scholarly journals Monoclonal antibodies identify a group of nuclear pore complex glycoproteins.

1987 ◽  
Vol 104 (5) ◽  
pp. 1143-1156 ◽  
Author(s):  
C M Snow ◽  
A Senior ◽  
L Gerace
Keyword(s):  
Monoclonal Antibodies ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Peptide Mapping ◽  
Polyclonal Antibodies ◽  
Integral Membrane Protein ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Surface ◽  
Punctate Pattern

Using monoclonal antibodies we identified a group of eight polypeptides of rat liver nuclear envelopes that have common epitopes. Most or all of these proteins are structurally distinct, as shown by tryptic peptide mapping and analysis with polyclonal antibodies. While these polypeptides are relatively tightly bound to nuclear membranes, only one is an integral membrane protein. The eight antigens cofractionate with the nuclear pore complex under various conditions of ionic strength and detergent. It can be seen by immunofluorescence microscopy that the monoclonal antibodies reacting with these antigens stain the nuclear surface of interphase cells in a finely punctate pattern. When the nuclear envelope is disassembled and subsequently reformed during mitosis, the proteins are reversibly dispersed throughout the cytoplasm in the form of minute foci. By EM immunogold localization on isolated nuclear envelopes, the monoclonal antibodies label exclusively the nuclear pore complex, at both its nucleoplasmic and cytoplasmic margins. Considered together, our biochemical and localization data indicate that the eight nuclear envelope polypeptides are pore complex components. As shown in the accompanying paper (Holt, G. D., C. M. Snow, A. Senior, R. S. Haltiwanger, L. Gerace, and G. W. Hart, J. Cell Biol., 104:1157-1164) these eight polypeptides contain a novel form of glycosylation, O-linked N-acetylglucosamine. The relative abundance and disposition of these O-linked glycoproteins in the pore complex are consistent with their having a role in nucleocytoplasmic transport.

scholarly journals Lamin A, lamin B, and lamin B receptor analogues in yeast.

1989 ◽  
Vol 108 (6) ◽  
pp. 2069-2082 ◽  
Author(s):  
S D Georgatos ◽  
I Maroulakou ◽  
G Blobel
Keyword(s):  
Nuclear Envelope ◽  
Peptide Mapping ◽  
Polyclonal Antibodies ◽  
Structural Similarity ◽  
Integral Membrane Protein ◽  
Lamin A ◽  
Nuclear Fraction ◽  
Yeast Saccharomyces Cerevisiae ◽  
Lamin B ◽  
Lamin B Receptor

Previous studies have shown that turkey erythrocyte lamin B is anchored to the nuclear envelope via a 58-kD integral membrane protein termed p58 or lamin B receptor (Worman H. J., J. Yuan, G. Blobel, and S. D. Georgatos. 1988. Proc. Natl. Acad. Sci. USA. 85:8531-8534). We now identify a p58 analogue in the yeast Saccharomyces cerevisiae. Turkey erythrocyte lamin B binds to yeast urea-extracted nuclear envelopes with high affinity, associating predominantly with a 58-kD polypeptide. This yeast polypeptide is recognized by polyclonal antibodies against turkey p58, partitions entirely with the nuclear fraction, remains membrane bound after urea extraction of the nuclear envelopes, and is structurally similar to turkey p58 by peptide mapping criteria. Using polyclonal antibodies against turkey erythrocyte lamins A and B, we also identify two yeast lamin forms. The yeast lamin B analogue has a molecular mass of 66 kD and is structurally related to erythrocyte lamin B. Moreover, the yeast lamin B analogue partitions exclusively with the nuclear envelope fraction, is quantitatively removed from the envelopes by urea extraction, and binds to turkey lamin A and vimentin. As many higher eukaryotic lamin B forms, the yeast analogue is chemically heterogeneous comprising two serologically related species with different charge characteristics. Antibodies against turkey lamin A detect a 74-kD yeast protein, slightly larger than the turkey lamin A. It is more abundant than the yeast lamin B analogue and partitions between a soluble cytoplasmic fraction and a nuclear envelope fraction. The yeast lamin A analogue can be extracted from the nuclear envelope by urea, shows structural similarity to turkey and rat lamin A, and binds to isolated turkey lamin B. These data indicate that analogues of typical nuclear lamina components (lamins A and B, as well as lamin B receptor) are present in yeast and behave as their vertebrate counterparts.

