scholarly journals Vps4 triggers sequential subunit exchange in ESCRT-III polymers that drives membrane constriction and fission

2019 ◽  
Author(s):  
Anna-Katharina Pfitzner ◽  
Vincent Mercier ◽  
Aurélien Roux
Keyword(s):  
Membrane Remodeling ◽  
Unknown Mechanism ◽  
Subunit Exchange ◽  
Membrane Deformation ◽  
Membrane Fission ◽  
Membrane Protrusions

AbstractESCRT-III is a ubiquitous complex which catalyzes membrane fission from within membrane necks via an as yet unknown mechanism. Here, we reconstituted in vitro the ESCRT-III complex onto membranes. We show that based on variable affinities between ESCRT-III proteins and the ATPase Vps4, subunits are recruited to the membrane in a Vps4-driven sequence that starts with Snf7 and ends with Did2 and Ist1 which, together, form a fission-active subcomplex. Sequential recruitment of ESCRT-III subunits is coupled to membrane remodeling. Binding of Did2 promoted the formation of membrane protrusions which later constricted and underwent fission mediated by the recruitment of Ist1. Overall, our results provide a mechanism to explain how a sequence of ESCRT-III subunits drives membrane deformation and fission.

scholarly journals Membrane Remodeling by a Bacterial Phospholipid-Methylating Enzyme

mBio ◽  
2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Linna Danne ◽  
Meriyem Aktas ◽  
Andreas Unger ◽  
Wolfgang A. Linke ◽  
Ralf Erdmann ◽  
...  
Keyword(s):  
Biological Membranes ◽  
Bacterial Membrane ◽  
Membrane Remodeling ◽  
Membrane Deformation ◽  
Bacterial Membranes ◽  
Bacterial Proteins ◽  
Complex Morphology ◽  
Membrane Biosynthesis

ABSTRACT Membrane deformation by proteins is a universal phenomenon that has been studied extensively in eukaryotes but much less in prokaryotes. In this study, we discovered a membrane-deforming activity of the phospholipid N-methyltransferase PmtA from the plant-pathogenic bacterium Agrobacterium tumefaciens. PmtA catalyzes the successive three-step N-methylation of phosphatidylethanolamine to phosphatidylcholine. Here, we defined the lipid and protein requirements for the membrane-remodeling activity of PmtA by a combination of transmission electron microscopy and liposome interaction studies. Dependent on the lipid composition, PmtA changes the shape of spherical liposomes either into filaments or small vesicles. Upon overproduction of PmtA in A. tumefaciens, vesicle-like structures occur in the cytoplasm, dependent on the presence of the anionic lipid cardiolipin. The N-terminal lipid-binding α-helix (αA) is involved in membrane deformation by PmtA. Two functionally distinct and spatially separated regions in αA can be distinguished. Anionic interactions by positively charged amino acids on one face of the helix are responsible for membrane recruitment of the enzyme. The opposite hydrophobic face of the helix is required for membrane remodeling, presumably by shallow insertion into the lipid bilayer. IMPORTANCE The ability to alter the morphology of biological membranes is known for a small number of some bacterial proteins. Our study adds the phospholipid N-methyltransferase PmtA as a new member to the category of bacterial membrane-remodeling proteins. A combination of in vivo and in vitro methods reveals the molecular requirements for membrane deformation at the protein and phospholipid level. The dual functionality of PmtA suggests a contribution of membrane biosynthesis enzymes to the complex morphology of bacterial membranes. IMPORTANCE The ability to alter the morphology of biological membranes is known for a small number of some bacterial proteins. Our study adds the phospholipid N-methyltransferase PmtA as a new member to the category of bacterial membrane-remodeling proteins. A combination of in vivo and in vitro methods reveals the molecular requirements for membrane deformation at the protein and phospholipid level. The dual functionality of PmtA suggests a contribution of membrane biosynthesis enzymes to the complex morphology of bacterial membranes.

scholarly journals GABARAP Like-1 enrichment on membranes: Direct observation of trans-homo-oligomerization between membranes and curvature-dependent partitioning into membrane tubules

2018 ◽  
Author(s):  
Isabelle Motta ◽  
Nathan Nguyen ◽  
Helene Gardavot ◽  
Diana Richerson ◽  
Frederic Pincet ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Membrane Remodeling ◽  
Membrane Deformation ◽  
Curvature Sensing ◽  
Local Areas ◽  
Test Elements ◽  
Atg8 Protein ◽  
Planar Regions ◽  
Curved Membranes

