scholarly journals The NIMA Kinase Is Required To Execute Stage-Specific Mitotic Functions after Initiation of Mitosis

2013 ◽  
Vol 13 (1) ◽  
pp. 99-109 ◽  
Author(s):  
Meera Govindaraghavan ◽  
Alisha A. Lad ◽  
Stephen A. Osmani
Keyword(s):  
Spindle Assembly Checkpoint ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Cycle Phase ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Content Type ◽  
Mitotic Entry ◽  
Spindle Pole Bodies ◽  
Pore Complexes

ABSTRACTThe G2-M transition inAspergillus nidulansrequires the NIMA kinase, the founding member of the Nek kinase family. Inactivation of NIMA results in a late G2arrest, while overexpression of NIMA is sufficient to promote mitotic events independently of cell cycle phase. Endogenously tagged NIMA-GFP has dynamic mitotic localizations appearing first at the spindle pole body and then at nuclear pore complexes before transitioning to within nuclei and the mitotic spindle and back at the spindle pole bodies at mitotic exit, suggesting that it functions sequentially at these locations. Since NIMA is indispensable for mitotic entry, it has been difficult to determine the requirement of NIMA for subaspects of mitosis. We show here that when NIMA is partially inactivated, although mitosis can be initiated, a proportion of cells fail to successfully generate two daughter nuclei. We further define the mitotic defects to show that normal NIMA function is required for the formation of a bipolar spindle, nuclear pore complex disassembly, completion of chromatin segregation, and the normal structural rearrangements of the nuclear envelope required to generate two nuclei from one. In the remaining population of cells that enter mitosis with inadequate NIMA, two daughter nuclei are generated in a manner dependent on the spindle assembly checkpoint, indicating highly penetrant defects in mitotic progression without sufficient NIMA activity. This study shows that NIMA is required not only for mitotic entry but also sequentially for successful completion of stage-specific mitotic events.

scholarly journals The Three Fungal Transmembrane Nuclear Pore Complex Proteins of Aspergillus nidulans Are Dispensable in the Presence of an Intact An-Nup84-120 Complex

2009 ◽  
Vol 20 (2) ◽  
pp. 616-630 ◽  
Author(s):  
Hui-Lin Liu ◽  
Colin P.C. De Souza ◽  
Aysha H. Osmani ◽  
Stephen A. Osmani
Keyword(s):  
High Temperature ◽  
Nuclear Envelope ◽  
Aspergillus Nidulans ◽  
Nuclear Pore Complex ◽  
Mitotic Spindle ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Spindle Pole Bodies ◽  
Pore Complexes

In Aspergillus nidulans nuclear pore complexes (NPCs) undergo partial mitotic disassembly such that 12 NPC proteins (Nups) form a core structure anchored across the nuclear envelope (NE). To investigate how the NPC core is maintained, we affinity purified the major core An-Nup84-120 complex and identified two new fungal Nups, An-Nup37 and An-ELYS, previously thought to be vertebrate specific. During mitosis the An-Nup84-120 complex locates to the NE and spindle pole bodies but, unlike vertebrate cells, does not concentrate at kinetochores. We find that mutants lacking individual An-Nup84-120 components are sensitive to the membrane destabilizer benzyl alcohol (BA) and high temperature. Although such mutants display no defects in mitotic spindle formation, they undergo mitotic specific disassembly of the NPC core and transient aggregation of the mitotic NE, suggesting the An-Nup84-120 complex might function with membrane. Supporting this, we show cells devoid of all known fungal transmembrane Nups (An-Ndc1, An-Pom152, and An-Pom34) are viable but that An-ndc1 deletion combined with deletion of individual An-Nup84-120 components is either lethal or causes sensitivity to treatments expected to destabilize membrane. Therefore, the An-Nup84-120 complex performs roles, perhaps at the NPC membrane as proposed previously, that become essential without the An-Ndc1 transmembrane Nup.

scholarly journals Tts1, the fission yeast homologue of the TMEM33 family, functions in NE remodeling during mitosis

2014 ◽  
Vol 25 (19) ◽  
pp. 2970-2983 ◽  
Author(s):  
Dan Zhang ◽  
Snezhana Oliferenko
Keyword(s):  
Fission Yeast ◽  
Structural Features ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
C Terminus ◽  
Spindle Pole Bodies ◽  
Evolutionarily Conserved ◽  
Pore Complexes ◽  
Insight Into

