T lymphocytes lacking the lymph node-homing receptors L-selectin and CCR7 do not migrate to lymph nodes in the steady state. Instead, we found here that lymph nodes draining sites of mature dendritic cells or adjuvant inoculation recruited L-selectin-negative CCR7- effector and memory CD8+ T cells. This recruitment required CXCR3 expression on T cells and occurred through high endothelial venules in concert with lumenal expression of the CXCR3 ligand CXCL9. In reactive lymph nodes, recruited T cells established stable interactions with and killed antigen-bearing dendritic cells, limiting the ability of these dendritic cells to activate naive CD4+ and CD8+ T cells. The inducible recruitment of blood-borne effector and memory T cells to lymph nodes may represent a mechanism for terminating primary and limiting secondary immune responses.

Chemokines coordinate leukocyte trafficking by promoting oligomerization and signaling by G protein-coupled receptors; however, it is not known which amino acid residues of the receptors participate in this process. Bioinformatic analysis predicted that Ile52 in transmembrane region-1 (TM1) and Val150 in TM4 of the chemokine receptor CCR5 are key residues in the interaction surface between CCR5 molecules. Mutation of these residues generated nonfunctional receptors that could not dimerize or trigger signaling. In vitro and in vivo studies in human cell lines and primary T cells showed that synthetic peptides containing these residues blocked responses induced by the CCR5 ligand CCL5. Fluorescence resonance energy transfer showed the presence of preformed, ligand-stabilized chemokine receptor oligomers. This is the first description of the residues involved in chemokine receptor dimerization, and indicates a potential target for the modification of chemokine responses.

Recent observations using multiphoton intravital microscopy (MP-IVM) have uncovered an unexpectedly high lymphocyte motility within peripheral lymph nodes (PLNs). Lymphocyte-expressed intracellular signaling molecules governing interstitial movement remain largely unknown. Here, we used MP-IVM of murine PLNs to examine interstitial motility of lymphocytes lacking the Rac guanine exchange factor DOCK2 and phosphoinositide-3-kinase (PI3K)γ, signaling molecules that act downstream of G protein–coupled receptors, including chemokine receptors (CKRs). T and B cells lacking DOCK2 alone or DOCK2 and PI3Kγ displayed markedly reduced motility inside T cell area and B cell follicle, respectively. Lack of PI3Kγ alone had no effect on migration velocity but resulted in increased turning angles of T cells. As lymphocyte egress from PLNs requires the sphingosine-1-phosphate (S1P) receptor 1, a Gαi protein–coupled receptor similar to CKR, we further analyzed whether DOCK2 and PI3Kγ contributed to S1P-triggered signaling events. S1P-induced cell migration was significantly reduced in T and B cells lacking DOCK2, whereas T cell–expressed PI3Kγ contributed to F-actin polymerization and protein kinase B phosphorylation but not migration. These findings correlated with delayed lymphocyte egress from PLNs in the absence of DOCK2 but not PI3Kγ, and a markedly reduced cell motility of DOCK2-deficient T cells in close proximity to efferent lymphatic vessels. In summary, our data support a central role for DOCK2, and to a lesser extent T cell–expressed PI3Kγ, for signal transduction during interstitial lymphocyte migration and S1P-mediated egress.

The JAK/STAT (Janus kinase / signaling transducer and activator of transcription) signaling pathway is implicated in converting stationary epithelial cells to migratory cells. In mammals, migratory responses are activated by chemoattractant proteins, including chemokines. We found that by binding to seven-transmembrane G-protein-coupled receptors, chemokines activate the JAK/STAT pathway to trigger chemotactic responses. We show that chemokinemediated JAK/STAT activation is critical for G-protein induction and for phospholipase C-g dependent Ca 2+ flux; in addition, pharmacological inhibition of JAK or mutation of the JAK kinase domain causes defects in both responses. Furthermore, G § i association with the receptor is dependent on JAK activation, and the chemokine-mediated Ca 2+ flux that requires phospholipase C-g activity takes place downstream of JAK kinases. The chemokines thus employ a mechanism that links heterologous signaling pathways -G proteins and tyrosine kinases -in a network that may be essential for mediating their pleiotropic responses.