AbstractThe Atg8/LC3/GABARAP protein family has been implicated in membrane remodeling events on the growing autophagosome. In particular, each of these proteins can form a protein-lipid conjugate that has been shown in vitro to drive liposome aggregation and in some cases membrane fusion. Furthermore, yeast Atg8 has been described as a curvature sensing protein, through its natural capacity to concentrate on highly curved membranes. A key advance with yeast Atg8, was the introduction of Giant Unilamellar Vesicles (GUVs) as an in vitro support that could allow membrane deformation and tethering to be observed by simple microscopy. Further, micromanipulation of an individual GUV could be used to create local areas of curvature to follow Atg8 partitioning. Here, we use a recently developed method to decorate GUVs with the mammalian Atg8 protein GABARAPL1 and establish the generality of the observations made on yeast Atg8. Then we apply double micromanipulation, the capture and positioning of two independently prepared GUVs, to test elements of the mechanism, speed and reversibility of mammalian Atg8 protein-mediated tethering. We find that the membranes adhere through GABARAPL1/GABARAPL1 homotypic trans-interactions. On a single membrane with two regions with significantly different curvatures we observed that the regions of higher curvature can be enriched up to 10 times in GABARAPL1 compared to the planar regions. This approach has the potential to allow the formation and study of specific topographically-controlled interfaces involving Atg8-proteins and their targets on apposing membranes.

scholarly journals Association of hemoglobin Saint Etienne (alpha2beta295F8 His replaced by G1n) with hemoglobins A and F. Synthesis and subunit exchange in vitro.

1976 ◽  
Vol 251 (14) ◽  
pp. 4346-4354
Author(s):  
J F Godeau ◽  
Y G Beuzard ◽  
J Cacheleux ◽  
C P Brizard ◽  
A Gibaud ◽  
...  
Keyword(s):  
Subunit Exchange

scholarly journals The role of VPS4 in ESCRT-III polymer remodeling

2019 ◽  
Vol 47 (1) ◽  
pp. 441-448 ◽  
Author(s):  
Christophe Caillat ◽  
Sourav Maity ◽  
Nolwenn Miguet ◽  
Wouter H. Roos ◽  
Winfried Weissenhorn
Keyword(s):  
Active Role ◽  
Mechanistic Models ◽  
Membrane Remodeling ◽  
Filament Formation ◽  
Enveloped Viruses ◽  
Endosomal Sorting ◽  
Membrane Fission ◽  
Dynamic Assembly ◽  
Inside Out

Abstract The endosomal sorting complex required for transport-III (ESCRT-III) and VPS4 catalyze a variety of membrane-remodeling processes in eukaryotes and archaea. Common to these processes is the dynamic recruitment of ESCRT-III proteins from the cytosol to the inner face of a membrane neck structure, their activation and filament formation inside or at the membrane neck and the subsequent or concomitant recruitment of the AAA-type ATPase VPS4. The dynamic assembly of ESCRT-III filaments and VPS4 on cellular membranes induces constriction of membrane necks with large diameters such as the cytokinetic midbody and necks with small diameters such as those of intraluminal vesicles or enveloped viruses. The two processes seem to use different sets of ESCRT-III filaments. Constriction is then thought to set the stage for membrane fission. Here, we review recent progress in understanding the structural transitions of ESCRT-III proteins required for filament formation, the functional role of VPS4 in dynamic ESCRT-III assembly and its active role in filament constriction. The recent data will be discussed in the context of different mechanistic models for inside-out membrane fission.

scholarly journals FAM92A1 is a BAR domain protein required for mitochondrial ultrastructure and function

2018 ◽  
Vol 218 (1) ◽  
pp. 97-111 ◽  
Author(s):  
Liang Wang ◽  
Ziyi Yan ◽  
Helena Vihinen ◽  
Ove Eriksson ◽  
Weihuan Wang ◽  
...  
Keyword(s):  
Mitochondrial Function ◽  
Molecular Mechanisms ◽  
Membrane Curvature ◽  
Mitochondrial Morphology ◽  
Membrane Remodeling ◽  
Mitochondrial Ultrastructure ◽  
Bar Domain ◽  
Bar Domain Proteins ◽  
Domain Protein