Reticulon-like REEP4 at the inner nuclear membrane promotes nuclear pore complex formation

2021 ◽  
Vol 221 (2) ◽  
Author(s):  
Banafsheh Golchoubian ◽  
Andreas Brunner ◽  
Helena Bragulat-Teixidor ◽  
Annett Neuner ◽  
Busra A. Akarlar ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Nuclear Membrane ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Inner Nuclear Membrane ◽  
Pore Complexes ◽  
Late Stages ◽  
Protein Complex Assembly

Nuclear pore complexes (NPCs) are channels within the nuclear envelope that mediate nucleocytoplasmic transport. NPCs form within the closed nuclear envelope during interphase or assemble concomitantly with nuclear envelope reformation in late stages of mitosis. Both interphase and mitotic NPC biogenesis require coordination of protein complex assembly and membrane deformation. During early stages of mitotic NPC assembly, a seed for new NPCs is established on chromatin, yet the factors connecting the NPC seed to the membrane of the forming nuclear envelope are unknown. Here, we report that the reticulon homology domain protein REEP4 not only localizes to high-curvature membrane of the cytoplasmic endoplasmic reticulum but is also recruited to the inner nuclear membrane by the NPC biogenesis factor ELYS. This ELYS-recruited pool of REEP4 promotes NPC assembly and appears to be particularly important for NPC formation during mitosis. These findings suggest a role for REEP4 in coordinating nuclear envelope reformation with mitotic NPC biogenesis.

scholarly journals A short perinuclear amphipathic α-helix in Apq12 promotes nuclear pore complex biogenesis

2021 ◽  
Author(s):  
Elmar Schiebel ◽  
Wanlu Zhang ◽  
Azqa Khan ◽  
Jlenia Vitale ◽  
Annett Neuner ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Phosphatidic Acid ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Nuclear Membrane ◽  
Integral Membrane Protein ◽  
Nuclear Pore ◽  
Binding Properties ◽  
Perinuclear Space ◽  
Α Helix

The integral membrane protein Apq12 is an important nuclear envelope (NE)/ER modulator that cooperates with the nuclear pore complex (NPC) biogenesis factors Brl1 and Brr6. How Apq12 executes these functions is unknown. Here we identified a short amphipathic α-helix (AαH) in Apq12 that links the two transmembrane domains in the perinuclear space and has liposome-binding properties. Cells expressing an APQ12 (apq12-ah) version in which AαH is disrupted show NPC biogenesis and NE integrity defects, without impacting upon Apq12-ah topology or NE/ER localization. Overexpression of APQ12 but not apq12-ah triggers striking over-proliferation of the outer nuclear membrane (ONM)/ER and promotes accumulation of phosphatidic acid (PA) at the NE. Apq12 and Apq12-ah both associate with NPC biogenesis intermediates and removal of AαH increases both Brl1 levels and the interaction between Brl1 and Brr6. We conclude that the short amphipathic α-helix of Apq12 regulates the function of Brl1 and Brr6 and promotes PA accumulation at the NE during NPC biogenesis.

scholarly journals Nuclear Envelope Regulation of Oncogenic Processes: Roles in Pancreatic Cancer

Epigenomes ◽  
2018 ◽  
Vol 2 (3) ◽  
pp. 15 ◽  
Author(s):  
Claudia Preston ◽  
Randolph Faustino
Keyword(s):  
Pancreatic Cancer ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Structural Integrity ◽  
Radiation Treatment ◽  
Tumor Biology ◽  
Nuclear Lamina ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Linc Complex

