Stathmin Regulates Microtubule Dynamics and Microtubule Organizing Center Polarization in Activated T Cells

While it is well established that microtubules (MTs) facilitate various stages of virus replication, how viruses actively control MT dynamics and functions remains less-well understood. Recent work has begun to reveal how several viruses exploit End-Binding (EB) proteins and their associated microtubule plus-end tracking proteins (+TIPs), in particular to enable loading of viral particles onto MTs for retrograde transport during early stages of infection. But distinct from other viruses studied to date, at mid-to-late stages of its unusually protracted replication cycle human cytomegalovirus (HCMV) increases the expression of all three EB family members. This occurs coincident with the formation of a unique structure termed the Assembly Compartment (AC), which serves as a Golgi-derived MT organizing center. Together, the AC and distinct EB proteins enable HCMV to increase the formation of dynamic and acetylated microtubule subsets to regulate distinct aspects of the viral replication cycle. Here, we reveal that HCMV also exploits EB-independent +TIP pathways by specifically increasing the expression of Transforming Acidic Coiled Coil protein 3 (TACC3) to recruit the MT polymerase, chTOG from initial sites of MT nucleation in the AC out into the cytosol, thereby increasing dynamic MT growth. Preventing TACC3 increases or depleting chTOG impaired MT polymerization, resulting in defects in early versus late endosome organization in and around the AC as well as defects in viral trafficking and spread. Our findings provide the first example of a virus that actively exploits EB-independent +TIP pathways to regulate MT dynamics and control late stages of virus replication. Importance Diverse viruses rely on host cell microtubule networks in order to transport viral particles within the dense cytoplasmic environment and to control the broader architecture of the cell to facilitate their replication. Yet precisely how viruses regulate the dynamic behavior and function of microtubule filaments remains poorly defined. We recently showed that the Assembly Compartment (AC) formed by human cytomegalovirus (HCMV) acts as a Golgi-derived microtubule organizing center. Here, we show that at mid-to-late stages of infection, HCMV increases the expression of Transforming Acidic Coiled Coil protein 3 (TACC3) in order to control the localization of the microtubule polymerase, chTOG. This in turn enables HCMV to generate dynamic microtubule subsets that organize endocytic vesicles in and around the AC and facilitate the transport of new viral particles released into the cytosol. Our findings reveal the first instance of viral targeting of TACC3 to control microtubule dynamics and virus spread.

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Imaging of plasmacytoid dendritic cell interactions with T cells

Blood ◽

10.1182/blood-2008-02-139865 ◽

2009 ◽

Vol 113 (1) ◽

pp. 75-84 ◽

Cited By ~ 33

Author(s):

María Mittelbrunn ◽

Gloria Martínez del Hoyo ◽

María López-Bravo ◽

Noa B. Martín-Cofreces ◽

Alix Scholer ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Plasmacytoid Dendritic Cell ◽

Time Lapse ◽

Cell Interactions ◽

Type I ◽

Microtubule Organizing Center ◽

Immune Synapse ◽

Organizing Center

Abstract Plasmacytoid dendritic cells (pDCs) efficiently produce type I interferon and participate in adaptive immune responses, although the molecular interactions between pDCs and antigen-specific T cells remain unknown. This study examines immune synapse (IS) formation between murine pDCs and CD4+ T cells. Mature pDCs formed canonical ISs, involving relocation to the contact site of the microtubule-organizing center, F-actin, protein kinase C-θ, and pVav, and activation of early signaling molecules in T cells. However, immature pDCs were less efficient at forming conjugates with T cells and inducing IS formation, microtubule-organizing center translocation, and T-cell signaling and activation. Time-lapse videomicroscopy and 2-photon in vivo imaging of pDC–T-cell interactions revealed that immature pDCs preferentially mediated transient interactions, whereas mature pDCs promoted more stable contacts. Our data indicate that, under steady-state conditions, pDCs preferentially establish transient contacts with naive T cells and show a very modest immunogenic capability, whereas on maturation, pDCs are able to form long-lived contacts with T cells and significantly enhance their capacity to activate these lymphocytes.

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Faculty Opinions recommendation of Cytokine secretion by CD4+ T cells at the immunological synapse requires Cdc42-dependent local actin remodeling but not microtubule organizing center polarity.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.717965358.793466415 ◽

2012 ◽

Author(s):

Cosima Tatiana Baldari

Keyword(s):

T Cells ◽

Cd4 T Cells ◽

Immunological Synapse ◽

Cytokine Secretion ◽

Microtubule Organizing Center ◽

Actin Remodeling ◽

Organizing Center

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Centrosome-dependent microtubule organization sets the conditions for axon formation

10.1101/2020.12.29.424696 ◽

2020 ◽

Author(s):

Durga Praveen Meka ◽

Oliver Kobler ◽

Souhaila Wuesthoff ◽

Birgit Schwanke ◽

Christoph Krisp ◽

...

