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Parvoviruses cause nuclear envelope breakdown by activating key enzymes of mitosis.

Porwal M, Cohen S, Snoussi K, Popa-Wagner R, Anderson F, Dugot-Senant N, Wodrich H, Dinsart C, Kleinschmidt JA, Panté N, Kann M - PLoS Pathog. (2013)

Bottom Line: Activation and coordination of the different activities is poorly understood and moreover complicated as some factors translocate between cytoplasm and nucleus in preparatory phases.Consistent with Ca⁺⁺ efflux from the lumen between inner and outer nuclear membrane we found that Ca⁺⁺ was essential for nuclear disassembly by activating PKC.PKC activation then triggered activation of cdk-2, which became further activated by caspase-3.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Virology, University of Giessen, Giessen, Germany ; Univ. de Bordeaux, Microbiologie fondamentale et Pathogénicité, UMR 5234, Bordeaux, France ; CNRS, Microbiologie fondamentale et Pathogénicité, UMR 5234, Bordeaux, France.

ABSTRACT
Disassembly of the nuclear lamina is essential in mitosis and apoptosis requiring multiple coordinated enzymatic activities in nucleus and cytoplasm. Activation and coordination of the different activities is poorly understood and moreover complicated as some factors translocate between cytoplasm and nucleus in preparatory phases. Here we used the ability of parvoviruses to induce nuclear membrane breakdown to understand the triggers of key mitotic enzymes. Nuclear envelope disintegration was shown upon infection, microinjection but also upon their application to permeabilized cells. The latter technique also showed that nuclear envelope disintegration was independent upon soluble cytoplasmic factors. Using time-lapse microscopy, we observed that nuclear disassembly exhibited mitosis-like kinetics and occurred suddenly, implying a catastrophic event irrespective of cell- or type of parvovirus used. Analyzing the order of the processes allowed us to propose a model starting with direct binding of parvoviruses to distinct proteins of the nuclear pore causing structural rearrangement of the parvoviruses. The resulting exposure of domains comprising amphipathic helices was required for nuclear envelope disintegration, which comprised disruption of inner and outer nuclear membrane as shown by electron microscopy. Consistent with Ca⁺⁺ efflux from the lumen between inner and outer nuclear membrane we found that Ca⁺⁺ was essential for nuclear disassembly by activating PKC. PKC activation then triggered activation of cdk-2, which became further activated by caspase-3. Collectively our study shows a unique interaction of a virus with the nuclear envelope, provides evidence that a nuclear pool of executing enzymes is sufficient for nuclear disassembly in quiescent cells, and demonstrates that nuclear disassembly can be uncoupled from initial phases of mitosis.

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Schematic presentation of the parvoviral interaction with the nuclear envelope.A. Upon arrival at the nuclear pores parvoviral capsids (blue icosahedra) interact directly with at least three Nup (Nup358: dark grey; Nup153: light grey, Nup62: middle grey). B. This interaction causes exposure of VP1u on the surface of the virus. C. VP1u exposure allows permeabilization of the nuclear membrane, which is indicated by the dotted line of the NE (black lines). The permeabilization causes efflux of Ca++ indicated by the arrows, causing elevated local Ca++ in the nuclear periphery. D. Ca++ activated PKCα causing exposure of the catalytic domain (orange; PKCα-i: inactive; PKCα-a: active), which than phosphorylates lamins (blue; PKCα phosphorylation indicated by orange P). Further PKCα inactivates indirectly cdk-2, subsequently activating cdk-1 (indirect activations indicated by a dotted circle; cdks in red, cdk-i: inactive, cdk-a: active). Further activation is mediated by caspase-3, likely in an indirect manner. E. Active cdk1 hyper phosphorylates lamins, shown by red P. Hyper phosphorylation subsequently causes lamin depolymerisation. On combination with the spread of membrane disintegration the LBR (green) dissociates from the NE. F. After significant membrane disintegration even larger structures as chromatin (red waved lines) can escape the nucleus.
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ppat-1003671-g011: Schematic presentation of the parvoviral interaction with the nuclear envelope.A. Upon arrival at the nuclear pores parvoviral capsids (blue icosahedra) interact directly with at least three Nup (Nup358: dark grey; Nup153: light grey, Nup62: middle grey). B. This interaction causes exposure of VP1u on the surface of the virus. C. VP1u exposure allows permeabilization of the nuclear membrane, which is indicated by the dotted line of the NE (black lines). The permeabilization causes efflux of Ca++ indicated by the arrows, causing elevated local Ca++ in the nuclear periphery. D. Ca++ activated PKCα causing exposure of the catalytic domain (orange; PKCα-i: inactive; PKCα-a: active), which than phosphorylates lamins (blue; PKCα phosphorylation indicated by orange P). Further PKCα inactivates indirectly cdk-2, subsequently activating cdk-1 (indirect activations indicated by a dotted circle; cdks in red, cdk-i: inactive, cdk-a: active). Further activation is mediated by caspase-3, likely in an indirect manner. E. Active cdk1 hyper phosphorylates lamins, shown by red P. Hyper phosphorylation subsequently causes lamin depolymerisation. On combination with the spread of membrane disintegration the LBR (green) dissociates from the NE. F. After significant membrane disintegration even larger structures as chromatin (red waved lines) can escape the nucleus.