Pancreatic cancer is an aggressive and intractable malignancy with high mortality. This is due in part to a high resistance to chemotherapeutics and radiation treatment conferred by diverse regulatory mechanisms. Among these, constituents of the nuclear envelope play a significant role in regulating oncogenesis and pancreatic tumor biology, and this review focuses on three specific components and their roles in cancer. The LINC complex is a nuclear envelope component formed by proteins with SUN and KASH domains that interact in the periplasmic space of the nuclear envelope. These interactions functionally and structurally couple the cytoskeleton to chromatin and facilitates gene regulation informed by cytoplasmic activity. Furthermore, cancer cell invasiveness is impacted by LINC complex biology. The nuclear lamina is adjacent to the inner nuclear membrane of the nuclear envelope and can actively regulate chromatin in addition to providing structural integrity to the nucleus. A disrupted lamina can impart biophysical compromise to nuclear structure and function, as well as form dysfunctional micronuclei that may lead to genomic instability and chromothripsis. In close relationship to the nuclear lamina is the nuclear pore complex, a large megadalton structure that spans both outer and inner membranes of the nuclear envelope. The nuclear pore complex mediates bidirectional nucleocytoplasmic transport and is comprised of specialized proteins called nucleoporins that are overexpressed in many cancers and are diagnostic markers for oncogenesis. Furthermore, recent demonstration of gene regulatory functions for discrete nucleoporins independent of their nuclear trafficking function suggests that these proteins may contribute more to malignant phenotypes beyond serving as biomarkers. The nuclear envelope is thus a complex, intricate regulator of cell signaling, with roles in pancreatic tumorigenesis and general oncogenic transformation.

scholarly journals Nup180, a novel nuclear pore complex protein localizing to the cytoplasmic ring and associated fibrils.

1993 ◽  
Vol 123 (6) ◽  
pp. 1345-1354 ◽  
Author(s):  
N Wilken ◽  
U Kossner ◽  
J L Senécal ◽  
U Scheer ◽  
M C Dabauvalle
Keyword(s):  
Nuclear Pore Complex ◽  
Mammalian Cells ◽  
Protein Transport ◽  
Nucleocytoplasmic Transport ◽  
Transport Processes ◽  
Nuclear Pore ◽  
Nuclear Surface ◽  
Apparent Lack

Using an autoimmune serum from a patient with overlap connective tissue disease we have identified by biochemical and immunocytochemical approaches an evolutionarily conserved nuclear pore complex (NPC) protein with an estimated molecular mass of 180 kD and an isoelectric point of approximately 6.2 which we have designated as nup180. Extraction of isolated nuclear envelopes with 2 M urea and chromatography of the solubilized proteins on WGA-Sepharose demonstrated that nup180 is a peripheral membrane protein and does not react with WGA. Affinity-purified antibodies yielded a punctate immunofluorescent pattern of the nuclear surface of mammalian cells and stained brightly the nuclear envelope of cryosectioned Xenopus oocytes. Nuclei reconstituted in vitro in Xenopus egg extract were also stained in the characteristic punctate fashion. Immunogold EM localized nup180 exclusively to the cytoplasmic ring of NPCs and short fibers emanating therefrom into the cytoplasm. Antibodies to nup180 did not inhibit nuclear protein transport in vivo nor in vitro. Despite the apparent lack of involvement in NPC assembly or nucleocytoplasmic transport processes, the conservation of nup180 across species and its exclusive association with the NPC cytoplasmic ring suggests an important, though currently undefined function for this novel NPC protein.

scholarly journals A Nup133-dependent NPC-anchored network tethers centrosomes to the nuclear envelope in prophase

2011 ◽  
Vol 192 (5) ◽  
pp. 855-871 ◽  
Author(s):  
Stéphanie Bolhy ◽  
Imène Bouhlel ◽  
Elisa Dultz ◽  
Tania Nayak ◽  
Michela Zuccolo ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Mitotic Spindle ◽  
Interaction Network ◽  
Nuclear Pore ◽  
Nuclear Surface ◽  
Cellular Mechanisms ◽  
Mitotic Entry ◽  
Dynactin Complex ◽  
Dependent Pathway

