Age-dependent deterioration of nuclear pore assembly in mitotic cells decreases transport dynamics

AbstractNuclear transport is facilitated by the Nuclear Pore Complex (NPC) and is essential for life in eukaryotes. The NPC is a long-lived and exceptionally large structure. We asked whether NPC function is compromised in ageing mitotic cells. By imaging of single yeast cells during ageing, we show that the abundance of several NPC components and NPC assembly factors decreases while signs of misassembled NPCs appear. Consequently, nuclear permeability decreases, resulting in decreased dynamics of transcription factor shuttling and increased nuclear compartmentalisation. In support that declining NPC quality control is important in mitotic ageing, we find that the transport kinetics observed in ageing is mimicked in an NPC assembly mutant. Additionally, the single cell life histories reveal that cells that better maintain NPC function are longer lived. We conclude that assembly and quality control of NPCs are major challenges for ageing mitotic cells.

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Age-dependent deterioration of nuclear pore assembly in mitotic cells decreases transport dynamics

eLife ◽

10.7554/elife.48186 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 11

Author(s):

Irina L Rempel ◽

Matthew M Crane ◽

David J Thaller ◽

Ankur Mishra ◽

Daniel PM Jansen ◽

...

Keyword(s):

Life Histories ◽

Yeast Cells ◽

Nuclear Pore ◽

Functional Changes ◽

Transport Dynamics ◽

Cell Life ◽

Age Dependent ◽

Nuclear Pore Assembly ◽

Nuclear Compartmentalization ◽

Mitotic Cells

Nuclear transport is facilitated by the Nuclear Pore Complex (NPC) and is essential for life in eukaryotes. The NPC is a long-lived and exceptionally large structure. We asked whether NPC quality control is compromised in aging mitotic cells. Our images of single yeast cells during aging, show that the abundance of several NPC components and NPC assembly factors decreases. Additionally, the single-cell life histories reveal that cells that better maintain those components are longer lived. The presence of herniations at the nuclear envelope of aged cells suggests that misassembled NPCs are accumulated in aged cells. Aged cells show decreased dynamics of transcription factor shuttling and increased nuclear compartmentalization. These functional changes are likely caused by the presence of misassembled NPCs, as we find that two NPC assembly mutants show similar transport phenotypes as aged cells. We conclude that NPC interphase assembly is a major challenge for aging mitotic cells.

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Author response: Age-dependent deterioration of nuclear pore assembly in mitotic cells decreases transport dynamics

10.7554/elife.48186.029 ◽

2019 ◽

Cited By ~ 2

Author(s):

Irina L Rempel ◽

Matthew M Crane ◽

David J Thaller ◽

Ankur Mishra ◽

Daniel PM Jansen ◽

...

Keyword(s):

Author Response ◽

Nuclear Pore ◽

Transport Dynamics ◽

Age Dependent ◽

Nuclear Pore Assembly ◽

Mitotic Cells

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Decision letter: Age-dependent deterioration of nuclear pore assembly in mitotic cells decreases transport dynamics

10.7554/elife.48186.028 ◽

2019 ◽

Keyword(s):

Nuclear Pore ◽

Transport Dynamics ◽

Age Dependent ◽

Nuclear Pore Assembly ◽

Mitotic Cells

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Specialization of chromatin-bound nuclear pore complexes promotes yeast aging

10.1101/2021.06.28.450139 ◽

2021 ◽

Author(s):

Anne C Meinema ◽

Theo Aspert ◽

Sung Sik Lee ◽

Gilles Charvin ◽

Yves Barral

Keyword(s):

Quality Control ◽

Nuclear Pore Complex ◽

Mrna Export ◽

Yeast Cells ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Replicative Lifespan ◽

Yeast Aging ◽

Key Drivers ◽

Pore Complexes

The nuclear pore complex (NPC) mediates nearly all exchanges between nucleus and cytoplasm, and changes composition in many species as the organism ages. However, how these changes arise and whether they contribute themselves to aging is poorly understood. We show that in replicatively aging yeast cells attachment of DNA circles to NPCs drives the displacement of the NPCs’ nuclear basket and cytoplasmic complexes. Remodeling of the NPC resulted from the regulation of basket components by SAGA, rather than from damages. These changes affected NPC interaction with mRNA export factors, without affecting the residence of import factors or engaging the NPC quality control machinery. Mutations preventing NPC remodeling extended the replicative lifespan of the cells. Thus, our data indicate that DNA circles accumulating in the mother cell drive aging at least in part by triggering NPC specialization. We suggest that antagonistic pleiotropic effects of NPC specialization are key drivers of aging.

