Nuclear envelope rupture is induced by actin-based nucleus confinement

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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Nuclear envelope organization in Dictyostelium discoideum

The International Journal of Developmental Biology ◽

10.1387/ijdb.190184rg ◽

2019 ◽

Vol 63 (8-9-10) ◽

pp. 509-519 ◽

Cited By ~ 3

Author(s):

Petros Batsios ◽

Ralph Gräf ◽

Michael P. Koonce ◽

Denis A. Larochelle ◽

Irene Meyer

Keyword(s):

Nuclear Envelope ◽

Nuclear Membrane ◽

Nuclear Lamina ◽

Major Constituent ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Linc Complex ◽

Family Protein ◽

Import And Export ◽

Pore Complexes

The nuclear envelope consists of the outer and the inner nuclear membrane, the nuclear lamina and the nuclear pore complexes, which regulate nuclear import and export. The major constituent of the nuclear lamina of Dictyostelium is the lamin NE81. It can form filaments like B-type lamins and it interacts with Sun1, as well as with the LEM/HeH-family protein Src1. Sun1 and Src1 are nuclear envelope transmembrane proteins involved in the centrosome-nucleus connection and nuclear envelope stability at the nucleolar regions, respectively. In conjunction with a KASH-domain protein, Sun1 usually forms a so-called LINC complex. Two proteins with functions reminiscent of KASH-domain proteins at the outer nuclear membrane of Dictyostelium are known; interaptin which serves as an actin connector and the kinesin Kif9 which plays a role in the microtubule-centrosome connector. However, both of these lack the conserved KASH-domain. The link of the centrosome to the nuclear envelope is essential for the insertion of the centrosome into the nuclear envelope and the appropriate spindle formation. Moreover, centrosome insertion is involved in permeabilization of the mitotic nucleus, which ensures access of tubulin dimers and spindle assembly factors. Our recent progress in identifying key molecular players at the nuclear envelope of Dictyostelium promises further insights into the mechanisms of nuclear envelope dynamics.

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Mena regulates the LINC complex to control actin–nuclear lamina associations, trans-nuclear membrane signalling and cancer gene expression

10.1101/2021.08.31.458340 ◽

2021 ◽

Author(s):

Frederic Li Mow Chee ◽

Bruno Beernaert ◽

Alexander E P Loftus ◽

Yatendra Kumar ◽

Billie G C Griffith ◽

...

Keyword(s):

Gene Expression ◽

Nuclear Envelope ◽

Cancer Progression ◽

Nuclear Membrane ◽

Regulatory Protein ◽

Nuclear Periphery ◽

Nuclear Lamina ◽

Actin Binding ◽

Cancer Gene ◽

Linc Complex

Interactions between cells and the extracellular matrix, mediated by integrin adhesion complexes (IACs), play key roles in cancer progression and metastasis. We investigated systems-level changes in the integrin adhesome during metastatic progression of a patient-derived cutaneous squamous cell carcinoma (cSCC), and found that the actin regulatory protein Mena is enriched in IACs in metastatic cSCC cells. Mena is connected within a subnetwork of actin-binding proteins to the LINC complex component nesprin-2, with which it interacts and co-localises at the nuclear envelope of metastatic cells. Moreover, Mena potentiates the interactions of nesprin-2 with the actin cytoskeleton and the nuclear lamina. CRISPR-mediated Mena depletion causes altered nuclear morphology, reduces tyrosine phosphorylation of the nuclear membrane protein emerin and downregulates expression of the immunomodulatory gene PTX3 via the recruitment of its enhancer to the nuclear periphery. We have uncovered an unexpected novel role for Mena at the nuclear membrane, where it controls the LINC complex, nuclear architecture, chromatin repositioning and cancer gene expression. This is the first description of an adhesion protein regulating gene transcription via direct signalling across the nuclear envelope.

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

The Journal of Cell Biology ◽

10.1083/jcb.201609102 ◽

2016 ◽

Vol 215 (1) ◽

pp. 5-8 ◽

Cited By ~ 30

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.