Centrosomes are closely associated with the nuclear envelope (NE) throughout the cell cycle and this association is maintained in prophase when they separate to establish the future mitotic spindle. At this stage, the kinetochore constituents CENP-F, NudE, NudEL, dynein, and dynactin accumulate at the NE. We demonstrate here that the N-terminal domain of the nuclear pore complex (NPC) protein Nup133, although largely dispensable for NPC assembly, is required for efficient anchoring of the dynein/dynactin complex to the NE in prophase. Nup133 exerts this function through an interaction network via CENP-F and NudE/EL. We show that this molecular chain is critical for maintaining centrosome association with the NE at mitotic entry and contributes to this process without interfering with the previously described RanBP2–BICD2-dependent pathway of centrosome anchoring. Finally, our study reveals that tethering of centrosomes to the nuclear surface at the G2/M transition contributes, along with other cellular mechanisms, to early stages of bipolar spindle assembly.

scholarly journals REEP4 is recruited to the inner nuclear membrane by ELYS and promotes nuclear pore complex formation

2020 ◽  
Author(s):  
Banafsheh Golchoubian ◽  
Andreas Brunner ◽  
Helena Bragulat-Teixidor ◽  
Busra A. Akarlar ◽  
Nurhan Ozlu ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Nuclear Membrane ◽  
Nucleocytoplasmic Transport ◽  
Local Deformation ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Membrane Remodeling ◽  
Inner Nuclear Membrane ◽  
Pore Complexes

AbstractNuclear pore complexes (NPCs) are channels within the nuclear envelope that mediate nucleocytoplasmic transport. NPCs assemble either into the closed nuclear envelope during interphase or concomitantly with nuclear envelope reformation during anaphase. Both, interphase and post-mitotic NPC biogenesis require local deformation of membrane. Yet, the factors that control proper membrane remodeling for post-mitotic NPC assembly are unknown. Here, we report that the reticulon homology domain-protein REEP4 localizes not only to high-curvature membrane of the cytoplasmic endoplasmic reticulum (ER) but also to the inner nuclear membrane (INM). We show that REEP4 is recruited to the INM by the NPC biogenesis factor ELYS and promotes NPC assembly. REEP4 contributes mainly to anaphase NPC assembly, suggesting that REEP4 has an unexpected role in coordinating nuclear envelope reformation with post-mitotic NPC biogenesis.

Temperature-induced changes in nuclear pore complex frequencies nuclear envelope surface areas and nuclear volumes in light-synchronized Euglena

1981 ◽  
Vol 59 (10) ◽  
pp. 802-809 ◽  
Author(s):  
John S. Greenwood ◽  
John N. A. Lott
Keyword(s):  
Nuclear Envelope ◽  
Surface Area ◽  
Nuclear Pore Complex ◽  
Temperature Shift ◽  
Mean Value ◽  
Nuclear Pore ◽  
Nuclear Surface ◽  
Envelope Surface ◽  
The Third ◽  
Surface Areas

An autotrophic culture of Euglena, synchronized using a day: night (D:N), 14:10-h cycle, was subjected to a 21.5 → 31.5 °C temperature shift and then to a reversed shift in temperature after three D:N cycles at 31.5 °C. Nuclear pore complex (NPC) number per square micrometre and nuclear surface area and volume determinations were made on G1 cells at various intervals. Cells sampled immediately prior to the 21.5 → 31.5 °C shift had a mean value of 37.68 NPC∙μm−2 nuclear envelope surface area, 30.40 NPCs/μm2 after three D:N cycles at.31.5 °C and 39.98 NPCs/μm2 after three D:N cycles at the resumed culture temperature of 21.5 °C. Thus temperature changes affect NPC numbers per square micrometre and these changes are reversible.Mean nuclear surface area was 125.76 μm2 immediately prior to the 21.5 → 31.5 °C shift, and decreased over two D:N cycles at 31.5 °C to 101.30 μm2 by the end of the third D:N cycle. Nuclear envelope surface area, one and two D:N cycles after the 31.5 → 21.5 °C shift, was approximately equal that prior to the 21.5 → 31.5 °C shift. After the third D:N cycle, however, nuclear surface area had increased to 173.05 μm2. The changes in nuclear surface area resulted in large differences in the estimates of the total number of NPCs per nucleus. Euglena immediately prior to the 21.5 → 31.5 °C temperature shift had 4739 NPCs/nucleus; immediately prior to the 31.5 → 21.5 °C shift had 3079 NPCs/nucleus; and had 6919 NPCs/nucleus at 21.5 °C and three D:N cycles after the 31.5 → 21.5 °C shift. Estimates of the number of NPCs per cubic micrometre of nuclear volume were almost identical between these samples.

