Low Ligand Requirement for Deletion and Lack of Synapses in Positive Selection Enforce the Gauntlet of Thymic T Cell Maturation

Howard Hughes Medical Institute and The Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Immunity (Impact Factor: 21.56). 12/2008; 29(5):734-45. DOI: 10.1016/j.immuni.2008.09.014
Source: PubMed


Immature double-positive (CD4(+)CD8(+)) thymocytes respond to negatively selecting peptide-MHC ligands by forming an immune synapse that sustains contact with the antigen-presenting cell (APC). Using fluorescently labeled peptides, we showed that as few as two agonist ligands could promote APC contact and subsequent apoptosis in reactive thymocytes. Furthermore, we showed that productive signaling for positive selection, as gauged by nuclear translocation of a green fluorescent protein (GFP)-labeled NFATc construct, did not involve formation of a synapse between thymocytes and selecting epithelial cells in reaggregate thymus cultures. Antibody blockade of endogenous positively selecting ligands prevented NFAT nuclear accumulation in such cultures and reversed NFAT accumulation in previously stimulated thymocytes. Together, these data suggest a "gauntlet" model in which thymocytes mature by continually acquiring and reacquiring positively selecting signals without sustained contact with epithelial cells, thereby allowing them to sample many cell surfaces for potentially negatively selecting ligands.

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    • " by a T cell and an APC , we acquired a differ - ential interference contrast ( DIC ) image at the focal plane , a z - stack DIC image , a z - stack allophycocyanin image , and two consecutive z - stacks of QD images . Each z - stack is composed of 1 mm sections ( 21 of these ) or 0 . 8 mm sections ( 26 of these ) visualized across the whole APC ( Ebert et al . , 2008 ; Irvine et al . , 2002 ; Purbhoo et al . , 2004 ) . The same z - stack DIC and allophycocyanin images were collected for the quantum allophycocyanin calibration beads ( Bangs Labora - tories ) during each T cell sensitivity experiment for fluorescence calibration and cytokine release quantification ( Figures S3B and S3C ) . Details are"
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