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A Balance Between Intermediate Filaments and Microtubules Maintains Nuclear Architecture in the Cardiomyocyte

Circulation Research ◽

10.1161/circresaha.119.315582 ◽

2020 ◽

Vol 126 (3) ◽

Cited By ~ 6

Author(s):

Julie Heffler ◽

Parisha P. Shah ◽

Patrick Robison ◽

Sai Phyo ◽

Kimberly Veliz ◽

...

Keyword(s):

Nuclear Envelope ◽

Intermediate Filaments ◽

Genome Organization ◽

Contractile Function ◽

Nuclear Architecture ◽

Super Resolution ◽

Nuclear Lamina ◽

Linc Complex ◽

Spectrin Repeat ◽

Balance Of Forces

Rationale: Mechanical forces are transduced to nuclear responses via the linkers of the nucleoskeleton and cytoskeleton (LINC) complex, which couples the cytoskeleton to the nuclear lamina and associated chromatin. While disruption of the LINC complex can cause cardiomyopathy, the relevant interactions that bridge the nucleoskeleton to cytoskeleton are poorly understood in the cardiomyocyte, where cytoskeletal organization is unique. Furthermore, while microtubules and desmin intermediate filaments associate closely with cardiomyocyte nuclei, the importance of these interactions is unknown. Objective: Here, we sought to determine how cytoskeletal interactions with the LINC complex regulate nuclear homeostasis in the cardiomyocyte. Methods and Results: To this end, we acutely disrupted the LINC complex, microtubules, actin, and intermediate filaments and assessed the consequences on nuclear morphology and genome organization in rat ventricular cardiomyocytes via a combination of super-resolution imaging, biophysical, and genomic approaches. We find that a balance of dynamic microtubules and desmin intermediate filaments is required to maintain nuclear shape and the fidelity of the nuclear envelope and lamina. Upon depletion of desmin (or nesprin [nuclear envelope spectrin repeat protein]-3, its binding partner in the LINC complex), polymerizing microtubules collapse the nucleus and drive infolding of the nuclear membrane. This results in DNA damage, a loss of genome organization, and broad transcriptional changes. The collapse in nuclear integrity is concomitant with compromised contractile function and may contribute to the pathophysiological changes observed in desmin-related myopathies. Conclusions: Disrupting the tethering of desmin to the nucleus results in a loss of nuclear homeostasis and rapid alterations to cardiomyocyte function. Our data suggest that a balance of forces imposed by intermediate filaments and microtubules is required to maintain nuclear structure and genome organization in the cardiomyocyte.

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

Molecular Biology of the Cell ◽

10.1091/mbc.e14-11-1530 ◽

2015 ◽

Vol 26 (10) ◽

pp. 1918-1934 ◽

Cited By ~ 35

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.

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Integrity of the Linker of Nucleoskeleton and Cytoskeleton Is Required for Efficient Herpesvirus Nuclear Egress

Journal of Virology ◽

10.1128/jvi.00330-17 ◽

2017 ◽

Vol 91 (19) ◽

Cited By ~ 8

Author(s):

Barbara G. Klupp ◽

Teresa Hellberg ◽

Harald Granzow ◽

Kati Franzke ◽

Beatriz Dominguez Gonzalez ◽

...

Keyword(s):

