure at the distal surface and marks the path of nuclear migration prior to pollen mitosis. The GPMS consists of numerous microtubules that extend between the plasma membrane and nuclear envelope. The nucleus becomes displaced toward the generative pole and flattened in association with microtubules of the GPMS. Initiation of the GPMS is marked by a localized proliferation of ER and clearing of large organelles from the generative pole.

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.

scholarly journals Counterregulation of clathrin-mediated endocytosis by the actin and microtubular cytoskeleton in human neutrophils

2009 ◽  
Vol 296 (4) ◽  
pp. C857-C867 ◽  
Author(s):  
Silvia M. Uriarte ◽  
Neelakshi R. Jog ◽  
Gregory C. Luerman ◽  
Samrath Bhimani ◽  
Richard A. Ward ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Actin Cytoskeleton ◽  
Human Neutrophils ◽  
Functional Responses ◽  
Membrane Expression ◽  
Granule Exocytosis ◽  
Plasma Membrane Expression ◽  
Microtubular Cytoskeleton ◽  
Latrunculin A ◽  
Azurophil Granule

We have recently reported that disruption of the actin cytoskeleton enhanced N-formylmethionyl-leucyl-phenylalanine (fMLP)-stimulated granule exocytosis in human neutrophils but decreased plasma membrane expression of complement receptor 1 (CR1), a marker of secretory vesicles. The present study was initiated to determine if reduced CR1 expression was due to fMLP-stimulated endocytosis, to determine the mechanism of this endocytosis, and to examine its impact on neutrophil functional responses. Stimulation of neutrophils with fMLP or ionomycin in the presence of latrunculin A resulted in the uptake of Alexa fluor 488-labeled albumin and transferrin and reduced plasma membrane expression of CR1. These effects were prevented by preincubation of the cells with sucrose, chlorpromazine, or monodansylcadaverine (MDC), inhibitors of clathrin-mediated endocytosis. Sucrose, chlorpromazine, and MDC also significantly inhibited fMLP- and ionomycin-stimulated specific and azurophil granule exocytosis. Disruption of microtubules with nocodazole inhibited endocytosis and azurophil granule exocytosis stimulated by fMLP in the presence of latrunculin A. Pharmacological inhibition of phosphatidylinositol 3-kinase, ERK1/2, and PKC significantly reduced fMLP-stimulated transferrin uptake in the presence of latrunculin A. Blockade of clathrin-mediated endocytosis had no significant effect on fMLP-stimulated phosphorylation of ERK1/2 in neutrophils pretreated with latrunculin A. From these data, we conclude that the actin cytoskeleton functions to limit microtubule-dependent, clathrin-mediated endocytosis in stimulated human neutrophils. The limitation of clathrin-mediated endocytosis by actin regulates the extent of both specific and azurophilic granule exocytosis.

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.

scholarly journals Signal Transduction across the Nuclear Envelope: Role of the LINC Complex in Bidirectional Signaling

Cells ◽  
2019 ◽  
Vol 8 (2) ◽  
pp. 124 ◽  
Author(s):  
Miki Hieda
Keyword(s):  
Signal Transduction ◽  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Cell Cycle Progression ◽  
Structural Integrity ◽  
Short Review ◽  
Linc Complex ◽  
Cellular Functions ◽  
Bidirectional Signaling ◽  
Extracellular Signal

The primary functions of the nuclear envelope are to isolate the nucleoplasm and its contents from the cytoplasm as well as maintain the spatial and structural integrity of the nucleus. The nuclear envelope also plays a role in the transfer of various molecules and signals to and from the nucleus. To reach the nucleus, an extracellular signal must be transmitted across three biological membranes: the plasma membrane, as well as the inner and outer nuclear membranes. While signal transduction across the plasma membrane is well characterized, signal transduction across the nuclear envelope, which is essential for cellular functions such as transcriptional regulation and cell cycle progression, remains poorly understood. As a physical entity, the nuclear envelope, which contains more than 100 proteins, functions as a binding scaffold for both the cytoskeleton and the nucleoskeleton, and acts in mechanotransduction by relaying extracellular signals to the nucleus. Recent results show that the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex, which is a conserved molecular bridge that spans the nuclear envelope and connects the nucleoskeleton and cytoskeleton, is also capable of transmitting information bidirectionally between the nucleus and the cytoplasm. This short review discusses bidirectional signal transduction across the nuclear envelope, with a particular focus on mechanotransduction.

scholarly journals NETosis proceeds by cytoskeleton and endomembrane disassembly and PAD4-mediated chromatin decondensation and nuclear envelope rupture

