Nuclear envelope breakdown in mammalian cells involves stepwise lamina disassembly and microtubule-drive deformation of the nuclear membrane

1997 ◽  
Vol 110 (17) ◽  
pp. 2129-2140 ◽  
Author(s):  
S.D. Georgatos ◽  
A. Pyrpasopoulou ◽  
P.A. Theodoropoulos
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Mitotic Spindle ◽  
Mammalian Cells ◽  
Nuclear Lamina ◽  
Early Prophase ◽  
Nuclear Surface ◽  
Late Prophase ◽  
Lamin B ◽  
Nuclear Envelope Breakdown

We have studied nuclear envelope disassembly in mammalian cells by morphological methods. The first signs of nuclear lamina depolymerization become evident in early prophase as A-type lamins start dissociating from the nuclear lamina and diffuse into the nucleoplasm. While B-type lamins are still associated with the inner nuclear membrane, two symmetrical indentations develop on antidiametric sites of the nuclear envelope. These indentations accommodate the sister centrosomes and associated astral microtubules. At mid- to late prophase, elongating microtubules apparently push on the nuclear surface and eventually penetrate the nucleus. At this point the nuclear envelope becomes freely permeable to large ligands, as indicated by experiments with digitonin-treated cells and by the massive release of solubilized A-type lamins into the cytoplasm. At the prophase/prometaphase transition, the B-type lamina is fragmented, but ‘islands’ of lamin B polymer can still be discerned on the tips of congressing chromosomes. Finally, at metaphase, the lamin B polymer breaks down into small pieces, which tend to concentrate in the area of the mitotic spindle. Nuclear envelope breakdown is not prevented when the microtubules are depolymerized by nocodazole; however, the mode of nuclear lamina fragmentation in the absence of microtubules is markedly different from the normal one and involves multiple raffles and gaps, which develop rapidly along the entire surface of the nuclear envelope. These data suggest that nuclear envelope disassembly is a stepwise process in which the microtubules play an important part.

Sequential PKC- and Cdc2-mediated phosphorylation events elicit zebrafish nuclear envelope disassembly

1999 ◽  
Vol 112 (6) ◽  
pp. 977-987 ◽  
Author(s):  
P. Collas
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Nuclear Lamina ◽  
Cdc2 Kinase ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Nuclear Envelope Breakdown ◽  
Simultaneous Inhibition ◽  
Integral Proteins

Molecular markers of the zebrafish inner nuclear membrane (NEP55) and nuclear lamina (L68) were identified, partially characterized and used to demonstrate that disassembly of the zebrafish nuclear envelope requires sequential phosphorylation events by first PKC, then Cdc2 kinase. NEP55 and L68 are immunologically and functionally related to human LAP2beta and lamin B, respectively. Exposure of zebrafish nuclei to meiotic cytosol elicits rapid phosphorylation of NEP55 and L68, and disassembly of both proteins. L68 phosphorylation is completely inhibited by simultaneous inhibition of Cdc2 and PKC and only partially blocked by inhibition of either kinase. NEP55 phosphorylation is completely prevented by inhibition or immunodepletion of cytosolic Cdc2. Inhibition of cAMP-dependent kinase, MEK or CaM kinase II does not affect NEP55 or L68 phosphorylation. In vitro, nuclear envelope disassembly requires phosphorylation of NEP55 and L68 by both mammalian PKC and Cdc2. Inhibition of either kinase is sufficient to abolish NE disassembly. Furthermore, novel two-step phosphorylation assays in cytosol and in vitro indicate that PKC-mediated phosphorylation of L68 prior to Cdc2-mediated phosphorylation of L68 and NEP55 is essential to elicit nuclear envelope breakdown. Phosphorylation elicited by Cdc2 prior to PKC prevents nuclear envelope disassembly even though NEP55 is phosphorylated. The results indicate that sequential phosphorylation events elicited by PKC, followed by Cdc2, are required for zebrafish nuclear disassembly. They also argue that phosphorylation of inner nuclear membrane integral proteins is not sufficient to promote nuclear envelope breakdown, and suggest a multiple-level regulation of disassembly of nuclear envelope components during meiosis and at mitosis.

scholarly journals Loss of a-Type Lamin Expression Compromises Nuclear Envelope Integrity Leading to Muscular Dystrophy

