Identification of self through two-dimensional chemistry and synapses.

Cells in the immune and nervous systems communicate through informational synapses. The two-dimensional chemistry underlying the process of synapse formation is beginning to be explored using fluorescence imaging and mechanical techniques. Early analysis of two-dimensional kinetic rates (k(on) and k(off)) and equilibrium constants (K(d)) provides a number of biological insights. First, there are two regimes for adhesion-one disordered with slow k(on) and the other self-ordered with 10(4)-fold faster k(on). Despite huge variation in two-dimensional k(on), the two-dimensional k(off) is like k(off) in solution, and two-dimensional k(off) is more closely related to intrinsic properties of the interaction than the two-dimensional k(on). Thus difference in k(off) can be used to set signaling thresholds. Early signaling complexes are compartmentalized to generate synergistic signaling domains. Immune antigen receptor components have a role in neural synapse editing. This suggests significant parallels in informational synapse formation based on common two-dimensional chemistry and signaling strategies.

The immunological synapse and CD28-CD80 interactions.

According to the two-signal model of T cell activation, costimulatory molecules augment T cell receptor (TCR) signaling, whereas adhesion molecules enhance TCR-MHC-peptide recognition. The structure and binding properties of CD28 imply that it may perform both functions, blurring the distinction between adhesion and costimulatory molecules. Our results show that CD28 on naïve T cells does not support adhesion and has little or no capacity for directly enhancing TCR-MHC-peptide interactions. Instead of being dependent on costimulatory signaling, we propose that a key function of the immunological synapse is to generate a cellular microenvironment that favors the interactions of potent secondary signaling molecules, such as CD28.

The immunological synapse.

The adaptive immune response is initiated by the interaction of T cell antigen receptors with major histocompatibility complex molecule-peptide complexes in the nanometer scale gap between a T cell and an antigen-presenting cell, referred to as an immunological synapse. In this review we focus on the concept of immunological synapse formation as it relates to membrane structure, T cell polarity, signaling pathways, and the antigen-presenting cell. Membrane domains provide an organizational principle for compartmentalization within the immunological synapse. T cell polarization by chemokines increases T cell sensitivity to antigen. The current model is that signaling and formation of the immunological synapse are tightly interwoven in mature T cells. We also extend this model to natural killer cell activation, where the inhibitory NK synapse provides a striking example in which inhibition of signaling leaves the synapse in its nascent, inverted state. The APC may also play an active role in immunological synapse formation, particularly for activation of naïve T cells.

A supramolecular basis for CD45 tyrosine phosphatase regulation in sustained T cell activation.

Transmembrane protein tyrosine phosphatases, such as CD45, can act as both positive and negative regulators of cellular signaling. CD45 positively modulates T cell receptor (TCR) signaling by constitutively priming p56lck through the dephosphorylation of the C-terminal negative regulatory phosphotyrosine site. However, CD45 can also exert negative effects on cellular processes, including events triggered by integrin-mediated adhesion. To better understand these opposing actions of tyrosine phosphatases, the subcellular compartmentalization of CD45 was imaged by using laser scanning confocal microscopy during functional TCR signaling of live T lymphocytes. On antigen engagement, CD45 was first excluded from the central region of the interface between the T cell and the antigen-presenting surface where CD45 would inhibit integrin activation. Subsequently, CD45 was recruited back to the center of the contact to an area adjacent to the site of sustained TCR engagement. Thus, CD45 is well positioned within a supramolecular assembly in the vicinity of the engaged TCR, where CD45 would be able to maintain src-kinase activity for the duration of TCR engagement.

Cutting edge: hierarchy of chemokine receptor and TCR signals regulating T cell migration and proliferation.