2020 ◽  
Vol 117 (13) ◽  
pp. 7326-7337 ◽  
Author(s):  
Hawa Racine Thiam ◽  
Siu Ling Wong ◽  
Rong Qiu ◽  
Mark Kittisopikul ◽  
Amir Vahabikashi ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Time Lapse ◽  
Extracellular Dna ◽  
Sterile Inflammation ◽  
Human Neutrophils ◽  
Chromatin Decondensation ◽  
Membrane Rupture ◽  
Cellular Events

Neutrophil extracellular traps (NETs) are web-like DNA structures decorated with histones and cytotoxic proteins that are released by activated neutrophils to trap and neutralize pathogens during the innate immune response, but also form in and exacerbate sterile inflammation. Peptidylarginine deiminase 4 (PAD4) citrullinates histones and is required for NET formation (NETosis) in mouse neutrophils. While the in vivo impact of NETs is accumulating, the cellular events driving NETosis and the role of PAD4 in these events are unclear. We performed high-resolution time-lapse microscopy of mouse and human neutrophils and differentiated HL-60 neutrophil-like cells (dHL-60) labeled with fluorescent markers of organelles and stimulated with bacterial toxins or Candida albicans to induce NETosis. Upon stimulation, cells exhibited rapid disassembly of the actin cytoskeleton, followed by shedding of plasma membrane microvesicles, disassembly and remodeling of the microtubule and vimentin cytoskeletons, ER vesiculation, chromatin decondensation and nuclear rounding, progressive plasma membrane and nuclear envelope (NE) permeabilization, nuclear lamin meshwork and then NE rupture to release DNA into the cytoplasm, and finally plasma membrane rupture and discharge of extracellular DNA. Inhibition of actin disassembly blocked NET release. Mouse and dHL-60 cells bearing genetic alteration of PAD4 showed that chromatin decondensation, lamin meshwork and NE rupture and extracellular DNA release required the enzymatic and nuclear localization activities of PAD4. Thus, NETosis proceeds by a stepwise sequence of cellular events culminating in the PAD4-mediated expulsion of DNA.

scholarly journals DrosophilaklaroidEncodes a SUN Domain Protein Required for Klarsicht Localization to the Nuclear Envelope and Nuclear Migration in the Eye

Fly ◽  
2007 ◽  
Vol 1 (2) ◽  
pp. 75-85 ◽  
Author(s):  
Martin P. Kracklauer ◽  
Susan M.L. Banks ◽  
Xuanhua Xie ◽  
Yaning Wu ◽  
Janice A. Fischer
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Migration ◽  
Domain Protein ◽  
Sun Domain

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 Live-Cell Coimaging of the Genomic RNAs and Gag Proteins of Two Lentiviruses

2010 ◽  
Vol 84 (13) ◽  
pp. 6352-6366 ◽  
Author(s):  
Iris Kemler ◽  
Anne Meehan ◽  
Eric M. Poeschla
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Spatial Dynamics ◽  
Live Cells ◽  
Genomic Rna ◽  
Genomic Rnas ◽  
Gag Proteins ◽  
Immunodeficiency Virus ◽  
Genome Association ◽  
Hiv 1

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) Gag and genomic RNA determinants required for encapsidation are well established, but where and when encapsidation occurs in the cell is unknown. We constructed MS2 phage coat protein labeling systems to track spatial dynamics of primate and nonprimate lentiviral genomic RNAs (HIV-1 and feline immunodeficiency virus [FIV]) vis-à-vis their Gag proteins in live cells. Genomic RNAs of both lentiviral genera were observed to traffic into the cytoplasm, and this was Rev dependent. In transit, FIV Gag and genomic RNA accumulated independently of each other at the nuclear envelope, and focal colocalizations of genomic RNA with an intact packaging signal (ψ) and Gag were observed to extend outward from the cytoplasmic face. In contrast, although HIV-1 genomic RNA was detected at the nuclear envelope, HIV-1 Gag was not. For both lentiviruses, genomic RNAs were seen at the plasma membrane if and only if Gag was present and ψ was intact. In addition, HIV-1 and FIV genomes accumulated with Gag in late endosomal foci, again, only ψ dependently. Thus, lentiviral genomic RNAs require specific Gag binding to accumulate at the plasma membrane, packaged genomes cointernalize with Gag into the endosomal pathway, and plasma membrane RNA incorporation by Gag does not trigger committed lentiviral particle egress from the cell. Based on the FIV results, we hypothesize that the Gag-genome association may initiate at the nuclear envelope.

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