Temporal Proteomics Profiling of Lipid Rafts in CCR6-Activated T Cells Reveals the Integration of Actin Cytoskeleton Dynamics

2010 ◽  
Vol 9 (1) ◽  
pp. 283-297 ◽  
Author(s):  
Shu-Ling Lin ◽  
Chih-Wei Chien ◽  
Chia-Li Han ◽  
Eric S.-W. Chen ◽  
Shao-Hsuan Kao ◽  
...  
Keyword(s):  
T Cells ◽  
Actin Cytoskeleton ◽  
Lipid Rafts ◽  
Activated T Cells ◽  
Cytoskeleton Dynamics

Spatial control of actin-based motility through plasmalemmal PtdIns(4,5)P2-rich raft assemblies.

2005 ◽  
Vol 72 ◽  
pp. 119-127 ◽  
Author(s):  
Tamara Golub ◽  
Caroni Pico
Keyword(s):  
Protein Kinase ◽  
Cell Surface ◽  
Actin Cytoskeleton ◽  
Lipid Rafts ◽  
Patch Clustering ◽  
Pkc Protein Kinase C ◽  
Membrane Bound ◽  
And Function ◽  
Cytoskeleton Dynamics ◽  
Syndrome Protein

The interactions of cells with their environment involve regulated actin-based motility at defined positions along the cell surface. Sphingolipid- and cholesterol-dependent microdomains (rafts) order proteins at biological membranes, and have been implicated in most signalling processes at the cell surface. Many membrane-bound components that regulate actin cytoskeleton dynamics and cell-surface motility associate with PtdIns(4,5)P2-rich lipid rafts. Although raft integrity is not required for substrate-directed cell spreading, or to initiate signalling for motility, it is a prerequisite for sustained and organized motility. Plasmalemmal rafts redistribute rapidly in response to signals, triggering motility. This process involves the removal of rafts from sites that are not interacting with the substrate, apparently through endocytosis, and a local accumulation at sites of integrin-mediated substrate interactions. PtdIns(4,5)P2-rich lipid rafts can assemble into patches in a process depending on PtdIns(4,5)P2, Cdc42 (cell-division control 42), N-WASP (neural Wiskott-Aldrich syndrome protein) and actin cytoskeleton dynamics. The raft patches are sites of signal-induced actin assembly, and their accumulation locally promotes sustained motility. The patches capture microtubules, which promote patch clustering through PKA (protein kinase A), to steer motility. Raft accumulation at the cell surface, and its coupling to motility are influenced greatly by the expression of intrinsic raft-associated components that associate with the cytosolic leaflet of lipid rafts. Among them, GAP43 (growth-associated protein 43)-like proteins interact with PtdIns(4,5)P2 in a Ca2+/calmodulin and PKC (protein kinase C)-regulated manner, and function as intrinsic determinants of motility and anatomical plasticity. Plasmalemmal PtdIns(4,5)P2-rich raft assemblies thus provide powerful organizational principles for tight spatial and temporal control of signalling in motility.

scholarly journals Fyn-Binding Protein (Fyb)/Slp-76–Associated Protein (Slap), Ena/Vasodilator-Stimulated Phosphoprotein (Vasp) Proteins and the Arp2/3 Complex Link T Cell Receptor (Tcr) Signaling to the Actin Cytoskeleton

2000 ◽  
Vol 149 (1) ◽  
pp. 181-194 ◽  
Author(s):  
Matthias Krause ◽  
Antonio S. Sechi ◽  
Marlies Konradt ◽  
David Monner ◽  
Frank B. Gertler ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Actin Cytoskeleton ◽  
T Cell Receptor ◽  
Cell Receptor ◽  
T Cell Signaling ◽  
Tcr Signaling ◽  
Activated T Cells ◽  
Antigen Presenting ◽  
Syndrome Protein

T cell receptor (TCR)-driven activation of helper T cells induces a rapid polarization of their cytoskeleton towards bound antigen presenting cells (APCs). We have identified the Fyn- and SLP-76–associated protein Fyb/SLAP as a new ligand for Ena/ vasodilator-stimulated phosphoprotein (VASP) homology 1 (EVH1) domains. Upon TCR engagement, Fyb/SLAP localizes at the interface between T cells and anti-CD3–coated beads, where Evl, a member of the Ena/VASP family, Wiskott-Aldrich syndrome protein (WASP) and the Arp2/3 complex are also found. In addition, Fyb/SLAP is restricted to lamellipodia of spreading platelets. In activated T cells, Fyb/SLAP associates with Ena/VASP family proteins and is present within biochemical complexes containing WASP, Nck, and SLP-76. Inhibition of binding between Fyb/SLAP and Ena/VASP proteins or WASP and the Arp2/3 complex impairs TCR-dependent actin rearrangement, suggesting that these interactions play a key role in linking T cell signaling to remodeling of the actin cytoskeleton.