Mitochondrial function is closely linked to its dynamic membrane ultrastructure. The mitochondrial inner membrane (MIM) can form extensive membrane invaginations known as cristae, which contain the respiratory chain and ATP synthase for oxidative phosphorylation. The molecular mechanisms regulating mitochondrial ultrastructure remain poorly understood. The Bin-Amphiphysin-Rvs (BAR) domain proteins are central regulators of diverse cellular processes related to membrane remodeling and dynamics. Whether BAR domain proteins are involved in sculpting membranes in specific submitochondrial compartments is largely unknown. In this study, we report FAM92A1 as a novel BAR domain protein localizes to the matrix side of the MIM. Loss of FAM92A1 caused a severe disruption to mitochondrial morphology and ultrastructure, impairing organelle bioenergetics. Furthermore, FAM92A1 displayed a membrane-remodeling activity in vitro, inducing a high degree of membrane curvature. Collectively, our findings uncover a role for a BAR domain protein as a critical organizer of the mitochondrial ultrastructure that is indispensable for mitochondrial function.

scholarly journals Unrestrained ESCRT-III drives chromosome fragmentation and micronuclear catastrophe

2019 ◽  
Author(s):  
Marina Vietri ◽  
Sebastian W. Schultz ◽  
Aurélie Bellanger ◽  
Carl M. Jones ◽  
Camilla Raiborg ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Genome Stability ◽  
Membrane Deformation ◽  
Chromosome Fragmentation ◽  
Inner Nuclear Membrane ◽  
Membrane Rupture ◽  
Membrane Fission ◽  
Stable Association ◽  
Inner Nuclear Membrane Protein

AbstractThe ESCRT-III membrane fission machinery1,2 restores nuclear envelope integrity during mitotic exit3,4 and interphase5,6. Whereas primary nuclei resealing takes minutes, micronuclear envelope ruptures appear irreversible and result in catastrophic collapse associated with chromosome fragmentation and rearrangements (chromothripsis), thought to be a major driving force in cancer development7-10. Despite its importance11-13, the mechanistic underpinnings of nuclear envelope sealing in primary nuclei and the defects observed in micronuclei remain largely unknown. Here we show that CHMP7, the nucleator of ESCRT-III filaments at the nuclear envelope3,14, and the inner nuclear membrane protein LEMD215 act as a compartmentalization sensor detecting the loss of nuclear integrity. In cells with intact nuclear envelope, CHMP7 is actively excluded from the nucleus to preclude its binding to LEMD2. Nuclear influx of CHMP7 results in stable association with LEMD2 at the inner nuclear membrane that licenses local polymerization of ESCRT-III. Tight control of nuclear CHMP7 levels is critical, as induction of nuclear CHMP7 mutants is sufficient to induce unrestrained growth of ESCRT-III foci at the nuclear envelope, causing dramatic membrane deformation, local DNA torsional stress, single-stranded DNA formation and fragmentation of the underlying chromosomes. At micronuclei, membrane rupture is not associated with repair despite timely recruitment of ESCRT-III. Instead, micronuclei inherently lack the capacity to restrict accumulation of CHMP7 and LEMD2. This drives unrestrained ESCRT-III recruitment, membrane deformation and DNA defects that strikingly resemble those at primary nuclei upon induction of nuclear CHMP7 mutants. Preventing ESCRT-III recruitment suppresses membrane deformation and DNA damage, without restoring nucleocytoplasmic compartmentalization. We propose that the ESCRT-III nuclear integrity surveillance machinery is a double-edged sword, as its exquisite sensitivity ensures rapid repair at primary nuclei while causing unrestrained polymerization at micronuclei, with catastrophic consequences for genome stability16-18.

scholarly journals Vesicle Size Regulates Nanotube Formation in the Cell

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Qian Peter Su ◽  
Wanqing Du ◽  
Qinghua Ji ◽  
Boxin Xue ◽  
Dong Jiang ◽  
...  
Keyword(s):  
Single Molecule ◽  
Model Systems ◽  
Organelle Biogenesis ◽  
Membrane Deformation ◽  
Vesicle Size ◽  
Cellular Processes ◽  
Vesicle Recycling ◽  
Physiological Relevance

