Adhesion shapes T cells for prompt and sustained T-cell receptor signalling.

During T-cell migration, cell polarity is orchestrated by chemokine receptors and adhesion molecules and involves the functional redistribution of molecules and organelles towards specific cell compartments. In contrast, it is generally believed that the cell polarity established when T cells meet antigen-presenting cells (APCs) is controlled by the triggered T-cell receptor (TCR). Here, we show that, during activation of human T lymphocytes by APCs, chemokines and LFA-1 establish cell polarity independently of TCR triggering. Chemokine-induced LFA-1 activation results in fast recruitment of MTOC and mitochondria towards the potential APC, a process required to amplify TCR Ca(2+) signalling at the upcoming immunological synapse, to promote nuclear translocation of transcriptional factor NFATc2 and boost CD25 expression. Our data show that the initial adhesive signals delivered by chemokines and LFA-1 shape and prepare T cells for antigen recognition.

CD28 interaction with filamin-A controls lipid raft accumulation at the T-cell immunological synapse.

During physiological T-cell stimulation by antigen presenting cells (APCs), a major T-cell membrane rearrangement is known to occur leading to the organization of 'supramolecular activation clusters' at the immunological synapse. A possible role for the synapse is the generation of membrane compartments where signalling may be organized and propagated. Thus, engagement of the costimulatory molecule CD28 at the immunological synapse promotes the organization of a signalling compartment by inducing cytoskeletal changes and lipid raft accumulation. We identified the actin-binding protein Filamin-A (FLNa) as a novel molecular partner of CD28. We found that, after physiological stimulation, CD28 associated with and recruited FLNa into the immunological synapse, where FLNa organized CD28 signalling. FLNa knockdown by short interfering RNA (siRNA) inhibited CD28-mediated raft accumulation at the immunological synapse and T-cell costimulation. Together, our data indicate that CD28 binding to FLNa is required to induce the T-cell cytoskeletal rearrangements leading to recruitment of lipid microdomains and signalling mediators into the immunological synapse.

Signaling amplification at the immunological synapse.

The immunological synapse is a dynamic structure, formed between a T cell and one or more antigen-presenting cells, characterized by lipid and protein segregation, signaling compartmentalization, and bidirectional information exchange through soluble and membrane-bound transmitters. In addition, the immunological synapse is the site where signals delivered by the T-cell receptors, adhesion molecules, as well as costimulatory and coinhibitory receptors are decoded and integrated. Signaling modulation and tunable activation thresholds allow T cells to interpret the context in which the antigen is presented, recognize infectious stimuli, and finally decide between activation and tolerance. In this review, we discuss some strategies used by membrane receptors to tune activation signals in T cells.

CXCR4-CCR5: a couple modulating T cell functions.

Chemokines and their receptors direct leukocyte migration among blood, lymph and tissues. Evidence has recently accumulated that, besides their chemotactic functions, chemokine receptors are highly versatile players that fine tune immune responses. During human T cell activation by antigen-presenting cells, the chemokine receptors CCR5 and CXCR4 are recruited into the immunological synapse, where they deliver costimulatory signals. However, the molecular mechanisms allowing signaling versatility of chemokine receptors are unknown. Here, we describe the functional interaction between CXCR4 and CCR5 to exert specific biological functions and modulate T lymphocyte responses. We demonstrate that simultaneous expression and cooperation between CCR5 and CXCR4 are required for chemokine-induced T cell costimulation at the immunological synapse. In addition, we provide evidence for a physical association of the two receptors in a signaling complex that activates distinct T cell functions. We suggest that cooperation between receptors represents one key strategy for the functional plasticity of chemokines.

Chemokines: coded messages for T-cell missions.

Chemokines and their receptors control leukocyte migration and homing throughout the body in both physiological and pathological conditions. In the context of the adaptive immune system, which requires high efficiency and control, chemokines and chemokine receptors represent a versatile code that orchestrates the "who, where and when" of the immune response by providing the spatio-temporal guidance for T-cell development, priming and effector functions. In addition to their chemotactic properties, chemokines can directly modulate T-cell responses by amplifying signals at the immune synapse and tuning Th1/Th2 polarization. In this review we will discuss the role of chemokines in T-cell biology, following an ideal pilgrimage that spans the key steps of the T-cell life.

Functional single-cell analysis of T-cell activation by supported lipid bilayer-tethered ligands on arrays of nanowells.

Supported lipid bilayers are an important biomolecular tool for characterizing immunological synapses. Immobilized bilayers presenting tethered ligands on planar substrates have yielded both spatio-temporal and structural insights into how T cell receptors (TCRs) reorganize during the initial formation of synapses upon recognition of peptide antigens bound to major histocompatibility complex (MHC) molecules. The prototypical configuration of these assays, however, limits the extent to which the kinetics and structure of the supramolecular activation clusters of the synapse (that occur in seconds or minutes) can be related to subsequent complex cellular responses, such as cytokine secretion and proliferation, occurring over hours to days. Here we describe a new method that allows correlative measures of both attributes with single-cell resolution by using immobilized lipid bilayers and tethered ligands on the surface of dense arrays of subnanoliter wells. This modification allows each nanowell to function as an artificial antigen-presenting cell (APC), and the synapses formed upon contact can be imaged by fluorescence microscopy. We show that the lipid bilayers remain stable and mobile on the surface of the PDMS, and that modifying the ligands tethered to the bilayer alters the structure of the resulting synapses in expected ways. Finally, we demonstrate that this approach allows the subsequent characterization of secreted cytokines from the activated human T cell clones by microengraving in both antigen- and pan-specific manners. This new technique should allow detailed investigations on how biophysical and structural aspects of the synapse influence the activation of individual T cells and their complex functional responses.

T cells and their partners: The chemokine dating agency.

Chemokines and their receptors have long been recognized as key molecules directing leukocyte migration between blood, lymph and tissues. Evidence accumulated in recent years indicates that, in addition to their chemotactic functions, chemokine receptors are highly versatile players fine-tuning immune responses. Chemokine receptors and ligands have been implicated in dendritic-cell maturation, signal transmission at the immunological synapse between T lymphocytes and their cellular partners, and in the polarization of immune responses. These findings identify new roles for chemokines in T-cell triggering and activation of effector functions, and suggest that these small cytokines represent key conductors of adaptive immunity.