scholarly journals Vav1/Rac-dependent actin cytoskeleton reorganization is required for lipid raft clustering in T cells

2001 ◽  
Vol 155 (3) ◽  
pp. 331-338 ◽  
Author(s):  
Martin Villalba ◽  
Kun Bi ◽  
Fernando Rodriguez ◽  
Yoshihiko Tanaka ◽  
Stephen Schoenberger ◽  
...  
Keyword(s):  
T Cells ◽  
Actin Cytoskeleton ◽  
Lipid Rafts ◽  
Lipid Raft ◽  
Large Scale ◽  
Actin Polymerization ◽  
Dominant Negative ◽  
Membrane Microdomains ◽  
Jurkat T Cells ◽  
Cytoskeleton Reorganization

Formation of the immunological synapse (IS) in T cells involves large scale molecular movements that are mediated, at least in part, by reorganization of the actin cytoskeleton. Various signaling proteins accumulate at the IS and are localized in specialized membrane microdomains, known as lipid rafts. We have shown previously that lipid rafts cluster and localize at the IS in antigen-stimulated T cells. Here, we provide evidence that lipid raft polarization to the IS depends on an intracellular pathway that involves Vav1, Rac, and actin cytoskeleton reorganization. Thus, lipid rafts did not translocate to the IS in Vav1-deficient (Vav1−/−) T cells upon antigen stimulation. Similarly, T cell receptor transgenic Jurkat T cells also failed to translocate lipid rafts to the IS when transfected with dominant negative Vav1 mutants. Raft polarization induced by membrane-bound cholera toxin cross-linking was also abolished in Jurkat T cells expressing dominant negative Vav1 or Rac mutants and in cells treated with inhibitors of actin polymerization. However, Vav overexpression that induced F-actin polymerization failed to induce lipid rafts clustering. Therefore, Vav is necessary, but not sufficient, to regulate lipid rafts clustering and polarization at the IS, suggesting that additional signals are required.

scholarly journals Structure and function of lipid rafts in human activated T cells

2005 ◽  
Vol 17 (6) ◽  
pp. 749-758 ◽  
Author(s):  
Shizue Tani-ichi ◽  
Koji Maruyama ◽  
Nami Kondo ◽  
Masakazu Nagafuku ◽  
Kazuya Kabayama ◽  
...  
Keyword(s):  
T Cells ◽  
Lipid Rafts ◽  
Structure And Function ◽  
Activated T Cells ◽  
And Function

scholarly journals Selective interaction of LAT (linker of activated T cells) with the open-active form of Lck in lipid rafts reveals a new mechanism for the regulation of Lck in T cells

2003 ◽  
Vol 371 (3) ◽  
pp. 907-915 ◽  
Author(s):  
Panagiotis S. KABOURIDIS
Keyword(s):  
T Cells ◽  
Lipid Rafts ◽  
Intramolecular Interaction ◽  
Active Form ◽  
Enzymic Activity ◽  
Open Form ◽  
Urea Denaturation ◽  
Activated T Cells ◽  
C Terminus ◽  
Raft Domains

In T cells, the lipid raft-associated Lck is strongly tyrosine phosphorylated and has reduced enzymic activity in contrast with the detergent-soluble pool, which has substantial activity. Lck tagged at the C-terminus (Lck/V5-His) was efficiently captured by epitope-specific reagents from the detergent-soluble fraction but not from lipid rafts. Binding was restored following urea denaturation, suggesting that Lck/V5-His is in a ‘closed’ conformation in these domains. In agreement with this hypothesis, the Tyr505 → Phe/V5-His and Arg154 → Lys/V5-His mutants, which disrupt the SH2-Tyr505 intramolecular interaction, were efficiently precipitated from lipid rafts. In contrast to Lck, Fyn/V5-His was precipitated equally well from both fractions. In the LAT (linker of activated T cells)-deficient J.CaM2 cells, Tyr505 phosphorylation of raft-associated Lck was reduced whereas its enzymic activity was elevated. This correlated with decreased levels of raft-localized Csk (C-terminal Src kinase) kinase. Increased tyrosine phosphorylation of Lck was restored in LAT-reconstituted J.CaM2 cells suggesting that LAT negatively regulates Lck activity in lipid rafts. Co-immunoprecipitation experiments from Tyr505 → Phe/V5-His-expressing cells revealed that LAT preferentially interacts with the ‘open’ form of Lck in T cell raft domains. These results demonstrate that, unlike the non-raft pool, Lck in lipid rafts has a ‘closed’-inactive structure, and that LAT plays a role in maintaining this conformation, possibly by facilitating critical associations within lipid rafts via its capacity to interact with the ‘open’ form of the kinase.

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