scholarly journals Nucleocytoplasmic Transport: Regulatory Mechanisms and the Implications in Neurodegeneration

2021 ◽  
Vol 22 (8) ◽  
pp. 4165
Author(s):  
Baojin Ding ◽  
Masood Sepehrimanesh
Keyword(s):  
Neurodegenerative Diseases ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Molecular Mechanisms ◽  
Nucleocytoplasmic Transport ◽  
Research Field ◽  
Regulatory Mechanisms ◽  
Nuclear Pore ◽  
Age Related ◽  
Lateral Sclerosis

Nucleocytoplasmic transport (NCT) across the nuclear envelope is precisely regulated in eukaryotic cells, and it plays critical roles in maintenance of cellular homeostasis. Accumulating evidence has demonstrated that dysregulations of NCT are implicated in aging and age-related neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), Alzheimer’s disease (AD), and Huntington disease (HD). This is an emerging research field. The molecular mechanisms underlying impaired NCT and the pathogenesis leading to neurodegeneration are not clear. In this review, we comprehensively described the components of NCT machinery, including nuclear envelope (NE), nuclear pore complex (NPC), importins and exportins, RanGTPase and its regulators, and the regulatory mechanisms of nuclear transport of both protein and transcript cargos. Additionally, we discussed the possible molecular mechanisms of impaired NCT underlying aging and neurodegenerative diseases, such as ALS/FTD, HD, and AD.

scholarly journals Nuclear pore complex glycoproteins contain cytoplasmically disposed O-linked N-acetylglucosamine.

1987 ◽  
Vol 104 (5) ◽  
pp. 1157-1164 ◽  
Author(s):  
G D Holt ◽  
C M Snow ◽  
A Senior ◽  
R S Haltiwanger ◽  
L Gerace ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Nuclear Envelope ◽  
Rat Liver ◽  
Nuclear Pore Complex ◽  
Nuclear Pore ◽  
Side Chains ◽  
Nuclear Membranes ◽  
Cell Biol ◽  
Direct Demonstration ◽  
Polypeptide Backbone

A novel form of protein-saccharide linkage consisting of single N-acetylglucosamine (GlcNAc) residues attached in O-linkages directly to the polypeptide backbone has been described (Holt, G. D., and G. W. Hart, 1986, J. Biol. Chem., 261:8049-8057). This modification was found on proteins distributed throughout the cell, although proteins bearing O-linked GlcNAc moieties were particularly abundant in the cytosolic and nuclear envelope fractions of rat liver. In the accompanying article (Snow, C. M., A. Senior, and L. Gerace, 1987, J. Cell. Biol., 104: 1143-1156), the authors describe monoclonal antibodies directed against eight proteins localized to the nuclear pore complex. These proteins occur on the cytoplasmic and nucleoplasmic (but not lumenal) sides of nuclear membranes. In this report, we demonstrate that all members of this group of pore complex proteins bear multiple O-linked GlcNAc residues. Further, we show that the O-linked GlcNAc moieties are linked via serine (and possibly threonine) side chains to these proteins. Perturbing the O-linked GlcNAc residues either by covalently attaching galactose to them or by releasing them with beta-N-acetylglucosaminidase strongly diminishes the immunoreactivity of the proteins with all of the monoclonal antibodies. However, the O-linked GlcNAc moieties are only part of the epitopes recognized, since O-GlcNAc-containing limit pronase fragments of nuclear pore complex proteins cannot be immunoprecipitated by these antibodies. These findings, taken together with those in the accompanying article, are a direct demonstration that proteins of the cytoplasm and nucleoplasm bear O-linked GlcNAc residues.

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