title: Ligand mobility modulates immunological synapse formation and T cell activation.
abstract: T cell receptor (TCR) engagement induces clustering and recruitment to the plasma membrane of many signaling molecules, including the protein tyrosine kinase zeta-chain associated protein of 70 kDa (ZAP70) and the adaptor SH2 domain-containing leukocyte protein of 76 kDa (SLP76). This molecular rearrangement results in formation of the immunological synapse (IS), a dynamic protein array that modulates T cell activation. The current study investigates the effects of apparent long-range ligand mobility on T cell signaling activity and IS formation. We formed stimulatory lipid bilayers on glass surfaces from binary lipid mixtures with varied composition, and characterized these surfaces with respect to diffusion coefficient and fluid connectivity. Stimulatory ligands coupled to these surfaces with similar density and orientation showed differences in their ability to activate T cells. On less mobile membranes, central supramolecular activation cluster (cSMAC) formation was delayed and the overall accumulation of CD3ζ at the IS was reduced. Analysis of signaling microcluster (MC) dynamics showed that ZAP70 MCs exhibited faster track velocity and longer trajectories as a function of increased ligand mobility, whereas movement of SLP76 MCs was relatively insensitive to this parameter. Actin retrograde flow was observed on all surfaces, but cell spreading and subsequent cytoskeletal contraction were more pronounced on mobile membranes. Finally, increased tyrosine phosphorylation and persistent elevation of intracellular Ca(2+) were observed in cells stimulated on fluid membranes. These results point to ligand mobility as an important parameter in modulating T cell responses.
authors.affiliation: Department of Chemistry, The Children's Hospital of Philadelphia and Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Chih-Jung
authors.initials: CJ
authors.lastname: Hsu
authors.forename: Wan-Ting
authors.initials: WT
authors.lastname: Hsieh
authors.forename: Abraham
authors.initials: A
authors.lastname: Waldman
authors.forename: Fiona
authors.initials: F
authors.lastname: Clarke
authors.forename: Eric S
authors.initials: ES
authors.lastname: Huseby
authors.forename: Janis K
authors.initials: JK
authors.lastname: Burkhardt
authors.forename: Tobias
authors.initials: T
authors.lastname: Baumgart
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D016000:Cluster Analysis; D006801:Humans; D054992:Immunological Synapses / Q000502:physiology*; D019169:Jurkat Cells; D008024:Ligands; D008051:Lipid Bilayers; D008055:Lipids / Q000737:chemistry; D008213:Lymphocyte Activation; D051379:Mice; D008810:Mice, Inbred C57BL; D018625:Microscopy, Atomic Force / Q000379:methods; D010750:Phosphoproteins / Q000378:metabolism; D011506:Proteins / Q000737:chemistry; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000166:cytology; D014443:Tyrosine / Q000737:chemistry; D051746:ZAP-70 Protein-Tyrosine Kinase / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D008024:Ligands; D008051:Lipid Bilayers; D008055:Lipids; D010750:Phosphoproteins; D011506:Proteins; C092947:SLP-76 signal Transducing adaptor proteins; D014443:Tyrosine; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human; D002118:Calcium