scholarly journals Membrane binding by CHMP7 coordinates ESCRT-III dependent nuclear envelope reformation

2016 ◽  
Author(s):  
Yolanda Olmos ◽  
Anna Perdrix ◽  
Jeremy G Carlton
Keyword(s):  
Nuclear Envelope ◽  
Membrane Binding ◽  
Point Mutations ◽  
Binding Activity ◽  
Mitotic Exit ◽  
Exit Point ◽  
Cellular Functions ◽  
Endosomal Sorting ◽  
N Terminus ◽  
And Function

AbstractAmongst other cellular functions, the Endosomal Sorting Complex Required for Transport-III (ESCRT-III) machinery controls nuclear envelope (NE) reformation during mitotic exit by sealing holes in the reforming NE. ESCRT-III also acts to repair this organelle upon migration-induced rupture. The ESCRT-III component CHMP7 is responsible for recruitment of ESCRT-III to the NE. Here, we show that the N-terminus of CHMP7, comprising tandem Winged Helix (WH)-domains, is a membrane-binding module. This activity allows CHMP7 to bind to the Endoplasmic Reticulum (ER), an organelle continuous with the NE, and provides a platform to direct NE-recruitment of ESCRT-III during mitotic exit. Point mutations that disrupt membrane-binding prevent CHMP7 localising to the ER and its subsequent enrichment at the reforming NE. These mutations prevent both assembly of downstream ESCRT-III components at the reforming NE and proper establishment of post-mitotic nucleo-cytoplasmic compartmentalisation. These data identify a novel membrane-binding activity within an ESCRT-III subunit that is essential for post-mitotic nuclear regeneration.One Sentence SummaryCHMP7’s atypical N-terminus is a membrane-binding module that allows assembly and function of ESCRT-III at the nuclear envelope during mitotic exit.

scholarly journals CDK1 controls CHMP7-dependent nuclear envelope reformation

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Alberto T Gatta ◽  
Yolanda Olmos ◽  
Caroline L Stoten ◽  
Qu Chen ◽  
Peter B Rosenthal ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Cell Cycle Control ◽  
Nucleocytoplasmic Transport ◽  
Control Programme ◽  
Mitotic Exit ◽  
Microtubule Network ◽  
Endosomal Sorting ◽  
Protein Biochemistry ◽  
M Phase ◽  
Membrane Sealing

Through membrane sealing and disassembly of spindle microtubules, the Endosomal Sorting Complex Required for Transport-III (ESCRT-III) machinery has emerged as a key player in the regeneration of a sealed nuclear envelope (NE) during mitotic exit, and in the repair of this organelle during interphase rupture. ESCRT-III assembly at the NE occurs transiently during mitotic exit and is initiated when CHMP7, an ER-localised ESCRT-II/ESCRT-III hybrid protein, interacts with the Inner Nuclear Membrane (INM) protein LEM2. Whilst classical nucleocytoplasmic transport mechanisms have been proposed to separate LEM2 and CHMP7 during interphase, it is unclear how CHMP7 assembly is suppressed in mitosis when NE and ER identities are mixed. Here, we use live cell imaging and protein biochemistry to examine the biology of these proteins during mitotic exit. Firstly, we show that CHMP7 plays an important role in the dissolution of LEM2 clusters that form at the NE during M-exit. Secondly, we show that CDK1 phosphorylates CHMP7 upon mitotic entry at Ser3 and Ser441 and that this phosphorylation reduces CHMP7's interaction with LEM2, limiting its assembly during M-phase. We show that spatiotemporal differences in the dephosphorylation of CHMP7 license its assembly at the NE during telophase, but restrict its assembly on the ER at this time. Without CDK1 phosphorylation, CHMP7 undergoes inappropriate assembly in the peripheral ER during M-exit, capturing LEM2 and downstream ESCRT-III components. Lastly, we establish that a microtubule network is dispensable for ESCRT-III assembly at the reforming nuclear envelope. These data identify a key cell-cycle control programme allowing ESCRT-III-dependent nuclear regeneration.

scholarly journals EFFECTORS OF THE SPINDLE ASSEMBLY CHECKPOINT BUT NOT THE MITOTIC EXIT NETWORK ARE CONFINED WITHIN THE NUCLEUS OF SACCHAROMYCES CEREVISIAE

