scholarly journals Rhes travels from cell to cell and transports Huntington disease protein via TNT-like protrusion

2019 ◽  
Vol 218 (6) ◽  
pp. 1972-1993 ◽  
Author(s):  
Manish Sharma ◽  
Srinivasa Subramaniam
Keyword(s):  
Huntington Disease ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Mutant Huntingtin ◽  
Tunneling Nanotubes ◽  
Neighboring Cell ◽  
Disease Protein ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Cell Cell

Tunneling nanotubes (TNT) are thin, membranous, tunnel-like cell-to-cell connections, but the mechanisms underlying their biogenesis or functional role remains obscure. Here, we report, Rhes, a brain-enriched GTPase/SUMO E3-like protein, induces the biogenesis of TNT-like cellular protrusions, “Rhes tunnels,” through which Rhes moves from cell to cell and transports Huntington disease (HD) protein, the poly-Q expanded mutant Huntingtin (mHTT). The formation of TNT-like Rhes tunnels requires the Rhes’s serine 33, C-terminal CAAX, and a SUMO E3-like domain. Electron microscopy analysis revealed that TNT-like Rhes tunnels appear continuous, cell–cell connections, and <200 nm in diameter. Live-cell imaging shows that Rhes tunnels establish contact with the neighboring cell and deliver Rhes-positive cargoes, which travel across the plasma membrane of the neighboring cell before entering it. The Rhes tunnels carry Rab5a/Lyso 20-positive vesicles and transport mHTT, but not normal HTT, mTOR, or wtTau proteins. SUMOylation-defective mHTT, Rhes C263S (cannot SUMOylate mHTT), or CRISPR/Cas9-mediated depletion of three isoforms of SUMO diminishes Rhes-mediated mHTT transport. Thus, Rhes promotes the biogenesis of TNT-like cellular protrusions and facilitates the cell–cell transport of mHTT involving SUMO-mediated mechanisms.

scholarly journals Live cell imaging and biophotonic methods reveal two types of mutant huntingtin inclusions

2013 ◽  
Vol 23 (9) ◽  
pp. 2324-2338 ◽  
Author(s):  
Nicholas S. Caron ◽  
Claudia L. Hung ◽  
Randy S. Atwal ◽  
Ray Truant
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Mutant Huntingtin

scholarly journals Biochemical and structural analysis of α-catenin in cell–cell contacts

2008 ◽  
Vol 36 (2) ◽  
pp. 141-147 ◽  
Author(s):  
Sabine Pokutta ◽  
Frauke Drees ◽  
Soichiro Yamada ◽  
W. James Nelson ◽  
William I. Weis
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Adherens Junction ◽  
Structural Data ◽  
Cell Contact ◽  
Related Protein ◽  
Imaging Studies ◽  
Actin Networks ◽  
Junction Formation ◽  
Cell Cell

Cadherins are transmembrane adhesion molecules that mediate homotypic cell–cell contact. In adherens junctions, the cytoplasmic domain of cadherins is functionally linked to the actin cytoskeleton through a series of proteins known as catenins. E-cadherin binds to β-catenin, which in turn binds to α-catenin to form a ternary complex. α-Catenin also binds to actin, and it was assumed previously that α-catenin links the cadherin–catenin complex to actin. However, biochemical, structural and live-cell imaging studies of the cadherin–catenin complex and its interaction with actin show that binding of β-catenin to α-catenin prevents the latter from binding to actin. Biochemical and structural data indicate that α-catenin acts as an allosteric protein whose conformation and activity changes depending on whether or not it is bound to β-catenin. Initial contacts between cells occur on dynamic lamellipodia formed by polymerization of branched actin networks, a process controlled by the Arp2/3 (actin-related protein 2/3) complex. α-Catenin can suppress the activity of Arp2/3 by competing for actin filaments. These findings lead to a model for adherens junction formation in which clustering of the cadherin–β-catenin complex recruits high levels of α-catenin that can suppress the Arp2/3 complex, leading to cessation of lamellipodial movement and formation of a stable contact. Thus α-catenin appears to play a central role in cell–cell contact formation.

scholarly journals Interaction of Huntington Disease Protein with Transcriptional Activator Sp1

2002 ◽  
Vol 22 (5) ◽  
pp. 1277-1287 ◽  
Author(s):  
Shi-Hua Li ◽  
Anna L. Cheng ◽  
Hui Zhou ◽  
Suzanne Lam ◽  
Manjula Rao ◽  
...  
Keyword(s):  
Huntington Disease ◽  
Cultured Cells ◽  
Growth Factor Receptor ◽  
Soluble Form ◽  
Mutant Huntingtin ◽  
Nuclear Effect ◽  
Polyglutamine Expansion ◽  
Disease Protein ◽  
Cellular Dysfunction

ABSTRACT Polyglutamine expansion causes Huntington disease (HD) and at least seven other neurodegenerative diseases. In HD, N-terminal fragments of huntingtin with an expanded glutamine tract are able to aggregate and accumulate in the nucleus. Although intranuclear huntingtin affects the expression of numerous genes, the mechanism of this nuclear effect is unknown. Here we report that huntingtin interacts with Sp1, a transcription factor that binds to GC-rich elements in certain promoters and activates transcription of the corresponding genes. In vitro binding and immunoprecipitation assays show that polyglutamine expansion enhances the interaction of N-terminal huntingtin with Sp1. In HD transgenic mice (R6/2) that express N-terminal-mutant huntingtin, Sp1 binds to the soluble form of mutant huntingtin but not to aggregated huntingtin. Mutant huntingtin inhibits the binding of nuclear Sp1 to the promoter of nerve growth factor receptor and suppresses its transcriptional activity in cultured cells. Overexpression of Sp1 reduces the cellular toxicity and neuritic extension defects caused by intranuclear mutant huntingtin. These findings suggest that the soluble form of mutant huntingtin in the nucleus may cause cellular dysfunction by binding to Sp1 and thus reducing the expression of Sp1-regulated genes.