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Faculty Opinions recommendation of Age-dependent deterioration of nuclear pore complexes causes a loss of nuclear integrity in postmitotic cells.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1148112.605236 ◽

2009 ◽

Author(s):

Joachim Seemann

Keyword(s):

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Age Dependent ◽

Postmitotic Cells ◽

Pore Complexes

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Stoichiometry and compositional plasticity of the yeast nuclear pore complex revealed by quantitative fluorescence microscopy

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1719398115 ◽

2018 ◽

Vol 115 (17) ◽

pp. E3969-E3977 ◽

Cited By ~ 36

Author(s):

Sasikumar Rajoo ◽

Pascal Vallotton ◽

Evgeny Onischenko ◽

Karsten Weis

Keyword(s):

Nuclear Pore Complex ◽

Yeast Cells ◽

Nuclear Pore ◽

Altered Expression ◽

Copy Numbers ◽

Quantitative Image ◽

Multiple Copies ◽

Almost All ◽

High Degree

The nuclear pore complex (NPC) is an eightfold symmetrical channel providing selective transport of biomolecules across the nuclear envelope. Each NPC consists of ∼30 different nuclear pore proteins (Nups) all present in multiple copies per NPC. Significant progress has recently been made in the characterization of the vertebrate NPC structure. However, because of the estimated size differences between the vertebrate and yeast NPC, it has been unclear whether the NPC architecture is conserved between species. Here, we have developed a quantitative image analysis pipeline, termed nuclear rim intensity measurement (NuRIM), to precisely determine copy numbers for almost all Nups within native NPCs of budding yeast cells. Our analysis demonstrates that the majority of yeast Nups are present at most in 16 copies per NPC. This reveals a dramatic difference to the stoichiometry determined for the human NPC, suggesting that despite a high degree of individual Nup conservation, the yeast and human NPC architecture is significantly different. Furthermore, using NuRIM, we examined the effects of mutations on NPC stoichiometry. We demonstrate for two paralog pairs of key scaffold Nups, Nup170/Nup157 and Nup192/Nup188, that their altered expression leads to significant changes in the NPC stoichiometry inducing either voids in the NPC structure or substitution of one paralog by the other. Thus, our results not only provide accurate stoichiometry information for the intact yeast NPC but also reveal an intriguing compositional plasticity of the NPC architecture, which may explain how differences in NPC composition could arise in the course of evolution.

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Insertion Defects of Mitochondrially Encoded Proteins Burden the Mitochondrial Quality Control System

Cells ◽

10.3390/cells7100172 ◽

2018 ◽

Vol 7 (10) ◽

pp. 172 ◽

Cited By ~ 2

Author(s):

Braulio Vargas Möller-Hergt ◽

Andreas Carlström ◽

Tamara Suhm ◽

Martin Ott

Keyword(s):

Quality Control ◽

Control System ◽

Mitochondrial Protein ◽

Quality Control System ◽

Yeast Cells ◽

Mitochondrial Quality Control ◽

Protein Insertion ◽

Mitochondrial Proteome ◽

Proteotoxic Stress ◽

Membrane Protein Insertion

The mitochondrial proteome contains proteins from two different genetic systems. Proteins are either synthesized in the cytosol and imported into the different compartments of the organelle or directly produced in the mitochondrial matrix. To ensure proteostasis, proteins are monitored by the mitochondrial quality control system, which will degrade non-native polypeptides. Defective mitochondrial membrane proteins are degraded by membrane-bound AAA-proteases. These proteases are regulated by factors promoting protein turnover or preventing their degradation. Here we determined genetic interactions between the mitoribosome receptors Mrx15 and Mba1 with the quality control system. We show that simultaneous absence of Mrx15 and the regulators of the i-AAA protease Mgr1 and Mgr3 provokes respiratory deficiency. Surprisingly, mutants lacking Mrx15 were more tolerant against proteotoxic stress. Furthermore, yeast cells became hypersensitive against proteotoxic stress upon deletion of MBA1. Contrary to Mrx15, Mba1 cooperates with the regulators of the m-AAA and i-AAA proteases. Taken together, these results suggest that membrane protein insertion and mitochondrial AAA-proteases are functionally coupled, possibly reflecting an early quality control step during mitochondrial protein synthesis.