2018 ◽  
Author(s):  
Lydia R Heasley ◽  
Jennifer G DeLuca ◽  
Steven M Markus
Keyword(s):  
Nuclear Envelope ◽  
Mammalian Cells ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Mitotic Checkpoint ◽  
Mitotic Exit ◽  
Cell Cycle Checkpoints ◽  
Anaphase Promoting Complex ◽  
Mitotic Exit Network ◽  
And Function

The Spindle Assembly Checkpoint (SAC) prevents erroneous chromosome segregation by delaying mitotic progression when chromosomes are incorrectly attached to the mitotic spindle. This delay is mediated by Mitotic Checkpoint Complexes (MCCs), which assemble at unattached kinetochores and repress the activity of the Anaphase Promoting Complex/Cyclosome (APC/C). The cellular localizations of MCCs are likely critical for proper SAC function, yet remain poorly defined. We recently demonstrated that in mammalian cells, in which the nuclear envelope disassembles during mitosis, MCCs diffuse throughout the spindle region and cytoplasm. Here, we employed binucleate yeast zygotes to examine the localization dynamics of SAC effectors required for MCC assembly and function in budding yeast, in which the nuclear envelope remains intact throughout mitosis. Our findings indicate that in yeast MCCs are confined to the nuclear compartment and excluded from the cytoplasm during mitosis. In contrast, we find that effectors of the Mitotic Exit Network (MEN) - a pathway that initiates disassembly of the anaphase spindle only when it is properly oriented - are in fact freely exchanged between multiple nuclei within a shared cytoplasm. Our study provides insight into how cell cycle checkpoints have evolved to function in diverse cellular contexts.

scholarly journals Human prion proteins with pathogenic mutations share common conformational changes resulting in enhanced binding to glycosaminoglycans

2007 ◽  
Vol 104 (18) ◽  
pp. 7546-7551 ◽  
Author(s):  
Shaoman Yin ◽  
Nancy Pham ◽  
Shuiliang Yu ◽  
Chaoyang Li ◽  
Poki Wong ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Prion Proteins ◽  
Prion Diseases ◽  
Point Mutations ◽  
Binding Activity ◽  
Binding Motif ◽  
Wild Type ◽  
Pathogenic Mutations ◽  
N Terminus ◽  
Ligand Binding Activity

Mutation in the prion gene PRNP accounts for 10–15% of human prion diseases. However, little is known about the mechanisms by which mutant prion proteins (PrPs) cause disease. Here we investigated the effects of 10 different pathogenic mutations on the conformation and ligand-binding activity of recombinant human PrP (rPrP). We found that mutant rPrPs react more strongly with N terminus-specific antibodies, indicative of a more exposed N terminus. The N terminus of PrP contains a glycosaminoglycan (GAG)-binding motif. Binding of GAG is important in prion disease. Accordingly, all mutant rPrPs bind more GAG, and GAG promotes the aggregation of mutant rPrPs more efficiently than wild-type recombinant normal cellular PrP (rPrPC). Furthermore, point mutations in PRNP also cause conformational changes in the region between residues 109 and 136, resulting in the exposure of a second, normally buried, GAG-binding motif. Importantly, brain-derived PrP from transgenic mice, which express a pathogenic mutant with nine extra octapeptide repeats, also binds more strongly to GAG than wild-type PrPC. Thus, several rPrPs with distinct pathogenic mutations have common conformational changes, which enhance binding to GAG. These changes may contribute to the pathogenesis of inherited prion diseases.

scholarly journals ESCRT-III-driven piecemeal micro-ER-phagy remodels the ER during recovery from ER stress

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Marisa Loi ◽  
Andrea Raimondi ◽  
Diego Morone ◽  
Maurizio Molinari
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Molecular Chaperones ◽  
Binding Activity ◽  
Catabolic Pathway ◽  
Unfolded Protein Responses ◽  
Genetic Analyses ◽  
Endosomal Sorting ◽  
Unfolded Protein ◽  
And Function