The fission yeast Schizosaccharomyces pombe undergoes “closed” mitosis in which the nuclear envelope (NE) stays intact throughout chromosome segregation. Here we show that Tts1, the fission yeast TMEM33 protein that was previously implicated in organizing the peripheral endoplasmic reticulum (ER), also functions in remodeling the NE during mitosis. Tts1 promotes insertion of spindle pole bodies (SPBs) in the NE at the onset of mitosis and modulates distribution of the nuclear pore complexes (NPCs) during mitotic NE expansion. Structural features that drive partitioning of Tts1 to the high-curvature ER domains are crucial for both aspects of its function. An amphipathic helix located at the C-terminus of Tts1 is important for ER shaping and modulating the mitotic NPC distribution. Of interest, the evolutionarily conserved residues at the luminal interface of the third transmembrane region function specifically in promoting SPB-NE insertion. Our data illuminate cellular requirements for remodeling the NE during “closed” nuclear division and provide insight into the structure and functions of the eukaryotic TMEM33 family.

scholarly journals The SUN protein Mps3 controls Ndc1 distribution and function on the nuclear membrane

2014 ◽  
Vol 204 (4) ◽  
pp. 523-539 ◽  
Author(s):  
Jingjing Chen ◽  
Christine J. Smoyer ◽  
Brian D. Slaughter ◽  
Jay R. Unruh ◽  
Sue L. Jaspersen
Keyword(s):  
Spindle Pole Body ◽  
Integral Membrane Protein ◽  
Spindle Pole ◽  
Correlation Spectroscopy ◽  
Yeast Cells ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
The Sun ◽  
Pore Complexes ◽  
And Function

In closed mitotic systems such as Saccharomyces cerevisiae, nuclear pore complexes (NPCs) and the spindle pole body (SPB) must assemble into an intact nuclear envelope (NE). Ndc1 is a highly conserved integral membrane protein involved in insertion of both complexes. In this study, we show that Ndc1 interacts with the SUN domain–containing protein Mps3 on the NE in live yeast cells using fluorescence cross-correlation spectroscopy. Genetic and molecular analysis of a series of new ndc1 alleles allowed us to understand the role of Ndc1–Mps3 binding at the NE. We show that the ndc1-L562S allele is unable to associate specifically with Mps3 and find that this mutant is lethal due to a defect in SPB duplication. Unlike other ndc1 alleles, the growth and Mps3 binding defect of ndc1-L562S is fully suppressed by deletion of POM152, which encodes a NPC component. Based on our data we propose that the Ndc1–Mps3 interaction is important for controlling the distribution of Ndc1 between the NPC and SPB.

scholarly journals Characterization of spindle pole body duplication reveals a regulatory role for nuclear pore complexes

2017 ◽  
Vol 216 (8) ◽  
pp. 2425-2442 ◽  
Author(s):  
Diana Rüthnick ◽  
Annett Neuner ◽  
Franziska Dietrich ◽  
Daniel Kirrmaier ◽  
Ulrike Engel ◽  
...  
Keyword(s):  
Spindle Pole Body ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Pole Body ◽  
Associated Proteins ◽  
Pore Complexes ◽  
Open Question ◽  
Cytoplasmic Side

The spindle pole body (SPB) of budding yeast duplicates once per cell cycle. In G1, the satellite, an SPB precursor, assembles next to the mother SPB (mSPB) on the cytoplasmic side of the nuclear envelope (NE). How the growing satellite subsequently inserts into the NE is an open question. To address this, we have uncoupled satellite growth from NE insertion. We show that the bridge structure that separates the mSPB from the satellite is a distance holder that prevents deleterious fusion of both structures. Binding of the γ-tubulin receptor Spc110 to the central plaque from within the nucleus is important for NE insertion of the new SPB. Moreover, we provide evidence that a nuclear pore complex associates with the duplicating SPB and helps to insert the SPB into the NE. After SPB insertion, membrane-associated proteins including the conserved Ndc1 encircle the SPB and retain it within the NE. Thus, uncoupling SPB growth from NE insertion unmasks functions of the duplication machinery.

scholarly journals Quantitative analysis of nuclear pore complex organization in Schizosaccharomyces pombe

2021 ◽  
Author(s):  
Joseph M Varberg ◽  
Jay Unruh ◽  
Andrew J Bestul ◽  
Azqa A Khan ◽  
Sue Jaspersen
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Spindle Pole Body ◽  
Cell Types ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Wide Range ◽  
Pore Complexes

The number, distribution and composition of nuclear pore complexes (NPCs) in the nuclear envelope (NE) varies between cell types and changes during cellular differentiation and in disease. To understand how NPC density and organization is controlled, we analyzed NPC number and distribution in the fission yeast Schizosaccharomyces pombe using structured illumination microscopy. The small size of yeast nuclei, genetic features of fungi and our robust image analysis pipeline allowed us to study NPCs in intact nuclei under multiple conditions. Our data revealed that NPC density is maintained across a wide range of nuclear sizes. Regions of reduced NPC density are observed over the nucleolus and surrounding the spindle pole body (SPB). Lem2-mediated tethering of the centromeres to the SPB is required to maintain NPC exclusion, which is important for timely mitotic progression. These findings provide a quantitative understanding of NPC number and distribution in S. pombe and show that interactions between the centromere and the NE influences local NPC distribution.