Mentions: In summary our data give evidence for a unique virus-mediated pathway that causes NEBD (see scheme Fig. 11). NEBD was observed for different PV and in cells ranging from human to Xenopus laevis implying an evolutionary well conserved phenomenon. Our model of PV host-interaction starts with the release of the viruses from the microtubules in the nuclear periphery, followed by (a) attachment to the NPC directly, which (b) subsequently causes exposure of VP1u. A yet undefined domain of VP1u then (c) permeabilizes the nuclear membranes leading to Ca++ efflux. (d) Ca++ activates nuclear PKCα, which phosphorylates lamins [56] and activates cdk-2, which becomes further activated by caspase-3 as described recently [63]. Activated cdk-2 – a key element of entry into mitosis - possibly leads to cdk-1 activation and (e) lamin A/C hyper phosphorylation [64]. (f) Lamin (hyper) phosphorylation than leads to lamin depolymerisation allowing entry and exit of large cargos from the nucleus by diffusion. At least in somatic cells the activation cascade must spread within the nucleus explaining why nuclear permeability increased suddenly, which is also a characteristic of mitosis.


Parvoviruses cause nuclear envelope breakdown by activating key enzymes of mitosis.

Porwal M, Cohen S, Snoussi K, Popa-Wagner R, Anderson F, Dugot-Senant N, Wodrich H, Dinsart C, Kleinschmidt JA, Panté N, Kann M - PLoS Pathog. (2013)

Schematic presentation of the parvoviral interaction with the nuclear envelope.A. Upon arrival at the nuclear pores parvoviral capsids (blue icosahedra) interact directly with at least three Nup (Nup358: dark grey; Nup153: light grey, Nup62: middle grey). B. This interaction causes exposure of VP1u on the surface of the virus. C. VP1u exposure allows permeabilization of the nuclear membrane, which is indicated by the dotted line of the NE (black lines). The permeabilization causes efflux of Ca++ indicated by the arrows, causing elevated local Ca++ in the nuclear periphery. D. Ca++ activated PKCα causing exposure of the catalytic domain (orange; PKCα-i: inactive; PKCα-a: active), which than phosphorylates lamins (blue; PKCα phosphorylation indicated by orange P). Further PKCα inactivates indirectly cdk-2, subsequently activating cdk-1 (indirect activations indicated by a dotted circle; cdks in red, cdk-i: inactive, cdk-a: active). Further activation is mediated by caspase-3, likely in an indirect manner. E. Active cdk1 hyper phosphorylates lamins, shown by red P. Hyper phosphorylation subsequently causes lamin depolymerisation. On combination with the spread of membrane disintegration the LBR (green) dissociates from the NE. F. After significant membrane disintegration even larger structures as chromatin (red waved lines) can escape the nucleus.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3814971&req=5