Abstract The endoplasmic reticulum (ER) produces about 40% of the nucleated cell’s proteome. ER size and content in molecular chaperones increase upon physiologic and pathologic stresses on activation of unfolded protein responses (UPR). On stress resolution, the mammalian ER is remodeled to pre-stress, physiologic size and function on activation of the LC3-binding activity of the translocon component SEC62. This elicits recov-ER-phagy, i.e., the delivery of the excess ER generated during the phase of stress to endolysosomes (EL) for clearance. Here, ultrastructural and genetic analyses reveal that recov-ER-phagy entails the LC3 lipidation machinery and proceeds via piecemeal micro-ER-phagy, where RAB7/LAMP1-positive EL directly engulf excess ER in processes that rely on the Endosomal Sorting Complex Required for Transport (ESCRT)-III component CHMP4B and the accessory AAA+ ATPase VPS4A. Thus, ESCRT-III-driven micro-ER-phagy emerges as a key catabolic pathway activated to remodel the mammalian ER on recovery from ER stress.

scholarly journals Cofilin Signaling in the CNS Physiology and Neurodegeneration

2021 ◽  
Vol 22 (19) ◽  
pp. 10727
Author(s):  
Jannatun Nayem Namme ◽  
Asim Kumar Bepari ◽  
Hirohide Takebayashi
Keyword(s):  
Protein Trafficking ◽  
Binding Activity ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Oxygen Species ◽  
Cellular Functions ◽  
Binding Partners ◽  
Cellular Factors ◽  
The Central Nervous System ◽  
And Function

All eukaryotic cells are composed of the cytoskeleton, which plays crucial roles in coordinating diverse cellular functions such as cell division, morphology, migration, macromolecular stabilization, and protein trafficking. The cytoskeleton consists of microtubules, intermediate filaments, and actin filaments. Cofilin, an actin-depolymerizing protein, is indispensable for regulating actin dynamics in the central nervous system (CNS) development and function. Cofilin activities are spatiotemporally orchestrated by numerous extra- and intra-cellular factors. Phosphorylation at Ser-3 by kinases attenuate cofilin’s actin-binding activity. In contrast, dephosphorylation at Ser-3 enhances cofilin-induced actin depolymerization. Cofilin functions are also modulated by various binding partners or reactive oxygen species. Although the mechanism of cofilin-mediated actin dynamics has been known for decades, recent research works are unveiling the profound impacts of cofilin dysregulation in neurodegenerative pathophysiology. For instance, oxidative stress-induced increase in cofilin dephosphorylation is linked to the accumulation of tau tangles and amyloid-beta plaques in Alzheimer’s disease. In Parkinson’s disease, cofilin activation by silencing its upstream kinases increases α-synuclein-fibril entry into the cell. This review describes the molecular mechanism of cofilin-mediated actin dynamics and provides an overview of cofilin’s importance in CNS physiology and pathophysiology.

scholarly journals CDK1 controls CHMP7-dependent nuclear envelope reformation

2020 ◽  
Author(s):  
Alberto T Gatta ◽  
Yolanda Olmos ◽  
Caroline L Stoten ◽  
Jeremy G Carlton
Keyword(s):  
Nuclear Envelope ◽  
Cell Cycle Control ◽  
Nucleocytoplasmic Transport ◽  
Control Programme ◽  
Mitotic Exit ◽  
Transport Mechanisms ◽  
Microtubule Network ◽  
Endosomal Sorting ◽  
Protein Biochemistry ◽  
Spindle Microtubules

AbstractThrough the process of annular fusion and disassembly of spindle microtubules, the Endosomal Sorting Complex Required for Transport-III (ESCRT-III) machinery has emerged as a key player in the regeneration of a sealed nuclear envelope during mitotic exit, and in the repair of this organelle during interphase rupture. ESCRT-III polymerisation at the nuclear envelope occurs transiently during mitotic exit and CHMP7, an ER-localised ESCRT-II/ESCRT-III hybrid protein, initiates assembly in a manner dependent upon the INM protein LEM2. Whilst classical nucleocytoplasmic transport mechanisms have been proposed to separate LEM2 and CHMP7 during interphase, it is unclear how CHMP7 assembly is suppressed in mitosis when NE and ER identities are mixed. Here, we use live cell imaging and protein biochemistry to examine the biology of these proteins during mitotic exit. We show that CHMP7 plays an important role in disaggregating LEM2 clusters at the reforming nuclear envelope during mitotic exit to allow proper nuclear regeneration. Further, we show that CDK1 phosphorylates CHMP7 upon mitotic entry and suppresses its polymerisation, preventing its inappropriate capture of LEM2 in the peripheral ER during mitotic exit. Lastly, we establish that a microtubule network is dispensable for ESCRT-III assembly at the reforming nuclear envelope. These data identify a key cell-cycle control programme allowing ESCRT-III-dependent nuclear regeneration.