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The Positioning and Dynamics of Origins of Replication in the Budding Yeast Nucleus

The Journal of Cell Biology ◽

10.1083/jcb.152.2.385 ◽

2001 ◽

Vol 152 (2) ◽

pp. 385-400 ◽

Cited By ~ 115

Author(s):

Patrick Heun ◽

Thierry Laroche ◽

M.K. Raghuraman ◽

Susan M. Gasser

Keyword(s):

Nuclear Envelope ◽

Chromatin Structure ◽

Fluorescent Protein ◽

Time Lapse ◽

Yeast Cells ◽

G1 Phase ◽

Nuclear Pore ◽

Origin Firing ◽

Green Fluorescent

We have analyzed the subnuclear position of early- and late-firing origins of DNA replication in intact yeast cells using fluorescence in situ hybridization and green fluorescent protein (GFP)–tagged chromosomal domains. In both cases, origin position was determined with respect to the nuclear envelope, as identified by nuclear pore staining or a NUP49-GFP fusion protein. We find that in G1 phase nontelomeric late-firing origins are enriched in a zone immediately adjacent to the nuclear envelope, although this localization does not necessarily persist in S phase. In contrast, early firing origins are randomly localized within the nucleus throughout the cell cycle. If a late-firing telomere-proximal origin is excised from its chromosomal context in G1 phase, it remains late-firing but moves rapidly away from the telomere with which it was associated, suggesting that the positioning of yeast chromosomal domains is highly dynamic. This is confirmed by time-lapse microscopy of GFP-tagged origins in vivo. We propose that sequences flanking late-firing origins help target them to the periphery of the G1-phase nucleus, where a modified chromatin structure can be established. The modified chromatin structure, which would in turn retard origin firing, is both autonomous and mobile within the nucleus.

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In Vivo Dynamics of Nuclear Pore Complexes in Yeast

The Journal of Cell Biology ◽

10.1083/jcb.136.6.1185 ◽

1997 ◽

Vol 136 (6) ◽

pp. 1185-1199 ◽

Cited By ~ 82

Author(s):

Mirella Bucci ◽

Susan R. Wente

Keyword(s):

Nuclear Envelope ◽

Recipient Cell ◽

Nucleocytoplasmic Transport ◽

Yeast Cells ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Central Location ◽

Nuclear Membranes ◽

Mutant Strains ◽

Pore Complexes

While much is known about the role of nuclear pore complexes (NPCs) in nucleocytoplasmic transport, the mechanism of NPC assembly into pores formed through the double lipid bilayer of the nuclear envelope is not well defined. To investigate the dynamics of NPCs, we developed a live-cell assay in the yeast Saccharomyces cerevisiae. The nucleoporin Nup49p was fused to the green fluorescent protein (GFP) of Aequorea victoria and expressed in nup49 null haploid yeast cells. When the GFP–Nup49p donor cell was mated with a recipient cell harboring only unlabeled Nup49p, the nuclei fused as a consequence of the normal mating process. By monitoring the distribution of the GFP–Nup49p, we could assess whether NPCs were able to move from the donor section of the nuclear envelope to that of the recipient nucleus. We observed that fluorescent NPCs moved and encircled the entire nucleus within 25 min after fusion. When assays were done in mutant kar1-1 strains, where nuclear fusion does not occur, GFP–Nup49p appearance in the recipient nucleus occurred at a very slow rate, presumably due to new NPC biogenesis or to exchange of GFP– Nup49p into existing recipient NPCs. Interestingly, in a number of existing mutant strains, NPCs are clustered together at permissive growth temperatures. This has been explained with two different hypotheses: by movement of NPCs through the double nuclear membranes with subsequent clustering at a central location; or, alternatively, by assembly of all NPCs at a central location (such as the spindle pole body) with NPCs in mutant cells unable to move away from this point. Using the GFP–Nup49p system with a mutant in the NPCassociated factor Gle2p that exhibits formation of NPC clusters only at 37°C, it was possible to distinguish between these two models for NPC dynamics. GFP– Nup49p-labeled NPCs, assembled at 23°C, moved into clusters when the cells were shifted to growth at 37°C. These results indicate that NPCs can move through the double nuclear membranes and, moreover, can do so to form NPC clusters in mutant strains. Such clusters may result by releasing NPCs from a nuclear tether, or by disappearance of a protein that normally prevents pore aggregation. This system represents a novel approach for identifying regulators of NPC assembly and movement in the future.

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