Lipid order and molecular assemblies in the plasma membrane of eukaryotic cells

2009 ◽  
Vol 37 (5) ◽  
pp. 1056-1060 ◽  
Author(s):  
Marek Cebecauer ◽  
Dylan M. Owen ◽  
Anna Markiewicz ◽  
Anthony I. Magee
Keyword(s):  
Plasma Membrane ◽  
Physical Properties ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Eukaryotic Cells ◽  
Dynamic Nature ◽  
Membrane Components ◽  
Technological Improvements

Multimolecular assemblies on the plasma membrane exhibit dynamic nature and are often generated during the activation of eukaryotic cells. The role of lipids and their physical properties in helping to control the existence of these structures is discussed. Technological improvements for live cell imaging of membrane components are also reviewed.

scholarly journals EB1 binding restricts STIM1 translocation to ER–PM junctions and regulates store-operated Ca2+ entry

2018 ◽  
Vol 217 (6) ◽  
pp. 2047-2058 ◽  
Author(s):  
Chi-Lun Chang ◽  
Yu-Ju Chen ◽  
Carlo Giovanni Quintanilla ◽  
Ting-Sung Hsieh ◽  
Jen Liou
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Cellular Functions ◽  
Binding Ability ◽  
Trapping Mechanism ◽  
Spatiotemporal Regulation

The endoplasmic reticulum (ER) Ca2+ sensor STIM1 forms oligomers and translocates to ER–plasma membrane (PM) junctions to activate store-operated Ca2+ entry (SOCE) after ER Ca2+ depletion. STIM1 also interacts with EB1 and dynamically tracks microtubule (MT) plus ends. Nevertheless, the role of STIM1–EB1 interaction in regulating SOCE remains unresolved. Using live-cell imaging combined with a synthetic construct approach, we found that EB1 binding constitutes a trapping mechanism restricting STIM1 targeting to ER–PM junctions. We further showed that STIM1 oligomers retain EB1 binding ability in ER Ca2+-depleted cells. By trapping STIM1 molecules at dynamic contacts between the ER and MT plus ends, EB1 binding delayed STIM1 translocation to ER–PM junctions during ER Ca2+ depletion and prevented excess SOCE and ER Ca2+ overload. Our study suggests that STIM1–EB1 interaction shapes the kinetics and amplitude of local SOCE in cellular regions with growing MTs and contributes to spatiotemporal regulation of Ca2+ signaling crucial for cellular functions and homeostasis.

scholarly journals EB1 binding provides a diffusion trap mechanism regulating STIM1 localization and Ca2+ signaling

2017 ◽  
Author(s):  
Chi-Lun Chang ◽  
Yu-Ju Chen ◽  
Jen Liou
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Time Lapse ◽  
Cellular Functions ◽  
Binding Ability ◽  
Spatiotemporal Regulation ◽  
Time Lapse Imaging

AbstractThe endoplasmic reticulum (ER) Ca2+ sensor STIM1 forms oligomers and translocates to ER-plasma membrane (PM) junctions to activate store-operated Ca2+ entry (SOCE) following ER Ca2+ depletion. STIM1 also directly interacts with end binding protein 1 (EB1) at microtubule (MT) plus-ends and resembles comet-like structures during time-lapse imaging. Nevertheless, the role of STIM1-EB1 interaction in regulating SOCE remains unresolved. Using live-cell imaging combined with pharmacological perturbation and a reconstitution approach, we revealed that EB1 binding constitutes a diffusion trap mechanism restricting STIM1 targeting to ER-PM junctions. We further showed that STIM1 oligomers retain EB1 binding ability in ER Ca2+-depleted cells. EB1 binding delayed the translocation of STIM1 oligomers to ER-PM junctions and recaptured STIM1 to prevent excess SOCE and ER Ca2+ overload. Thus, the counterbalance of EB1 binding and PM targeting of STIM1 shapes the kinetics and amplitude of local SOCE in regions with growing MTs, and contributes to precise spatiotemporal regulation of Ca2+ signaling crucial for cellular functions and homeostasis.SummarySTIM1 activates store-operated Ca2+ entry (SOCE) by translocating to endoplasmic reticulum-plasma membrane junctions. Chang et al. revealed that STIM1 localization and SOCE are regulated by a diffusion trap mechanism mediated by STIM1 binding to EB1 at growing microtubule ends.

scholarly journals Distinct acto/myosin-I structures associate with endocytic profiles at the plasma membrane

2008 ◽  
Vol 180 (6) ◽  
pp. 1219-1232 ◽  
Author(s):  
Fatima-Zahra Idrissi ◽  
Helga Grötsch ◽  
Isabel M. Fernández-Golbano ◽  
Cristina Presciatto-Baschong ◽  
Howard Riezman ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Immunoelectron Microscopy ◽  
Live Cell Imaging ◽  
Actin Polymerization ◽  
Cell Imaging ◽  
Live Cell ◽  
Endocytic Vesicle ◽  
Myosin I ◽  
Initial Membrane ◽  
Membrane Bending

Endocytosis in yeast requires actin and clathrin. Live cell imaging has previously shown that massive actin polymerization occurs concomitant with a slow 200-nm inward movement of the endocytic coat (Kaksonen, M., Y. Sun, and D.G. Drubin. 2003. Cell. 115:475–487). However, the nature of the primary endocytic profile in yeast and how clathrin and actin cooperate to generate an endocytic vesicle is unknown. In this study, we analyze the distribution of nine different proteins involved in endocytic uptake along plasma membrane invaginations using immunoelectron microscopy. We find that the primary endocytic profiles are tubular invaginations of up to 50 nm in diameter and 180 nm in length, which accumulate the endocytic coat components at the tip. Interestingly, significant actin labeling is only observed on invaginations longer than 50 nm, suggesting that initial membrane bending occurs before initiation of the slow inward movement. We also find that in the longest profiles, actin and the myosin-I Myo5p form two distinct structures that might be implicated in vesicle fission.