scholarly journals Structural and Functional Dissection of the Abp1 ADFH Actin-binding Domain Reveals Versatile In Vivo Adapter Functions

2005 ◽  
Vol 16 (7) ◽  
pp. 3128-3139 ◽  
Author(s):  
Omar Quintero-Monzon ◽  
Avital A. Rodal ◽  
Boris Strokopytov ◽  
Steven C. Almo ◽  
Bruce L. Goode
Keyword(s):  
Crystal Structure ◽  
Point Mutations ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Cellular Functions ◽  
Multidomain Protein ◽  
Actin Binding Domain ◽  
And Function ◽  
Actin Binding Site

Abp1 is a multidomain protein that regulates the Arp2/3 complex and links proteins involved in endocytosis to the actin cytoskeleton. All of the proposed cellular functions of Abp1 involve actin filament binding, yet the actin binding site(s) on Abp1 have not been identified, nor has the importance of actin binding for Abp1 localization and function in vivo been tested. Here, we report the crystal structure of the Saccharomyces cerevisiae Abp1 actin-binding actin depolymerizing factor homology (ADFH) domain and dissect its activities by mutagenesis. Abp1-ADFH domain and ADF/cofilin structures are similar, and they use conserved surfaces to bind actin; however, there are also key differences that help explain their differential effects on actin dynamics. Using point mutations, we demonstrate that actin binding is required for localization of Abp1 in vivo, the lethality caused by Abp1 overexpression, and the ability of Abp1 to activate Arp2/3 complex. Furthermore, we genetically uncouple ABP1 functions that overlap with SAC6, SLA1, and SLA2, showing they require distinct combinations of activities and interactions. Together, our data provide the first structural and functional view of the Abp1–actin interaction and show that Abp1 has distinct cellular roles as an adapter, linking different sets of ligands for each function.

Establishing a model to demonstrate physical and mathematical properties of chromatin fibres in fission yeast cells - Research in the Molecular Biophysics Lab at Hiroshima University

Impact ◽  
2018 ◽  
Vol 2018 (3) ◽  
pp. 89-91
Author(s):  
Shin-ichi Tate
Keyword(s):  
Molecular Biology ◽  
Yeast Cells ◽  
Molecular Architecture ◽  
Ministry Of Education ◽  
Cellular Functions ◽  
Sports Science ◽  
Cellular Events ◽  
And Function ◽  
Education Culture ◽  
Open The Doors

The field of molecular biology has provided great insights into the structure and function of key molecules. Thanks to this area of research, we can now grasp the biological details of DNA and have characterised an enormous number of molecules in massive data bases. These 'biological periodic tables' have allowed scientists to connect molecules to particular cellular events, furthering scientific understanding of biological processes. However, molecular biology has yet to answer questions regarding 'higher-order' molecular architecture, such as that of chromatin. Chromatin is the molecular material that serves as the building block for chromosomes, the structures that carry an organism's genetic information inside of the cell's nucleus. Understanding the physical properties of chromatin is crucial in developing a more thorough picture of how chromatin's structure relate to its key cellular functions. Moreover, by establishing a physical model of chromatin, scientists will be able to open the doors into the true inner workings of the cell nucleus. Professor Shin-ichi Tate and his team of researchers at Hiroshima University's Research Center for the Mathematics on Chromatin Live Dynamics (RcMcD), are attempting to do just that. Through a five-year grant funded by the Platform for Dynamic Approaches to Living Systems from the Ministry of Education, Culture, Sports, Science and Technology, Tate is aiming to gain a clearer understanding of the structure and dynamics of chromatin.

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