Shannon K. Bromley

A Novel Adaptor Protein Orchestrates Receptor Patterning and Cytoskeletal Polarity in T-Cell Contacts

Recognition of antigen by T cells requires the formation of a specialized junction between the T cell and the antigen-presenting cell. This junction is generated by the recruitment and the exclusion of specific proteins from the contact area. The mechanisms that regulate these events are unknown. Here we demonstrate that ligand engagement of the adhesion molecule, CD2, initiates a process of protein segregation, CD2 clustering, and cytoskeletal polarization. Although protein segregation was not dependent on the cytoplasmic domain of CD2, CD2 clustering and cytoskeletal polarization required an interaction of the CD2 cytoplasmic domain with a novel SH3-containing protein. This novel protein, called CD2AP, is likely to facilitate receptor patterning in the contact area by linking specific adhesion receptors to the cytoskeleton.

Orchestrating the orchestrators: chemokines in control of T cell traffic

The understanding of how chemokines orchestrate the trafficking and activity of immune cells has increased considerably. So far, over 50 chemokines and 20 chemokine receptors have been identified. Detailed analyses have demonstrated the function of chemokine receptors on T cell subsets, the temporal and spatial expression patterns of chemokines in vivo and the phenotypes of animals genetically deficient in one component or several components of the chemokine-chemokine receptor system. New microscopy modalities for studying the influence of chemokines on the migratory activity of T cells in the lymph node have also brought new insights. Here we review such advances with particular emphasis on control of the migration of T cell subsets in lymph nodes and in peripheral tissues in homeostasis and inflammation.

Chemokine receptor CCR7 guides T cell exit from peripheral tissues and entry into afferent lymphatics

T cell circulation between peripheral tissues and the lymphoid compartment is critical for immunosurveillance and host defense. However, the factors that determine whether T cells remain in peripheral tissue or return to the circulation are undefined. Here we demonstrate that the chemokine receptor CCR7 is a critical signal that determines T cell exit from peripheral tissue. Both CCR7− and CCR7+ effector T cells entered mouse asthmatic lung and while CCR7− T cells accumulated, CCR7+ T cells continued to migrate into afferent lymph. Delivery of both CCR7+ and CCR7− T cells directly into the airways showed that only CCR7+ T cells exited the lung and entered draining lymph nodes. Our study establishes a molecular basis for T cell exit from peripheral tissues.

Leukotriene B4 receptor BLT1 mediates early effector T cell recruitment

Leukotriene B4 (LTB4) was originally described as a potent lipid myeloid cell chemoattractant, rapidly generated from innate immune cells, that activates leukocytes through the G protein–coupled receptor BLT1. We report here that BLT1 is expressed on effector CD4+ T cells generated in vitro as well as in vivo when effector T cells migrate out of the lymphoid compartment and are recruited into peripheral tissues. BLT1 mediated LTB4-induced T helper type 1 (TH1) and TH2 cell chemotaxis and firm adhesion to endothelial cells under flow, as well as early CD4+ and CD8+ T cell recruitment into the airway in an asthma model. Our findings show that the LTB4-BLT1 pathway is involved in linking early immune system activation and early effector T cell recruitment.

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.

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.

CCR4-dependent regulatory T cell function in inflammatory bowel disease

Inflammatory bowel disease (IBD) is an idiopathic inflammatory disease of the intestine. CD4+ T lymphocytes play an important role in both initiating and regulating intestinal inflammatory immune responses. CD4+CD25+CD45RBlow regulatory T (T reg) cells are capable of preventing the development of colitis in a mouse model of IBD. The precise mechanism of T reg cell–mediated prevention of colitis in this model is unclear, and the role of chemokine receptors in the trafficking and function of T reg cells in this model has not been determined. We examined the role of the chemokine receptor CCR4 in in vivo trafficking and suppressive function of T reg cells in a mouse adoptive transfer model of IBD. CCR4-deficient T reg cells failed to accumulate in the mesenteric lymph nodes (MLNs) at early time points (2–5 d) after adoptive transfer, resulting in a failure to suppress the generation of pathogenic T cells and the development of colitis. Moreover, although CCR4-deficent T cells had equivalent in vitro suppressive activity and accumulated in MLNs at later time points (42–56 d), they were unable to suppress colitis. Our study demonstrates that CCR4 plays an important role in T reg cell trafficking in LNs and that this is critical for T reg cell suppressive function in vivo.

