scholarly journals SINC, a type III secreted protein of Chlamydia psittaci, targets the inner nuclear membrane of infected cells and uninfected neighbors

2015 ◽  
Vol 26 (10) ◽  
pp. 1918-1934 ◽  
Author(s):  
Sergio A. Mojica ◽  
Kelley M. Hovis ◽  
Matthew B. Frieman ◽  
Bao Tran ◽  
Ru-ching Hsia ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Nuclear Lamina ◽  
Chlamydia Psittaci ◽  
Hek293 Cells ◽  
Secreted Protein ◽  
Inner Nuclear Membrane ◽  
Type Iii ◽  
Infected Cells ◽  
Digitonin Permeabilization

SINC, a new type III secreted protein of the avian and human pathogen Chlamydia psittaci, uniquely targets the nuclear envelope of C. psittaci–infected cells and uninfected neighboring cells. Digitonin-permeabilization studies of SINC-GFP–transfected HeLa cells indicate that SINC targets the inner nuclear membrane. SINC localization at the nuclear envelope was blocked by importazole, confirming SINC import into the nucleus. Candidate partners were identified by proximity to biotin ligase-fused SINC in HEK293 cells and mass spectrometry (BioID). This strategy identified 22 candidates with high confidence, including the nucleoporin ELYS, lamin B1, and four proteins (emerin, MAN1, LAP1, and LBR) of the inner nuclear membrane, suggesting that SINC interacts with host proteins that control nuclear structure, signaling, chromatin organization, and gene silencing. GFP-SINC association with the native LEM-domain protein emerin, a conserved component of nuclear “lamina” structure, or with a complex containing emerin was confirmed by GFP pull down. Our findings identify SINC as a novel bacterial protein that targets the nuclear envelope with the capability of globally altering nuclear envelope functions in the infected host cell and neighboring uninfected cells. These properties may contribute to the aggressive virulence of C. psittaci.

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 Fibroblasts lacking nuclear lamins do not have nuclear blebs or protrusions but nevertheless have frequent nuclear membrane ruptures

2018 ◽  
Vol 115 (40) ◽  
pp. 10100-10105 ◽  
Author(s):  
Natalie Y. Chen ◽  
Paul Kim ◽  
Thomas A. Weston ◽  
Lovelyn Edillo ◽  
Yiping Tu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Membrane Integrity ◽  
Nuclear Lamina ◽  
Virtual Absence ◽  
Inner Nuclear Membrane ◽  
Nuclear Lamins ◽  
Transmission Electron ◽  
Nuclear Lamin

The nuclear lamina, an intermediate filament meshwork lining the inner nuclear membrane, is formed by the nuclear lamins (lamins A, C, B1, and B2). Defects or deficiencies in individual nuclear lamin proteins have been reported to elicit nuclear blebs (protrusions or outpouchings of the nuclear envelope) and increase susceptibility for nuclear membrane ruptures. It is unclear, however, how a complete absence of nuclear lamins would affect nuclear envelope morphology and nuclear membrane integrity (i.e., whether nuclear membrane blebs or protrusions would occur and, if not, whether cells would be susceptible to nuclear membrane ruptures). To address these issues, we generated mouse embryonic fibroblasts (MEFs) lacking all nuclear lamins. The nuclear lamin-deficient MEFs had irregular nuclear shapes but no nuclear blebs or protrusions. Despite a virtual absence of nuclear blebs, MEFs lacking nuclear lamins had frequent, prolonged, and occasionally nonhealing nuclear membrane ruptures. By transmission electron microscopy, the inner nuclear membrane in nuclear lamin-deficient MEFs have a “wavy” appearance, and there were discrete discontinuities in the inner and outer nuclear membranes. Nuclear membrane ruptures were accompanied by a large increase in DNA damage, as judged by γ-H2AX foci. Mechanical stress increased both nuclear membrane ruptures and DNA damage, whereas minimizing transmission of cytoskeletal forces to the nucleus had the opposite effects.

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 Characterization of A 54-kD protein of the inner nuclear membrane: evidence for cell cycle-dependent interaction with the nuclear lamina.