Chemokines play an important role in establishing the distribution of lymphocyte subpopulations in primary and secondary lymphoid tissues and in the recruitment of leukocytes to sites of inflammation. However, the potential of chemokines to down-regulate immune responses has not been demonstrated. We now show that certain chemokine gradients have the potential to suppress T cell activation by preventing formation of the immunological synapse, the specialized cell-cell junction that forms before a T cell can be fully activated. Our data reveals an immunosuppressive potential of chemokines engaging the CXCR3 and CCR7 receptors, but not the CXCR4, CCR2, CCR4, or CCR5 receptors. These results suggest a novel mechanism for T cell ignorance of agonist MHC-peptide complexes based on dominant chemokine gradients.

Relative contribution of LFA-1 and Mac-1 to neutrophil adhesion and migration.

To differentiate the unique and overlapping functions of LFA-1 and Mac-1, LFA-1-deficient mice were developed by targeted homologous recombination in embryonic stem cells, and neutrophil function was compared in vitro and in vivo with Mac-1-deficient, CD18-deficient, and wild-type mice. LFA-1-deficient mice exhibit leukocytosis but do not develop spontaneous infections, in contrast to CD18-deficient mice. After zymosan-activated serum stimulation, LFA-1-deficient neutrophils demonstrated activation, evidenced by up-regulation of surface Mac-1, but did not show increased adhesion to purified ICAM-1 or endothelial cells, similar to CD18-deficient neutrophils. Adhesion of Mac-1-deficient neutrophils significantly increased with stimulation, although adhesion was lower than for wild-type neutrophils. Evaluation of the strength of adhesion through LFA-1, Mac-1, and CD18 indicated a marked reduction in firm attachment, with increasing shear stress in LFA-1-deficient neutrophils, similar to CD18-deficient neutrophils, and only a modest reduction in Mac-1-deficient neutrophils. Leukocyte influx in a subcutaneous air pouch in response to TNF-alpha was reduced by 67% and 59% in LFA-1- and CD18-deficient mice but increased by 198% in Mac-1-deficient mice. Genetic deficiencies demonstrate that both LFA-1 and Mac-1 contribute to adhesion of neutrophils to endothelial cells and ICAM-1, but adhesion through LFA-1 overshadows the contribution from Mac-1. Neutrophil extravasation in response to TNF-alpha in LFA-1-deficient mice dramatically decreased, whereas neutrophil extravasation in Mac-1-deficient mice markedly increased.

The immunological synapse: a molecular machine controlling T cell activation.

The specialized junction between a T lymphocyte and an antigen-presenting cell, the immunological synapse, consists of a central cluster of T cell receptors surrounded by a ring of adhesion molecules. Immunological synapse formation is now shown to be an active and dynamic mechanism that allows T cells to distinguish potential antigenic ligands. Initially, T cell receptor ligands were engaged in an outermost ring of the nascent synapse. Transport of these complexes into the central cluster was dependent on T cell receptor-ligand interaction kinetics. Finally, formation of a stable central cluster at the heart of the synapse was a determinative event for T cell proliferation.

Antigen receptor engagement delivers a stop signal to migrating T lymphocytes.

We investigated the role of the T cell antigen receptor (TcR) in control of T cell migration in an in vitro system. We used T cells from transgenic mice bearing a TcR for the lysozyme peptide 48-62 bound to I-A(k) (3A9). T cells from the 3A9 TcR transgenic mice crawled on purified intercellular adhesion molecule-1 substrates, but strikingly, stopped upon interaction with the physiological ligand, i.e., the mouse I-A(k) with covalently attached hen egg white lysozyme peptide residues 48-62 complex. TcR-triggered stopping was reversible by treatment with adhesion-strengthening phorbol esters. The microtubule organizing center of stopped cells was positioned adjacent to the site of stable cell anchorage. Direct conversion of lymphocyte function associated-1 to the high-affinity conformation with antibodies also stopped T cells in a similar manner to antigen. Thus, physiological TcR engagement triggers a stop signal through lymphocyte function associated-1. We propose that the stop signal is an early and essential event in T cell activation that also will play an important role in control of T cell migration.