Nuclear Membrane ◽

Dominant Negative ◽

Progeny Virus ◽

Linc Complex ◽

Outer Nuclear Membrane ◽

Inner Nuclear Membrane ◽

Nuclear Membranes ◽

Nuclear Egress ◽

Virion Envelope ◽

In Cells

ABSTRACT Herpesvirus capsids assemble in the nucleus, while final virion maturation proceeds in the cytoplasm. This requires that newly formed nucleocapsids cross the nuclear envelope (NE), which occurs by budding at the inner nuclear membrane (INM), release of the primary enveloped virion into the perinuclear space (PNS), and subsequent rapid fusion with the outer nuclear membrane (ONM). During this process, the NE remains intact, even at late stages of infection. In addition, the spacing between the INM and ONM is maintained, as is that between the primary virion envelope and nuclear membranes. The linker of nucleoskeleton and cytoskeleton (LINC) complex consists of INM proteins with a luminal SUN (Sad1/UNC-84 homology) domain connected to ONM proteins with a KASH (Klarsicht, ANC-1, SYNE homology) domain and is thought to be responsible for spacing the nuclear membranes. To investigate the role of the LINC complex during herpesvirus infection, we generated cell lines constitutively expressing dominant negative (dn) forms of SUN1 and SUN2. Ultrastructural analyses revealed a significant expansion of the PNS and the contiguous intracytoplasmic lumen, most likely representing endoplasmic reticulum (ER), especially in cells expressing dn-SUN2. After infection, primary virions accumulated in these expanded luminal regions, also very distant from the nucleus. The importance of the LINC complex was also confirmed by reduced progeny virus titers in cells expressing dn-SUN2. These data show that the intact LINC complex is required for efficient nuclear egress of herpesviruses, likely acting to promote fusion of primary enveloped virions with the ONM. IMPORTANCE While the viral factors for primary envelopment of nucleocapsids at the inner nuclear membrane are known to the point of high-resolution structures, the roles of cellular components and regulators remain enigmatic. Furthermore, the machinery responsible for fusion with the outer nuclear membrane is unsolved. We show here that dominant negative SUN2 interferes with efficient herpesvirus nuclear egress, apparently by interfering with fusion between the primary virion envelope and outer nuclear membrane. This identifies a new cellular component important for viral egress and implicates LINC complex integrity in nonconventional nuclear membrane trafficking.

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Decreased torsinA or LINC Complex Function Rescues a Laminopathy in Caenorhabditis elegans

10.1101/2020.04.22.055988 ◽

2020 ◽

Author(s):

Gabriela Huelgas-Morales ◽

Mark Sanders ◽

Gemechu Mekonnen ◽

Tatsuya Tsukamoto ◽

David Greenstein

Keyword(s):

Nuclear Envelope ◽

Complex Function ◽

Nuclear Lamina ◽

Premature Aging ◽

Linc Complex ◽

Force Transmission ◽

Complex Activity ◽

Aaa Atpase ◽

C Elegans ◽

Associated Proteins

AbstractThe function of the nucleus depends on the integrity of the nuclear lamina, an intermediate filament network associated with the linker of nucleoskeleton and cytoskeleton (LINC) complex spanning the nuclear envelope. In turn, the AAA+ ATPase torsinA regulates force transmission from the cytoskeleton to the nucleus. In humans, mutations affecting nuclear envelope-associated proteins cause laminopathies, including progeria, myopathy, and dystonia. We report that decreasing the function of the C. elegans torsinA homolog, OOC-5, rescues the sterility and premature aging caused by a null mutation in the single worm lamin homolog, lmn-1. Loss of OOC-5 activity prevents nuclear collapse in lmn-1 mutants by disrupting the function of the LINC complex. These results suggest that LINC complex-transmitted forces damage nuclei with a compromised nuclear lamina.One Sentence SummaryInhibiting LINC complex activity prevents a progeric syndrome in C. elegans.

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Chromosome length and gene density contribute to micronuclear membrane stability

10.1101/2021.05.12.443914 ◽

2021 ◽

Author(s):

Anna Mammel ◽

Heather Z Huang ◽

Amanda L Gunn ◽

Emma Choo ◽

Emily M Hatch

Keyword(s):

Nuclear Envelope ◽

Membrane Integrity ◽

Nuclear Lamina ◽

Chromosome Length ◽

Gene Density ◽

Membrane Stability ◽

Nuclear Pore ◽

Membrane Rupture ◽

Lamin B1 ◽

Nuclear Envelope Assembly

Micronuclei are derived from missegregated chromosomes and frequently lose membrane integrity, leading to DNA damage, innate immune activation, and metastatic signaling. Here we demonstrate that two characteristics of the trapped chromosome, length and gene density, are key contributors to micronuclei membrane stability in human cells. Chromosome length is proportional to micronuclei size, and gene density has an additive effect with micronucleus size on membrane stability. We demonstrate that these results are not due to chromosome-specific differences in spindle position or initial nuclear pore complex recruitment during post-mitotic nuclear envelope assembly. We find that chromosome length and micronuclei size strongly correlate with lamin B1 and nuclear pore density in intact micronuclei. Unexpectedly, lamin B1 levels do not predict nuclear lamina organization and membrane stability. Instead, small gene-dense micronuclei have decreased nuclear lamina gaps compared to large micronuclei, despite very low levels of lamin B1. Our data strongly suggest that nuclear envelope composition defects previously correlated with membrane rupture only partly explain membrane stability in micronuclei. We propose that an unknown factor linked to gene density has a separate function that inhibits the appearance of nuclear lamina gaps and delays membrane rupture until late in the cell cycle.