Role of Doc2α in dense-core vesicle exocytosis in PC12 cells revealed by live cell imaging

2011 ◽  
Vol 71 ◽  
pp. e213
Author(s):  
Takashi Tsuboi ◽  
Yasunori Mori ◽  
Hideki Matsui ◽  
Ryo Aoki ◽  
Manami Oya ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Dense Core ◽  
Dense Core Vesicle ◽  
Vesicle Exocytosis

scholarly journals Role of cytoplasmic dynein in the axonal transport of microtubules and neurofilaments

2005 ◽  
Vol 168 (5) ◽  
pp. 697-703 ◽  
Author(s):  
Yan He ◽  
Franto Francis ◽  
Kenneth A. Myers ◽  
Wenqian Yu ◽  
Mark M. Black ◽  
...  
Keyword(s):  
Rna Interference ◽  
Axonal Transport ◽  
Heavy Chain ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Sympathetic Neurons ◽  
Cytoplasmic Dynein ◽  
Live Cell ◽  
Cytoskeletal Elements

Recent studies have shown that the transport of microtubules (MTs) and neurofilaments (NFs) within the axon is rapid, infrequent, asynchronous, and bidirectional. Here, we used RNA interference to investigate the role of cytoplasmic dynein in powering these transport events. To reveal transport of MTs and NFs, we expressed EGFP-tagged tubulin or NF proteins in cultured rat sympathetic neurons and performed live-cell imaging of the fluorescent cytoskeletal elements in photobleached regions of the axon. The occurrence of anterograde MT and retrograde NF movements was significantly diminished in neurons that had been depleted of dynein heavy chain, whereas the occurrence of retrograde MT and anterograde NF movements was unaffected. These results support a cargo model for NF transport and a sliding filament model for MT transport.

Live cell imaging of interleukin-6-induced targeting of “transcription factor” STAT3 to sequestering endosomes in the cytoplasm

2007 ◽  
Vol 293 (4) ◽  
pp. C1374-C1382 ◽  
Author(s):  
Fang Xu ◽  
Somshuvra Mukhopadhyay ◽  
Pravin B. Sehgal
Keyword(s):  
Plasma Membrane ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Direct Evidence ◽  
Live Cell ◽  
Dominant Negative ◽  
Adapter Protein ◽  
Toll Like Receptor ◽  
Caveolin 1 ◽  
Induced Signal

Signal transducer and activator of transcription (STAT) family transcription factors are classically viewed as transducing cytokine- and growth factor-activated signals from the plasma membrane to the cell nucleus for the purpose of activating transcription. We report live cell imaging studies of fluorescently labeled STAT3 expressed in Hep3B hepatocytes that reveal interleukin (IL)-6-activated targeting of STAT3 and PY-STAT3 to relatively long-lived sequestering endosomes in the cytoplasm. This targeting was rapid but transient, required phosphorylation and integrity of Tyr 705 in STAT3, and was blocked by nocodazole, geldanamycin, and indirubin E804 and by overexpression of wild-type caveolin-1. Strikingly, overexpression of the dominant-negative (DN) mutant K44A of the GTPase dynamin II led to marked constitutive accumulation of STAT3 in the endocytic compartment with depletion of the STAT3 nuclear pool. Subsets of the native and K44A-generated STAT3- and PY-STAT3-sequestering endosomes colocalized with MyD88, an adapter protein that integrates pathways of Toll-like receptor and IL-1 transcriptional signaling and stabilization of mRNAs. These data provide direct evidence for the cytokine-induced “signal transduction” by STAT3 from the plasma membrane to a cytoplasmic membrane destination for yet to be elucidated function(s) in the cytoplasm including prolongation of signaling and/or cross talk.

scholarly journals ARL3 activation requires the co-GEF BART and effector-mediated turnover

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Yasmin ElMaghloob ◽  
Begoña Sot ◽  
Michael J McIlwraith ◽  
Esther Garcia ◽  
Tamas Yelland ◽  
...  
Keyword(s):  
G Protein ◽  
Primary Cilium ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Guanine Exchange Factor ◽  
Bona Fide ◽  
Modified Proteins ◽  
Adp Ribosylation Factor

The ADP-ribosylation factor-like 3 (ARL3) is a ciliopathy G-protein which regulates the ciliary trafficking of several lipid-modified proteins. ARL3 is activated by its guanine exchange factor (GEF) ARL13B via an unresolved mechanism. BART is described as an ARL3 effector which has also been implicated in ciliopathies, although the role of its ARL3 interaction is unknown. Here, we show that, at physiological GTP:GDP levels, human ARL3GDP is weakly activated by ARL13B. However, BART interacts with nucleotide-free ARL3 and, in concert with ARL13B, efficiently activates ARL3. In addition, BART binds ARL3GTP and inhibits GTP dissociation, thereby stabilising the active G-protein; the binding of ARL3 effectors then releases BART. Finally, using live cell imaging, we show that BART accesses the primary cilium and colocalises with ARL13B. We propose a model wherein BART functions as a bona fide co-GEF for ARL3 and maintains the active ARL3GTP, until it is recycled by ARL3 effectors.

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