1991 ◽  
Vol 114 (3) ◽  
pp. 389-400 ◽  
Author(s):  
S M Bailer ◽  
H M Eppenberger ◽  
G Griffiths ◽  
E A Nigg
Keyword(s):  
Cell Cycle ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Cultured Cells ◽  
Nuclear Lamina ◽  
Biochemical Properties ◽  
Inner Nuclear Membrane ◽  
Cycle Dependent ◽  
Mitotic Phosphorylation

Using a mAb (R-7), we have characterized a 54-kD protein of the chicken nuclear envelope. Based on its biochemical properties and subnuclear distribution p54 is likely to be an integral membrane component specific to the inner nuclear membrane. Fractionation experiments indicate that p54 interacts, directly or indirectly, with the nuclear lamina, and analysis of p54 in cultured cells suggests that this interaction is controlled by cell cycle-dependent posttranslational modification, most likely phosphorylation. Modification of p54 results in a slightly reduced electrophoretic mobility, and it converts the protein from a detergent-resistant to a detergent-extractable form. Detergent solubilization of p54 can be induced in vivo by treating isolated nuclei or nuclear envelopes with highly purified cdc2 kinase, one of the most prominent kinases active in mitotic cells. These results suggest that mitotic phosphorylation of p54 might contribute to control nuclear envelope dynamics during mitosis in vivo.

scholarly journals Fate of the Inner Nuclear Membrane Protein Lamin B Receptor and Nuclear Lamins in Herpes Simplex Virus Type 1 Infection

2001 ◽  
Vol 75 (18) ◽  
pp. 8818-8830 ◽  
Author(s):  
Emily S. Scott ◽  
Peter O'Hare
Keyword(s):  
Herpes Simplex ◽  
Nuclear Membrane ◽  
Nuclear Lamina ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Lamin B Receptor ◽  
Infected Cells ◽  
Simplex Virus ◽  
Inner Nuclear Membrane Protein

ABSTRACT During herpesvirus egress, capsids bud through the inner nuclear membrane. Underlying this membrane is the nuclear lamina, a meshwork of intermediate filaments with which it is tightly associated. Details of alterations to the lamina and the inner nuclear membrane during infection and the mechanisms involved in capsid transport across these structures remain unclear. Here we describe the fate of key protein components of the nuclear envelope and lamina during herpes simplex virus type 1 (HSV-1) infection. We followed the distribution of the inner nuclear membrane protein lamin B receptor (LBR) and lamins A and B2 tagged with green fluorescent protein (GFP) in live infected cells. Together with additional results from indirect immunofluorescence, our studies reveal major morphologic distortion of nuclear-rim LBR and lamins A/C, B1, and B2. By 8 h p.i., we also observed a significant redistribution of LBR-GFP to the endoplasmic reticulum, where it colocalized with a subpopulation of cytoplasmic glycoprotein B by immunofluorescence. In addition, analysis by fluorescence recovery after photobleaching reveals that LBR-GFP exhibited increased diffusional mobility within the nuclear membrane of infected cells. This is consistent with the disruption of interactions between LBR and the underlying lamina. In addition to studying stably expressed GFP-lamins by fluorescence microscopy, we studied endogenous A- and B-type lamins in infected cells by Western blotting. Both approaches reveal a loss of lamins associated with virus infection. These data indicate major disruption of the nuclear envelope and lamina of HSV-1-infected cells and are consistent with a virus-induced dismantling of the nuclear lamina, possibly in order to gain access to the inner nuclear membrane.

scholarly journals Synthesis, transport and incorporation into the nuclear envelope of A-type lamins and inner nuclear membrane proteins

2011 ◽  
Vol 39 (6) ◽  
pp. 1758-1763 ◽  
Author(s):  
Jose M. González ◽  
Vicente Andrés
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Complex Structure ◽  
Nuclear Lamina ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Inner Nuclear Membrane ◽  
Perinuclear Space ◽  
Interphase Cells ◽  
Pore Complexes

The mammalian NE (nuclear envelope), which separates the nucleus from the cytoplasm, is a complex structure composed of nuclear pore complexes, the outer and inner nuclear membranes, the perinuclear space and the nuclear lamina (A- and B-type lamins). The NE is completely disassembled and reassembled at each cell division. In the present paper, we review recent advances in the understanding of the mechanisms implicated in the transport of inner nuclear membrane and nuclear lamina proteins from the endoplasmic reticulum to the nucleus in interphase cells and mitosis, with special attention to A-type lamins.