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Chromosome length and gene density contribute to micronuclear membrane stability

Life Science Alliance ◽

10.26508/lsa.202101210 ◽

2021 ◽

Vol 5 (2) ◽

pp. e202101210

Author(s):

Anna E Mammel ◽

Heather Z Huang ◽

Amanda L Gunn ◽

Emma Choo ◽

Emily M Hatch

Keyword(s):

Nuclear Envelope ◽

Membrane Integrity ◽

Nuclear Lamina ◽

Chromosome Length ◽

Gene Density ◽

Membrane Stability ◽

Nuclear Pore ◽

Membrane Rupture ◽

Lamin B1 ◽

Nuclear Envelope Assembly

Micronuclei are derived from missegregated chromosomes and frequently lose membrane integrity, leading to DNA damage, innate immune activation, and metastatic signaling. Here, we demonstrate that two characteristics of the trapped chromosome, length and gene density, are key contributors to micronuclei membrane stability and determine the timing of micronucleus rupture. We demonstrate that these results are not due to chromosome-specific differences in spindle position or initial protein recruitment during post-mitotic nuclear envelope assembly. Micronucleus size strongly correlates with lamin B1 levels and nuclear pore density in intact micronuclei, but, unexpectedly, lamin B1 levels do not completely predict nuclear lamina organization or membrane stability. Instead, small gene-dense micronuclei have decreased nuclear lamina gaps compared to large micronuclei, despite very low levels of lamin B1. Our data strongly suggest that nuclear envelope composition defects previously correlated with membrane rupture only partly explain membrane stability in micronuclei. We propose that an unknown factor linked to gene density has a separate function that inhibits the appearance of nuclear lamina gaps and delays membrane rupture until late in the cell cycle.

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Decreased mechanotransduction prevents nuclear collapse in aCaenorhabditis eleganslaminopathy

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2015050117 ◽

2020 ◽

Vol 117 (49) ◽

pp. 31301-31308

Author(s):

Gabriela Huelgas-Morales ◽

Mark Sanders ◽

Gemechu Mekonnen ◽

Tatsuya Tsukamoto ◽

David Greenstein

Keyword(s):

Nuclear Envelope ◽

Complex Function ◽

Nuclear Lamina ◽

Premature Aging ◽

Nuclear Pore ◽

Linc Complex ◽

Force Transmission ◽

Cell Nuclei ◽

Aaa Atpase ◽

C Elegans

The function of the nucleus depends on the integrity of the nuclear lamina, an intermediate filament network associated with the linker of nucleoskeleton and cytoskeleton (LINC) complex. The LINC complex spans the nuclear envelope and mediates nuclear mechanotransduction, the process by which mechanical signals and forces are transmitted across the nuclear envelope. In turn, the AAA+ ATPase torsinA is thought to regulate force transmission from the cytoskeleton to the nucleus. In humans, mutations affecting nuclear envelope-associated proteins cause laminopathies, including progeria, myopathy, and dystonia, though the extent to which endogenous mechanical stresses contribute to these pathologies is unclear. Here, we use theCaenorhabditis elegansgermline as a model to investigate mechanisms that maintain nuclear integrity as germ cell nuclei progress through meiotic development and migrate for gametogenesis—processes that require LINC complex function. We report that decreasing the function of theC. eleganstorsinA homolog, OOC-5, rescues the sterility and premature aging caused by a null mutation in the single worm lamin homolog. We show that decreasing OOC-5/torsinA activity prevents nuclear collapse in lamin mutants by disrupting the function of the LINC complex. At a mechanistic level, OOC-5/torsinA promotes the assembly or maintenance of the lamin-associated LINC complex and this activity is also important for interphase nuclear pore complex insertion into growing germline nuclei. These results demonstrate that LINC complex-transmitted forces damage nuclei with a compromised nuclear lamina. Thus, the torsinA–LINC complex nexus might comprise a therapeutic target for certain laminopathies by preventing damage from endogenous cellular forces.

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