Keyword(s):

Microtubule Dynamics ◽

Growth Speed ◽

Microtubule Organizing Center ◽

Microtubule Organization ◽

Axon Elongation ◽

Axon Development ◽

Organizing Center ◽

Cytoskeleton Remodeling ◽

Axon Specification ◽

Developing Neurons

AbstractMicrotubule remodeling is critical during axon development when the more stable microtubules populate the axon. It is not completely understood, however, how this local cytoskeleton remodeling is coordinated. The centrosome, the main microtubule-organizing center (MTOC), has been suggested to be crucial for axon specification 1–5. Conversely, it was proposed that axon elongation is independent of centrosomal functions 6. Here we report that microtubule dynamics in early neurons follow a radial organization which establishes the conditions for the axon formation. Using high-resolution microscopy of early developing neurons, we demonstrate that few somatic acetylated microtubules are restricted near the centrosome. At later stages, however, acetylated microtubules spread out in the soma and concentrate in the growing axon. Furthermore, live-imaging of the microtubule plus-end binding protein EB3 in early differentiating neurons shows that growing microtubules have increased length and growth speed near the MTOC, suggesting local differences that might favor axon selection. Importantly, due to the lack of somatic stable/acetylated microtubules in early developing neurons, disruption of the F-actin cytoskeleton does not induce multiple axons, as it does at later stages of differentiation. Finally, we demonstrate that overexpression of the centrosomal protein 120 (Cep120), known for promoting microtubule acetylation and stabilization, induces multiple axons, while its downregulation decreases the content of proteins regulating microtubule dynamics and stability, hence hampering axon formation. Collectively, our data show that early centrosome-dependent microtubule organization contributes to axon formation.

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A new calcium-activated dynein adaptor protein, CRACR2a, regulates clathrin-independent endocytic traffic in T cells

10.1101/354498 ◽

2018 ◽

Author(s):

Yuxiao Wang ◽

Walter Huynh ◽

Taylor D. Skokan ◽

Ronald D. Vale

Keyword(s):

T Cells ◽

Coiled Coil ◽

Adaptor Protein ◽

Adaptor Proteins ◽

Cytoplasmic Dynein ◽

Rab Gtpase ◽

Rab Gtpases ◽

Microtubule Organizing Center ◽

Activated T Cells

AbstractCytoplasmic dynein is a microtubule minus-end-directed motor that transports numerous intracellular cargoes. Mammalian dynein transport is initiated by coiled-coil adaptor proteins that 1) join dynein and its co-factor dynactin into a complex capable of processive motility, and 2) interact with a cargo-bound receptor, which is frequently a Rab GTPase on an organelle. Here, we report two novel dynein adaptors, CRACR2a and Rab45, which have a coiled-coil adaptor domain, a pair of EF hands, and a Rab GTPase domain fused into a single polypeptide. We find that CRACR2a-mediated dynein-dynactin motility is activated by calcium in vitro and in cells. In activated T cells, CRACR2a localizes to clathrin-independent endosomes that require microtubule-based transport to detach from the actin cortex and travel towards the microtubule organizing center. Together these results represent the first known examples of Rab GTPases that directly act as dynein adaptors and implicate CRACR2a-dynein in regulation of endocytic trafficking in T cells.

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LARG and mDia1 Link Gα12/13 to Cell Polarity and Microtubule Dynamics

Molecular Biology of the Cell ◽

10.1091/mbc.e06-11-1045 ◽

2008 ◽

Vol 19 (1) ◽

pp. 30-40 ◽

Cited By ~ 38

Author(s):

Polyxeni Goulimari ◽

Helga Knieling ◽

Ulrike Engel ◽

Robert Grosse

Keyword(s):

Cell Polarity ◽

Glycogen Synthase ◽

Cell Polarization ◽

Microtubule Dynamics ◽

Microtubule Organizing Center ◽

Embryonic Fibroblasts ◽

Extracellular Signals ◽

Receptor Signal ◽

Organizing Center ◽

G Protein Coupled

Regulation of cell polarity is a process observed in all cells. During directed migration, cells orientate their microtubule cytoskeleton and the microtubule-organizing-center (MTOC), which involves integrins and downstream Cdc42 and glycogen synthase kinase-3β activity. However, the contribution of G protein-coupled receptor signal transduction for MTOC polarity is less well understood. Here, we report that the heterotrimeric Gα12 and Gα13 proteins are necessary for MTOC polarity and microtubule dynamics based on studies using Gα12/13-deficient mouse embryonic fibroblasts. Cell polarization involves the Gα12/13-interacting leukemia-associated RhoGEF (LARG) and the actin-nucleating diaphanous formin mDia1. Interestingly, LARG associates with pericentrin and localizes to the MTOC and along microtubule tracks. We propose that Gα12/13 proteins exert essential functions linking extracellular signals to microtubule dynamics and cell polarity via RhoGEF and formin activity.

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