Recirculating Memory T Cells Are a Unique Subset of CD4+ T Cells with a Distinct Phenotype and Migratory Pattern

Several populations of memory T cells have been described that differ in their migration and function. In this study, we have identified a unique subset of memory T cells, which we have named recirculating memory T cells (TRCM). By exposing Kaede transgenic mouse skin to violet light, we tracked the fate of cutaneous T cells. One population of memory CD4+ T cells remained in the skin. A second population migrated from the skin into draining lymph nodes (LNs) in a CCR7-dependent manner. These migrating CD4+ T cells expressed a novel cell surface phenotype (CCR7int/+CD62LintCD69−CD103+/− E-selectin ligands+) that is distinct from memory T cell subsets described to date. Unlike memory T cell subsets that remain resident within tissues long-term, or that migrate either exclusively between lymphoid tissues or into peripheral nonlymphoid sites, CD4+ TRCM migrate from the skin into draining LNs. From the draining LNs, CD4+ TRCM reenter into the circulation, distal LNs, and sites of non-specific cutaneous inflammation. In addition, CD4+ TRCM upregulated CD40L and secreted IL-2 following polyclonal stimulation. Our results identify a novel subset of recirculating memory CD4+ T cells equipped to deliver help to both distal lymphoid and cutaneous tissues.

Stimulation of naive T-cell adhesion and immunological synapse formation by chemokine-dependent and -independent mechanisms

Chemokines adsorbed to the cell surface play an important role in the initial interactions of T cells with endothelial cells, and may also have a role in T‐cell interactions with dendritic cells. Therefore, we examined the effect of surface‐adsorbed chemokines on the interaction of naïve murine splenic T cells with supported bilayers containing intercellular adhesion molecule (ICAM)‐1, or with bone marrow‐derived cultured dendritic cells in the presence and absence of relevant MHC–peptide complexes. Naïve T cells formed immunological synapses, defined as a ring of lymphocyte function associated (LFA)‐1–ICAM‐1 interactions surrounding a central cluster of MHC–peptide complexes, on supported planar bilayers containing ICAM‐1 and relevant MHC–peptide complexes. Chemokines stimulated an increase in the percentage of naïve cells that adhered to ICAM‐1, but did not increase the average number of LFA‐1–ICAM‐1 interactions in the contact area. In contrast, relevant MHC–peptide complexes resulted in a small increase in the proportion of interacting T cells, but stimulated an 8‐fold increase in the number of LFA‐1–ICAM‐1 interactions in each contact formed. Naïve T cells displayed a significant basal adhesion to bone marrow dendritic cells that was further increased when relevant chemokines were adsorbed to the dendritic cell surface. However, basal and antigen‐stimulated T‐cell adhesion to dendritic cells was not sensitive to pertussis toxin. Thus, there are chemokine‐independent mechanisms that initiate adhesion between T cells and dendritic cells.

Polar redistribution of the sialoglycoprotein CD43: implications for T cell function.

Contact between T cells and APCs results in the orchestrated segregation of molecules at the cell-cell interface and formation of a specialized structure termed the immunological synapse. This model predicts the topological seclusion of large molecules such as CD43 from the site of closest contact between the T cell and APC, allowing for the close apposition of cell membranes and effective TCR engagement. Similarly, during T cell migration segregation of CD43 to the uropod is thought to aid integrin adhesion at the leading edge of the cell by removing steric hindrance. We show in this work that CD43 distribution on T cells is regulated by a membrane proximal ezrin binding site and that failure to displace CD43 from the immunological synapse has no inhibitory effects on primary T cell activation. We also report that CD43 expression at the contact zone between T cells and matrix does not negatively regulate motility but may regulate LFA-1 de-adhesion. These results suggest that the steric barrier model of CD43 is inadequate and that alternative mechanisms account for the negative regulatory properties of CD43.