1999 ◽  
Vol 147 (5) ◽  
pp. 913-920 ◽  
Author(s):  
Teresa Sullivan ◽  
Diana Escalante-Alcalde ◽  
Harshida Bhatt ◽  
Miriam Anver ◽  
Narayan Bhat ◽  
...  
Keyword(s):  
Muscular Dystrophy ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Mammalian Cells ◽  
Nuclear Architecture ◽  
Nuclear Lamina ◽  
Postnatal Growth ◽  
Developmentally Regulated ◽  
Inner Nuclear Membrane ◽  
Cardiac Muscles

The nuclear lamina is a protein meshwork lining the nucleoplasmic face of the inner nuclear membrane and represents an important determinant of interphase nuclear architecture. Its major components are the A- and B-type lamins. Whereas B-type lamins are found in all mammalian cells, A-type lamin expression is developmentally regulated. In the mouse, A-type lamins do not appear until midway through embryonic development, suggesting that these proteins may be involved in the regulation of terminal differentiation. Here we show that mice lacking A-type lamins develop to term with no overt abnormalities. However, their postnatal growth is severely retarded and is characterized by the appearance of muscular dystrophy. This phenotype is associated with ultrastructural perturbations to the nuclear envelope. These include the mislocalization of emerin, an inner nuclear membrane protein, defects in which are implicated in Emery-Dreifuss muscular dystrophy (EDMD), one of the three major X-linked dystrophies. Mice lacking the A-type lamins exhibit tissue-specific alterations to their nuclear envelope integrity and emerin distribution. In skeletal and cardiac muscles, this is manifest as a dystrophic condition related to EDMD.

scholarly journals Identification of a Golgi-associated protein that undergoes mitosis dependent phosphorylation and relocation.

1990 ◽  
Vol 110 (5) ◽  
pp. 1513-1523 ◽  
Author(s):  
I McMorrow ◽  
W E Souter ◽  
G Plopper ◽  
B Burke
Keyword(s):  
Monoclonal Antibody ◽  
Nuclear Envelope ◽  
Immunofluorescence Microscopy ◽  
Nuclear Lamina ◽  
Early Prophase ◽  
Perinuclear Region ◽  
Nuclear Envelope Breakdown ◽  
Peripheral Membrane Protein ◽  
Double Label ◽  
Late Telophase

By means of a monoclonal antibody (BH3), we have identified a 57-kD protein (p57) that in interphase is restricted largely to the perinuclear region of the cell. Double label immunofluorescence microscopy suggests localization of p57 to the Golgi complex and associated membranous structures. Protease protection experiments and chemical extractability indicate that p57 is a peripheral membrane protein exposed to the cytoplasm. p57 displays unique behavior during mitosis. At the end of G2 or in early prophase, p57 leaves the perinuclear region and accumulates very rapidly within the nucleus, at a time when the nuclear envelope is still intact and before nuclear lamina disassembly. This relocation of p57 coincides with its hyperphosphorylation on serine and threonine residues. After nuclear envelope breakdown p57 becomes uniformly distributed throughout the mitotic cytoplasm until in late telophase when it returns to its perinuclear location and is once again excluded from the nucleus. The behavior of p57 during mitosis suggests that it may play a role in the cellular reorganization evident during mitotic prophase.

scholarly journals A computational model for the formation of lamin-B mitotic spindle envelope and matrix

2014 ◽  
Vol 4 (3) ◽  
pp. 20130063 ◽  
Author(s):  
Changji Shi ◽  
Wilbur E. Channels ◽  
Yixian Zheng ◽  
Pablo A. Iglesias
Keyword(s):  
Computational Model ◽  
Molecular Species ◽  
Mitotic Spindle ◽  
Effective Diffusion ◽  
Nuclear Lamina ◽  
Nuclear Proteins ◽  
Lamin B ◽  
Nuclear Envelope Breakdown ◽  
Spindle Poles ◽  
Continuum Description

Recent reports show that, after nuclear envelope breakdown, lamin-B, a component of the nuclear lamina in interphase, localizes around the mitotic spindle as a membranous network. How this process occurs, however, and how it influences mitotic spindle morphogenesis is unclear. Here, we develop a computational model based on a continuum description to represent the abundance and location of various molecular species involved during mitosis, and use the model to test a number of hypotheses regarding the formation of the mitotic matrix. Our model illustrates that freely diffusible nuclear proteins can be captured and transported to the spindle poles by minus-end-directed microtubule (MT) motors. Moreover, simulations show that these proteins can be used to build a shell-like region that envelopes the mitotic spindle, which helps to improve the focusing of the mitotic spindle by spatially restricting MT polymerization and limiting the effective diffusion of the free MTs. Simulations also confirm that spatially dependent regulation of the spindle network through the Ran system improves spindle focusing and morphology. Our results agree with experimental observations that lamin-B reorganizes around the spindle and helps to maintain spindle morphology.