scholarly journals The nuclear pore complex–associated protein, Mlp2p, binds to the yeast spindle pole body and promotes its efficient assembly

2005 ◽  
Vol 170 (2) ◽  
pp. 225-235 ◽  
Author(s):  
Mario Niepel ◽  
Caterina Strambio-de-Castillia ◽  
Joseph Fasolo ◽  
Brian T. Chait ◽  
Michael P. Rout
Keyword(s):  
Nuclear Pore Complex ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Pole Body ◽  
Core Components ◽  
Synthetically Lethal ◽  
Pore Complexes ◽  
Vertebrate Protein

The two yeast proteins Mlp1p and Mlp2p (homologues of the vertebrate protein Tpr) are filamentous proteins attached to the nuclear face of nuclear pore complexes. Here we perform a proteomic analysis, which reveals that the two Mlps have strikingly different interacting partners, testifying to their different roles within the cell. We find that Mlp2p binds directly to Spc110p, Spc42p, and Spc29p, which are three core components of the spindle pole body (SPB), the nuclear envelope–associated yeast spindle organizer. We further show that SPB function is compromised in mlp2 mutants. Cells lacking Mlp2p form significantly smaller SPBs, accumulate aberrant SPB component-containing structures inside the nucleus, and have stochastic failures of cell division. In addition, depletion of Mlp2p is synthetically lethal with mutants impaired in SPB assembly. Based on these data, we propose that Mlp2p links the SPB to the peripheral Mlp assembly, and that this linkage is required for efficient incorporation of components into the SPB.

scholarly journals Saccharomyces cerevisiae Ndc1p Is a Shared Component of Nuclear Pore Complexes and Spindle Pole Bodies

1998 ◽  
Vol 143 (7) ◽  
pp. 1789-1800 ◽  
Author(s):  
Heidi J. Chial ◽  
Michael P. Rout ◽  
Thomas H. Giddings ◽  
Mark Winey
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Immunofluorescence Microscopy ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Indirect Immunofluorescence Microscopy ◽  
Spindle Pole Bodies ◽  
Shared Component ◽  
Pore Complexes ◽  
Mutant Cells

We report a novel connection between nuclear pore complexes (NPCs) and spindle pole bodies (SPBs) revealed by our studies of the Saccharomyces cerevisiae NDC1 gene. Although both NPCs and SPBs are embedded in the nuclear envelope (NE) in yeast, their known functions are quite distinct. Previous work demonstrated that NDC1 function is required for proper SPB duplication (Winey, M., M.A. Hoyt, C. Chan, L. Goetsch, D. Botstein, and B. Byers. 1993. J. Cell Biol. 122:743–751). Here, we show that Ndc1p is a membrane protein of the NE that localizes to both NPCs and SPBs. Indirect immunofluorescence microscopy shows that Ndc1p displays punctate, nuclear peripheral localization that colocalizes with a known NPC component, Nup49p. Additionally, distinct spots of Ndc1p localization colocalize with a known SPB component, Spc42p. Immunoelectron microscopy shows that Ndc1p localizes to the regions of NPCs and SPBs that interact with the NE. The NPCs in ndc1-1 mutant cells appear to function normally at the nonpermissive temperature. Finally, we have found that a deletion of POM152, which encodes an abundant but nonessential nucleoporin, suppresses the SPB duplication defect associated with a mutation in the NDC1 gene. We show that Ndc1p is a shared component of NPCs and SPBs and propose a shared function in the assembly of these organelles into the NE.

scholarly journals Nuclear Fusion and Genome Encounter during Yeast Zygote Formation

2009 ◽  
Vol 20 (12) ◽  
pp. 2932-2942 ◽  
Author(s):  
Alan Michael Tartakoff ◽  
Purnima Jaiswal
Keyword(s):  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Zygote Formation ◽  
Spindle Pole Bodies ◽  
Higher Eukaryotes ◽  
Underlying Mechanisms ◽  
Pore Complexes ◽  
The Impact ◽  
Chromosome Tethering

When haploid cells of Saccharomyces cerevisiae are crossed, parental nuclei congress and fuse with each other. To investigate underlying mechanisms, we have developed assays that evaluate the impact of drugs and mutations. Nuclear congression is inhibited by drugs that perturb the actin and tubulin cytoskeletons. Nuclear envelope (NE) fusion consists of at least five steps in which preliminary modifications are followed by controlled flux of first outer and then inner membrane proteins, all before visible dilation of the waist of the nucleus or coalescence of the parental spindle pole bodies. Flux of nuclear pore complexes occurs after dilation. Karyogamy requires both the Sec18p/NSF ATPase and ER/NE luminal homeostasis. After fusion, chromosome tethering keeps tagged parental genomes separate from each other. The process of NE fusion and evidence of genome independence in yeast provide a prototype for understanding related events in higher eukaryotes.

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