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.

scholarly journals Unrestrained ESCRT-III drives chromosome fragmentation and micronuclear catastrophe

2019 ◽  
Author(s):  
Marina Vietri ◽  
Sebastian W. Schultz ◽  
Aurélie Bellanger ◽  
Carl M. Jones ◽  
Camilla Raiborg ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Genome Stability ◽  
Membrane Deformation ◽  
Chromosome Fragmentation ◽  
Inner Nuclear Membrane ◽  
Membrane Rupture ◽  
Membrane Fission ◽  
Stable Association ◽  
Inner Nuclear Membrane Protein

AbstractThe ESCRT-III membrane fission machinery1,2 restores nuclear envelope integrity during mitotic exit3,4 and interphase5,6. Whereas primary nuclei resealing takes minutes, micronuclear envelope ruptures appear irreversible and result in catastrophic collapse associated with chromosome fragmentation and rearrangements (chromothripsis), thought to be a major driving force in cancer development7-10. Despite its importance11-13, the mechanistic underpinnings of nuclear envelope sealing in primary nuclei and the defects observed in micronuclei remain largely unknown. Here we show that CHMP7, the nucleator of ESCRT-III filaments at the nuclear envelope3,14, and the inner nuclear membrane protein LEMD215 act as a compartmentalization sensor detecting the loss of nuclear integrity. In cells with intact nuclear envelope, CHMP7 is actively excluded from the nucleus to preclude its binding to LEMD2. Nuclear influx of CHMP7 results in stable association with LEMD2 at the inner nuclear membrane that licenses local polymerization of ESCRT-III. Tight control of nuclear CHMP7 levels is critical, as induction of nuclear CHMP7 mutants is sufficient to induce unrestrained growth of ESCRT-III foci at the nuclear envelope, causing dramatic membrane deformation, local DNA torsional stress, single-stranded DNA formation and fragmentation of the underlying chromosomes. At micronuclei, membrane rupture is not associated with repair despite timely recruitment of ESCRT-III. Instead, micronuclei inherently lack the capacity to restrict accumulation of CHMP7 and LEMD2. This drives unrestrained ESCRT-III recruitment, membrane deformation and DNA defects that strikingly resemble those at primary nuclei upon induction of nuclear CHMP7 mutants. Preventing ESCRT-III recruitment suppresses membrane deformation and DNA damage, without restoring nucleocytoplasmic compartmentalization. We propose that the ESCRT-III nuclear integrity surveillance machinery is a double-edged sword, as its exquisite sensitivity ensures rapid repair at primary nuclei while causing unrestrained polymerization at micronuclei, with catastrophic consequences for genome stability16-18.

scholarly journals Nuclear envelope rupture is induced by actin-based nucleus confinement

2016 ◽  
Vol 215 (1) ◽  
pp. 27-36 ◽  
Author(s):  
Emily M. Hatch ◽  
Martin W. Hetzer
Keyword(s):  
Nuclear Envelope ◽  
Cancer Cells ◽  
Nuclear Membrane ◽  
Nuclear Lamina ◽  
Membrane Stability ◽  
Linc Complex ◽  
Actin Bundles ◽  
In Cells

Repeated rounds of nuclear envelope (NE) rupture and repair have been observed in laminopathy and cancer cells and result in intermittent loss of nucleus compartmentalization. Currently, the causes of NE rupture are unclear. Here, we show that NE rupture in cancer cells relies on the assembly of contractile actin bundles that interact with the nucleus via the linker of nucleoskeleton and cytoskeleton (LINC) complex. We found that the loss of actin bundles or the LINC complex did not rescue nuclear lamina defects, a previously identified determinant of nuclear membrane stability, but did decrease the number and size of chromatin hernias. Finally, NE rupture inhibition could be rescued in cells treated with actin-depolymerizing drugs by mechanically constraining nucleus height. These data suggest a model of NE rupture where weak membrane areas, caused by defects in lamina organization, rupture because of an increase in intranuclear pressure from actin-based nucleus confinement.

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