scholarly journals Localized zones of Rho and Rac activities drive initiation and expansion of epithelial cell–cell adhesion

2007 ◽  
Vol 178 (3) ◽  
pp. 517-527 ◽  
Author(s):  
Soichiro Yamada ◽  
W. James Nelson
Keyword(s):  
Cell Adhesion ◽  
Actin Filaments ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
De Novo ◽  
Actin Dynamics ◽  
Lateral Expansion ◽  
Actomyosin Contractility ◽  
Spatiotemporal Coordination ◽  
Cell Cell

Spatiotemporal coordination of cell–cell adhesion involving lamellipodial interactions, cadherin engagement, and the lateral expansion of the contact is poorly understood. Using high-resolution live-cell imaging, biosensors, and small molecule inhibitors, we investigate how Rac1 and RhoA regulate actin dynamics during de novo contact formation between pairs of epithelial cells. Active Rac1, the Arp2/3 complex, and lamellipodia are initially localized to de novo contacts but rapidly diminish as E-cadherin accumulates; further rounds of activation and down-regulation of Rac1 and Arp2/3 occur at the contacting membrane periphery, and this cycle repeats as a restricted membrane zone that moves outward with the expanding contact. The cortical bundle of actin filaments dissolves beneath the expanding contacts, leaving actin bundles at the contact edges. RhoA and actomyosin contractility are activated at the contact edges and are required to drive expansion and completion of cell–cell adhesion. We show that zones of Rac1 and lamellipodia activity and of RhoA and actomyosin contractility are restricted to the periphery of contacting membranes and together drive initiation, expansion, and completion of cell–cell adhesion.

scholarly journals High-temperature live-cell imaging of cytokinesis, cell motility and cell-cell adhesion in the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius

2020 ◽  
Author(s):  
Arthur Charles-Orszag ◽  
Samuel J. Lord ◽  
R. Dyche Mullins
Keyword(s):  
Cell Division ◽  
High Temperature ◽  
Cell Motility ◽  
Cell Biology ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Sulfolobus Acidocaldarius ◽  
Model Organisms ◽  
Cell Cell

Significant technical challenges have limited the study of extremophile cell biology. For example, the absence of methods for performing high-resolution, live-cell imaging at high temperatures (>50°C) has impeded the study of cell motility and cell division in thermophilic archaea such as model organisms from the genus Sulfolobus. Here we describe a system for imaging samples at 75°C using high numerical aperture, oil-immersion lenses. With this system we observed and quantified the dynamics of cell division in the model thermoacidophilic crenarchaeon Sulfolobus acidocaldarius. In addition, we observed previously undescribed dynamic cell shape changes, cell motility, and cell-cell interactions, shedding significant new light on the high-temperature lifestyle of this organism.

scholarly journals F-pili dynamics by live-cell imaging

2008 ◽  
Vol 105 (46) ◽  
pp. 17978-17981 ◽  
Author(s):  
Margaret Clarke ◽  
Lucinda Maddera ◽  
Robin L. Harris ◽  
Philip M. Silverman
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Recipient Cell ◽  
Cell Communication ◽  
Live Cell ◽  
Cell Contact ◽  
Gram Negative Bacteria ◽  
Donor Cells ◽  
Cell Cell

Bacteria have evolved numerous mechanisms for cell–cell communication, many of which have important consequences for human health. Among these is conjugation, the direct transfer of DNA from one cell to another. For Gram-negative bacteria, conjugation requires thin, flexible filaments (conjugative pili) that are elaborated by DNA donor cells. The structure, function, and especially the dynamics of conjugative pili are poorly understood. Here, we have applied live-cell imaging to characterize the dynamics of F-pili (conjugative pili encoded by the F plasmid of Escherichia coli). We establish that F-pili normally undergo cycles of extension and retraction in the absence of any obvious triggering event, such as contact with a recipient cell. When made, such contacts are able to survive the shear forces felt by bacteria in liquid media. Our data emphasize the role of F-pilus flexibility both in efficiently sampling a large volume surrounding donor cells in liquid culture and in establishing and maintaining cell–cell contact. Additionally and unexpectedly, we infer that extension and retraction are accompanied by rotation about the long axis of the filament.

Material analysis techniques using the ion mill

1996 ◽  
Vol 54 ◽  
pp. 1034-1035
Author(s):  
J. P. Benedict ◽  
R. M. Anderson ◽  
S. J. Klepeis
Keyword(s):  
Polished Surface ◽  
Surface Material ◽  
Analysis Techniques ◽  
Material Analysis ◽  
Optical Inspection ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Argon Ions ◽  
The Impact ◽  
Scanning Electron

Ion mills equipped with flood guns can perform two important functions in material analysis; they can either remove material or deposit material. The ion mill holder shown in Fig. 1 is used to remove material from the polished surface of a sample for further optical inspection or SEM ( Scanning Electron Microscopy ) analysis. The sample is attached to a pohshing stud type SEM mount and placed in the ion mill holder with the polished surface of the sample pointing straight up, as shown in Fig 2. As the holder is rotating in the ion mill, Argon ions from the flood gun are directed down at the top of the sample. The impact of Argon ions against the surface of the sample causes some of the surface material to leave the sample at a material dependent, nonuniform rate. As a result, the polished surface will begin to develop topography during milling as fast sputtering materials leave behind depressions in the polished surface.

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