ppat-1003671-g011: Schematic presentation of the parvoviral interaction with the nuclear envelope.A. Upon arrival at the nuclear pores parvoviral capsids (blue icosahedra) interact directly with at least three Nup (Nup358: dark grey; Nup153: light grey, Nup62: middle grey). B. This interaction causes exposure of VP1u on the surface of the virus. C. VP1u exposure allows permeabilization of the nuclear membrane, which is indicated by the dotted line of the NE (black lines). The permeabilization causes efflux of Ca++ indicated by the arrows, causing elevated local Ca++ in the nuclear periphery. D. Ca++ activated PKCα causing exposure of the catalytic domain (orange; PKCα-i: inactive; PKCα-a: active), which than phosphorylates lamins (blue; PKCα phosphorylation indicated by orange P). Further PKCα inactivates indirectly cdk-2, subsequently activating cdk-1 (indirect activations indicated by a dotted circle; cdks in red, cdk-i: inactive, cdk-a: active). Further activation is mediated by caspase-3, likely in an indirect manner. E. Active cdk1 hyper phosphorylates lamins, shown by red P. Hyper phosphorylation subsequently causes lamin depolymerisation. On combination with the spread of membrane disintegration the LBR (green) dissociates from the NE. F. After significant membrane disintegration even larger structures as chromatin (red waved lines) can escape the nucleus.
Mentions: In summary our data give evidence for a unique virus-mediated pathway that causes NEBD (see scheme Fig. 11). NEBD was observed for different PV and in cells ranging from human to Xenopus laevis implying an evolutionary well conserved phenomenon. Our model of PV host-interaction starts with the release of the viruses from the microtubules in the nuclear periphery, followed by (a) attachment to the NPC directly, which (b) subsequently causes exposure of VP1u. A yet undefined domain of VP1u then (c) permeabilizes the nuclear membranes leading to Ca++ efflux. (d) Ca++ activates nuclear PKCα, which phosphorylates lamins [56] and activates cdk-2, which becomes further activated by caspase-3 as described recently [63]. Activated cdk-2 – a key element of entry into mitosis - possibly leads to cdk-1 activation and (e) lamin A/C hyper phosphorylation [64]. (f) Lamin (hyper) phosphorylation than leads to lamin depolymerisation allowing entry and exit of large cargos from the nucleus by diffusion. At least in somatic cells the activation cascade must spread within the nucleus explaining why nuclear permeability increased suddenly, which is also a characteristic of mitosis.

Bottom Line: Activation and coordination of the different activities is poorly understood and moreover complicated as some factors translocate between cytoplasm and nucleus in preparatory phases.Consistent with Ca⁺⁺ efflux from the lumen between inner and outer nuclear membrane we found that Ca⁺⁺ was essential for nuclear disassembly by activating PKC.PKC activation then triggered activation of cdk-2, which became further activated by caspase-3.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Virology, University of Giessen, Giessen, Germany ; Univ. de Bordeaux, Microbiologie fondamentale et Pathogénicité, UMR 5234, Bordeaux, France ; CNRS, Microbiologie fondamentale et Pathogénicité, UMR 5234, Bordeaux, France.

ABSTRACT
Disassembly of the nuclear lamina is essential in mitosis and apoptosis requiring multiple coordinated enzymatic activities in nucleus and cytoplasm. Activation and coordination of the different activities is poorly understood and moreover complicated as some factors translocate between cytoplasm and nucleus in preparatory phases. Here we used the ability of parvoviruses to induce nuclear membrane breakdown to understand the triggers of key mitotic enzymes. Nuclear envelope disintegration was shown upon infection, microinjection but also upon their application to permeabilized cells. The latter technique also showed that nuclear envelope disintegration was independent upon soluble cytoplasmic factors. Using time-lapse microscopy, we observed that nuclear disassembly exhibited mitosis-like kinetics and occurred suddenly, implying a catastrophic event irrespective of cell- or type of parvovirus used. Analyzing the order of the processes allowed us to propose a model starting with direct binding of parvoviruses to distinct proteins of the nuclear pore causing structural rearrangement of the parvoviruses. The resulting exposure of domains comprising amphipathic helices was required for nuclear envelope disintegration, which comprised disruption of inner and outer nuclear membrane as shown by electron microscopy. Consistent with Ca⁺⁺ efflux from the lumen between inner and outer nuclear membrane we found that Ca⁺⁺ was essential for nuclear disassembly by activating PKC. PKC activation then triggered activation of cdk-2, which became further activated by caspase-3. Collectively our study shows a unique interaction of a virus with the nuclear envelope, provides evidence that a nuclear pool of executing enzymes is sufficient for nuclear disassembly in quiescent cells, and demonstrates that nuclear disassembly can be uncoupled from initial phases of mitosis.

Show MeSH
Related in: MedlinePlus