Harnessing actin dynamics for clathrin-mediated endocytosis

Perillo

et al. 2016

Biophysical Journal

Whereas important discoveries made by single-particle tracking have changed our view of the plasma membrane organization and motor protein dynamics in the past three decades, experimental studies of intracellular processes using singleparticle tracking are rather scarce because of the lack of three-dimensional (3D) tracking capacity. In this study we use a newly developed 3D single-particle tracking method termed TSUNAMI (Tracking of Single particles Using Nonlinear And Multiplexed Illumination) to investigate epidermal growth factor receptor (EGFR) trafficking dynamics in live cells at 16/43 nm (xy/z) spatial resolution, with track duration ranging from 2 to 10 min and vertical tracking depth up to tens of microns. To analyze the long 3D trajectories generated by the TSUNAMI microscope, we developed a trajectory analysis algorithm, which reaches 81% segment classification accuracy in control experiments (termed simulated movement experiments). When analyzing 95 EGF-stimulated EGFR trajectories acquired in live skin cancer cells, we find that these trajectories can be separated into three groups-immobilization (24.2%), membrane diffusion only (51.6%), and transport from membrane to cytoplasm (24.2%). When EGFRs are membrane-bound, they show an interchange of Brownian diffusion and confined diffusion. When EGFRs are internalized, transitions from confined diffusion to directed diffusion and from directed diffusion back to confined diffusion are clearly seen. This observation agrees well with the model of clathrin-mediated endocytosis.

Section: Monitoring Egfr Trafficking From Membrane To Cytoplasm In LImentioning

confidence: 98%

Segmentation of 3D Trajectories Acquired by TSUNAMI Microscope: An Application to EGFR Trafficking

Perillo

et al. 2016

Biophysical Journal

“…Polymerization of thin, long bundles of filaments created jointly by ARP2/3, formins and crosslinker activity creates filopodia, while generation of short, branched filaments by ARP2/3 and capping proteins over extended areas creates lamellipodia 22 . Pushing by small foci of branched actin can also work together with membrane-bending proteins to deform the membrane inwards for endocytosis 23 .…”

Section: [H1] Structure Of the Cortical Cytoskeletonmentioning

confidence: 99%

Cytoskeletal control of B cell responses to antigens

Tolar

2017

Nat Rev Immunol

118

The actin cytoskeleton is essential for cell mechanics and has increasingly been implicated in the regulation of cell signalling. In B cells, the actin cytoskeleton couples extensively to B cell receptor (BCR) signalling pathways and its defects can either enhance or suppress B cell activation. Recent insights from single-cell imaging and biophysical techniques suggest that actin orchestrates BCR signalling at the plasma membrane through effects on protein diffusion, and that it regulates antigen discrimination through the biomechanics of immune synapses. These mechanical functions also play a role in the adaptation of B cell subsets to specialized tasks during antibody responses.3

“…13). It has recently been proposed, for both amoeba and yeast, that myosin-I motors function not in transport per se, but rather in the regulation and organization of the actin nucleation-polymerization machinery 2,3 . Whether this might also be true for animal cells has not as yet been studied.…”

Section: Processivitymentioning

confidence: 99%

“…Most of the data referred to have been acquired from animal systems, except where mentioned otherwise. The basic properties of molecular motors, such as how they generate movement and tension, have been discussed in a compilation of reviews elsewhere 1 , although some of the salient features are summarized in BOX 1. For a discussion of the additional mechanisms by which motors produce movement, such as harnessing the power of actin polymerization, readers are referred to recent reviews 2,3 . The diversity of the motors that are engaged in these processes is reflected in the complexity of attachment modes to their diverse cargoes, which are summarized in BOX 2. More complete outlines of cargobinding mechanisms can be found in recent reviews [4][5][6] .…”

mentioning

confidence: 99%

Powering membrane traffic in endocytosis and recycling

Soldati

Schliwa

2006

Nat Rev Mol Cell Biol

311

279

| Early in evolution, the diversification of membrane-bound compartments that characterize eukaryotic cells was accompanied by the elaboration of molecular machineries that mediate intercompartmental communication and deliver materials to specific destinations. Molecular motors that move on tracks of actin filaments or microtubules mediate the movement of organelles and transport between compartments. The subjects of this review are the motors that power the transport steps along the endocytic and recycling pathways, their modes of attachment to cargo and their regulation.The emergence of eukaryotic cells was accompanied by the development of endomembrane systems that compartmentalize biochemical pathways and biosynthetic processes. An evolutionary trend towards increasing complexity and subcellular specialization necessitated the development of machineries for intercompartmental communication. Molecular assemblies evolved to confer specificity to the interactions of donor and acceptor compartments (budding, sorting, tethering and fusion). Cytoskeletal polymers such as actin filaments and microtubules, which help segregate genetic material and determine cell shape and subcellular architecture, were co-opted as tracks for transport. In animal cells, microtubules support long-range transport, whereas the more flexible actin filaments serve short-range movements both near the cell periphery and in the cell interior. Also, actin filaments can be woven into dense networks of short fibres through the action of crosslinkers and branchpromoting complexes. On the other hand, in many plant cells, bundled actin filaments can form extended tracks for long-distance transport. Owing to the gel-like nature of the cytoplasm, the Brownian movement of organelles is severely restricted and cargoes have to be transported to their destinations at the expense of energy. Furthermore, seemingly random, but motor-dependent, movement is proposed to increase the probability of vesicle collisions and therefore promote fusion events. Movement is powered by three ATP-dependent motors: myosin, kinesin and dynein. Over the course of eukaryotic evolution, these motors have diversified into a large number of families with specific tasks (FIG. 1).Here, we review the involvement of molecular motors in the movement of endosomes and in vesicular trafficking along the endocytic and recycling pathways. These pathways are summarized in FIG. 2. We do not, however,