Spatial and temporal dynamics of T cell receptor signaling with a photoactivatable agonist.
The precise timing of signals downstream of the T cell receptor (TCR) is poorly understood. To address this problem, we prepared major histocompatibility complexes containing an antigenic peptide that is biologically inert until exposed to ultraviolet (UV) light. UV irradiation of these complexes in contact with cognate T cells enabled the high-resolution temporal analysis of signaling. Phosphorylation of the LAT adaptor molecule was observed in 4 s, and diacylglycerol production and calcium flux was observed in 6-7 s. TCR activation also induced cytoskeletal polarization within 2 min. Antibody blockade of CD4 reduced the intensity of LAT phosphorylation and the speed of calcium flux. Furthermore, strong desensitization of diacylglycerol production, but not LAT phosphorylation, occurred shortly after TCR activation, suggesting that different molecular events play distinct signal-processing roles. These results establish the speed and localization of early signaling steps, and have important implications regarding the overall structure of the network.
Huse
Morgan
M
Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Klein
Lawrence O
LO
Girvin
Andrew T
AT
Faraj
Joycelyn M
JM
Li
Qi-Jing
QJ
Kuhns
Michael S
MS
Davis
Mark M
MM
D000595:Amino Acid Sequence; D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects / Q000276:immunology; D045304:Cytochromes c / Q000502:physiology; D000431:Ethanol / Q000031:analogs & derivatives / Q000494:pharmacology; D000949:Histocompatibility Antigens Class II / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology / Q000528:radiation effects*; D051379:Mice; D008969:Molecular Sequence Data; D009036:Moths / Q000201:enzymology; D009578:Nitrobenzenes / Q000494:pharmacology; D010455:Peptides / Q000819:agonists / Q000378:metabolism / Q000502:physiology; D019920:Photoaffinity Labels / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000819:agonists* / Q000378:metabolism / Q000502:physiology*; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects*; D014466:Ultraviolet Rays
D000949:Histocompatibility Antigens Class II; C024138:I-E-antigen; D009578:Nitrobenzenes; D010455:Peptides; D019920:Photoaffinity Labels; D011948:Receptors, Antigen, T-Cell; C063593:1-(2-nitrophenyl)ethanol; D000431:Ethanol; D045304:Cytochromes c