Abstract Intracellular membrane nanotube formation and its dynamics play important roles for cargo transportation and organelle biogenesis. Regarding the regulation mechanisms, while much attention has been paid on the lipid composition and its associated protein molecules, effects of the vesicle size has not been studied in the cell. Giant unilamellar vesicles (GUVs) are often used for in vitro membrane deformation studies, but they are much larger than most intracellular vesicles and the in vitro studies also lack physiological relevance. Here, we use lysosomes and autolysosomes, whose sizes range between 100 nm and 1 μm, as model systems to study the size effects on nanotube formation both in vivo and in vitro. Single molecule observations indicate that driven by kinesin motors, small vesicles (100–200 nm) are mainly transported along the tracks while a remarkable portion of large vesicles (500–1000 nm) form nanotubes. This size effect is further confirmed by in vitro reconstitution assays on liposomes and purified lysosomes and autolysosomes. We also apply Atomic Force Microscopy (AFM) to measure the initiation force for nanotube formation. These results suggest that the size-dependence may be one of the mechanisms for cells to regulate cellular processes involving membrane-deformation, such as the timing of tubulation-mediated vesicle recycling.

scholarly journals VPS4 triggers constriction and cleavage of ESCRT-III helical filaments

2019 ◽  
Vol 5 (4) ◽  
pp. eaau7198 ◽  
Author(s):  
Sourav Maity ◽  
Christophe Caillat ◽  
Nolwenn Miguet ◽  
Guidenn Sulbaran ◽  
Gregory Effantin ◽  
...  
Keyword(s):  
High Speed ◽  
Membrane Structures ◽  
Vesicle Budding ◽  
Endosomal Sorting ◽  
Force Microscopy ◽  
Cellular Processes ◽  
Membrane Fission ◽  
End Caps

Many cellular processes such as endosomal vesicle budding, virus budding, and cytokinesis require extensive membrane remodeling by the endosomal sorting complex required for transport III (ESCRT-III). ESCRT-III protein family members form spirals with variable diameters in vitro and in vivo inside tubular membrane structures, which need to be constricted to proceed to membrane fission. Here, we show, using high-speed atomic force microscopy and electron microscopy, that the AAA-type adenosine triphosphatase VPS4 constricts and cleaves ESCRT-III CHMP2A-CHMP3 helical filaments in vitro. Constriction starts asymmetrically and progressively decreases the diameter of CHMP2A-CHMP3 tubular structure, thereby coiling up the CHMP2A-CHMP3 filaments into dome-like end caps. Our results demonstrate that VPS4 actively constricts ESCRT-III filaments and cleaves them before their complete disassembly. We propose that the formation of ESCRT-III dome-like end caps by VPS4 within a membrane neck structure constricts the membrane to set the stage for membrane fission.

scholarly journals Rab1 recruits WHAMM during membrane remodeling but limits actin nucleation

2016 ◽  
Vol 27 (6) ◽  
pp. 967-978 ◽  
Author(s):  
Ashley J. Russo ◽  
Alyssa J. Mathiowetz ◽  
Steven Hong ◽  
Matthew D. Welch ◽  
Kenneth G. Campellone
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Actin Assembly ◽  
Membrane Remodeling ◽  
Actin Nucleation ◽  
Factor Interactions ◽  
Cytoskeletal Rearrangements ◽  
Distinct Strategy ◽  
Small G Proteins

Small G-proteins are key regulatory molecules that activate the actin nucleation machinery to drive cytoskeletal rearrangements during plasma membrane remodeling. However, the ability of small G-proteins to interact with nucleation factors on internal membranes to control trafficking processes has not been well characterized. Here we investigated roles for members of the Rho, Arf, and Rab G-protein families in regulating WASP homologue associated with actin, membranes, and microtubules (WHAMM), an activator of Arp2/3 complex–mediated actin nucleation. We found that Rab1 stimulated the formation and elongation of WHAMM-associated membrane tubules in cells. Active Rab1 recruited WHAMM to dynamic tubulovesicular structures in fibroblasts, and an active prenylated version of Rab1 bound directly to an N-terminal domain of WHAMM in vitro. In contrast to other G-protein–nucleation factor interactions, Rab1 binding inhibited WHAMM-mediated actin assembly. This ability of Rab1 to regulate WHAMM and the Arp2/3 complex represents a distinct strategy for membrane remodeling in which a Rab G-protein recruits the actin nucleation machinery but dampens its activity.

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