scholarly journals Nuclear envelope breakdown in starfish oocytes proceeds by partial NPC disassembly followed by a rapidly spreading fenestration of nuclear membranes

2003 ◽  
Vol 160 (7) ◽  
pp. 1055-1068 ◽  
Author(s):  
Péter Lénárt ◽  
Gwénaël Rabut ◽  
Nathalie Daigle ◽  
Arthur R. Hand ◽  
Mark Terasaki ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Phase Ii ◽  
Time Course ◽  
Nuclear Lamina ◽  
Ultrastructural Level ◽  
Nuclear Pore ◽  
Nuclear Surface ◽  
Light Microscopic Level ◽  
Nuclear Envelope Breakdown ◽  
Nuclear Membranes

Breakdown of the nuclear envelope (NE) was analyzed in live starfish oocytes using a size series of fluorescently labeled dextrans, membrane dyes, and GFP-tagged proteins of the nuclear pore complex (NPC) and the nuclear lamina. Permeabilization of the nucleus occurred in two sequential phases. In phase I the NE became increasingly permeable for molecules up to ∼40 nm in diameter, concurrent with a loss of peripheral nuclear pore components over a time course of 10 min. The NE remained intact on the ultrastructural level during this time. In phase II the NE was completely permeabilized within 35 s. This rapid permeabilization spread as a wave from one epicenter on the animal half across the nuclear surface and allowed free diffusion of particles up to ∼100 nm in diameter into the nucleus. While the lamina and nuclear membranes appeared intact at the light microscopic level, a fenestration of the NE was clearly visible by electron microscopy in phase II. We conclude that NE breakdown in starfish oocytes is triggered by slow sequential disassembly of the NPCs followed by a rapidly spreading fenestration of the NE caused by the removal of nuclear pores from nuclear membranes still attached to the lamina.

scholarly journals A comparison of the distribution of actin and tubulin in the mammalian mitotic spindle as seen by indirect immunofluorescence.

1977 ◽  
Vol 72 (3) ◽  
pp. 552-567 ◽  
Author(s):  
W Z Cande ◽  
E Lazarides ◽  
J R McIntosh
Keyword(s):  
Indirect Immunofluorescence ◽  
Mitotic Spindle ◽  
Metaphase Plate ◽  
Spindle Pole ◽  
Early Prophase ◽  
Chromosome Movement ◽  
Late Prophase ◽  
Small Ball ◽  
Nuclear Envelope Breakdown ◽  
Daughter Cells

Rabbit antibodies against actin and tubulin were used in an indirect immunofluorescence study of the structure of the mitotic spindle of PtK1 cells after lysis under conditions that preserve anaphase chromosome movement. During early prophase there is no antiactin staining associated with the mitotic centers, but by late prophase, as the spindle is beginning to form, a small ball of actin antigenicity is found beside the nucleus; After nuclear envelope breakdown, the actiactin stains the region around each mitotic center, and becomes organized into fibers that run between the chromosomes and the poles. Colchicine blocks this organization, but does not disrupt the staining at the poles. At metaphase the antiactin reveals a halo of ill-defined radius around each spindle pole and fibers that run from the poles to the metaphase plate. Antitubulin shows astral rays, fibers running from chromosomes to poles, and some fibers that run across the metaphase plate. At anaphase, there is a shortening of the antiactin-stained fibers, leaving a zone which is essentially free of actin-staining fluorescence between the separating chromosomes. Antitubulin stains the region between chromosomes and poles, but also reveals substantial fibers running through the zone between separating chromosomes. Cells fixed during cytokinesis show actin in the region of the cleavage furrow, while antitubulin reveals the fibrous spindle remnant that runs between daughter cells. These results suggest that actin is a component of the mammalian mitotic spindle, that the distribution of actin differs from that of tubulin and that the distributions of these two fibrous proteins change in different ways during anaphase.

Nuclear envelope disassembly in mitotic extract requires functional nuclear pores and a nuclear lamina

1998 ◽  
Vol 111 (9) ◽  
pp. 1293-1303 ◽  
Author(s):  
P. Collas
Keyword(s):  
Nuclear Envelope ◽  
Sea Urchin ◽  
Nuclear Membrane ◽  
Critical Role ◽  
Nuclear Lamina ◽  
Nuclear Pore ◽  
Nuclear Pores ◽  
Lamin B ◽  
Nuclear Membranes

Using sea urchin embryonic and in-vitro-assembled nuclei incubated in sea urchin mitotic extract, I provide evidence for a requirement for functional nuclear pores and a nuclear lamina for nuclear envelope disassembly in vitro. In interphase gastrula nuclei, lamin B interacts with p56, an integral protein of inner nuclear membrane cross-reacting with antibodies to human lamin B receptor. Incubation of gastrula nuclei in mitotic cytosol containing an ATP-generating system rapidly induces hyperphosphorylation of p56 and lamin B. Subsequently, p56-lamin B interactions are weakened and the two proteins segregate into distinct nuclear envelope-derived vesicles upon disassembly of nuclear membranes and of the lamina. Nuclear disassembly is accompanied by chromatin condensation. Blocking nuclear pore function with wheat germ agglutinin or antibodies to nucleoporins prevents p56 and lamin B hyperphosphorylation, nuclear membrane breakdown and lamina solubilization. These events are not rescued by permeabilization of nuclear membranes to molecules of 150, 000 Mr with lysolecithin. In-vitro-assembled nuclei containing nuclear membranes with functional pores but no lamina do not disassemble in mitotic cytosol in spite of p56 hyperphosphorylation. Nuclear import of soluble lamin B and reformation of a lamina in interphase extract restores nuclear disassembly in mitotic cytosol. The data indicate a role for functional nuclear pores in nuclear disassembly in vitro. They show that p56 hyperphosphorylation is not sufficient for nuclear membrane disassembly in mitotic cytosol and argue that the nuclear lamina plays a critical role in nuclear disassembly at mitosis.

Lamin B receptor: role on chromatin structure, cellular senescence and possibly aging

2020 ◽  
Vol 477 (14) ◽  
pp. 2715-2720
Author(s):  
Susana Castro-Obregón
Keyword(s):  
Cellular Senescence ◽  
Nuclear Membrane ◽  
Chromatin Structure ◽  
Cholesterol Synthesis ◽  
Reductase Activity ◽  
Chimeric Protein ◽  
Nuclear Lamina ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Lamin B Receptor

The nuclear envelope is composed by an outer nuclear membrane and an inner nuclear membrane, which is underlain by the nuclear lamina that provides the nucleus with mechanical strength for maintaining structure and regulates chromatin organization for modulating gene expression and silencing. A layer of heterochromatin is beneath the nuclear lamina, attached by inner nuclear membrane integral proteins such as Lamin B receptor (LBR). LBR is a chimeric protein, having also a sterol reductase activity with which it contributes to cholesterol synthesis. Lukasova et al. showed that when DNA is damaged by ɣ-radiation in cancer cells, LBR is lost causing chromatin structure changes and promoting cellular senescence. Cellular senescence is characterized by terminal cell cycle arrest and the expression and secretion of various growth factors, cytokines, metalloproteinases, etc., collectively known as senescence-associated secretory phenotype (SASP) that cause chronic inflammation and tumor progression when they persist in the tissue. Therefore, it is fundamental to understand the molecular basis for senescence establishment, maintenance and the regulation of SASP. The work of Lukasova et al. contributed to our understanding of cellular senescence establishment and provided the basis that lead to the further discovery that chromatin changes caused by LBR reduction induce an up-regulated expression of SASP factors. LBR dysfunction has relevance in several diseases and possibly in physiological aging. The potential bifunctional role of LBR on cellular senescence establishment, namely its role in chromatin structure together with its enzymatic activity contributing to cholesterol synthesis, provide a new target to develop potential anti-aging therapies.

scholarly journals Nuclear envelope rupture: Actin fibers are putting the squeeze on the nucleus

2016 ◽  
Vol 215 (1) ◽  
pp. 5-8 ◽  
Author(s):  
Jan Lammerding ◽  
Katarina Wolf
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Nuclear Lamina ◽  
Cell Biol ◽  
Biophysical Processes

Cells exhibit transient nuclear envelope ruptures during interphase, but the responsible biophysical processes remain unclear. In this issue, Hatch and Hetzer (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201603053) show that actin fibers constrict the nucleus, causing chromatin protrusions and nuclear membrane ruptures at sites with nuclear lamina defects.

Sign in / Sign up

Export Citation Format

Share Document