Optimal T Cell Activation and B Cell Antibody Responses In Vivo Require the Interaction between Leukocyte Function-Associated Antigen-1 and Kindlin-3.

Kindlin-3 is an important integrin regulator that is mutated in the rare genetic disorder, leukocyte adhesion deficiency type III, a disorder characterized by defective neutrophil trafficking and platelet function, leading to recurrent bacterial infections and bleeding. Kindlin-3 is also known to regulate T cell adhesion in vitro and trafficking in vivo, but whether the integrin/kindlin interaction regulates T or B cell activation in vivo is unclear. In this study, we used TTT/AAA ß2-integrin knock-in (KI) mice and TCR-transgenic (OT-II) KI mice, in which the integrin/kindlin connection is disrupted, to investigate the role of the integrin/kindlin interaction in T cell activation. We show that basal T cell activation status in these animals in vivo is normal, but they display reduced T cell activation by wild-type Ag-loaded dendritic cells in vitro. In addition, T cell activation in vivo is reduced. We also show that basal Ab levels are normal in TTT/AAA ß2-integrin KI mice, but B cell numbers in lymph nodes and IgG and IgM production after immunization are reduced. In conclusion, we show that the integrin/kindlin interaction is required for trafficking of immune cells, as well as for T cell activation and B cell Ab responses in vivo. These results imply that the immunodeficiency found in leukocyte adhesion deficiency type III patients, in addition to being caused by defects in neutrophil function, may be due, in part, to defects in lymphocyte trafficking and activation.

Loss of beta2-integrin-mediated cytoskeletal linkage reprogrammes dendritic cells to a mature migratory phenotype.

The actin cytoskeleton has been reported to restrict signalling in resting immune cells. Beta2-integrins, which mediate adhesion and cytoskeletal organization, are emerging as negative regulators of myeloid cell-mediated immune responses, but the molecular mechanisms involved are poorly understood. Here, we show that loss of the interaction between beta2-integrins and kindlin-3 abolishes the actin-linkage of integrins and the GM-CSF receptor in dendritic cells. This leads to increased GM-CSF receptor/Syk signalling, and to the induction of a transcriptional programme characteristic of mature, migratory dendritic cells, accumulation of migratory dendritic cells in lymphoid organs, and increased Th1 immune responses in vivo. We observe increased GM-CSF responses and increased survival in neutrophils where the interaction between integrin and the cytoskeleton is disrupted. Thus, ligand-reinforced beta2-integrin tail interactions restrict cytokine receptor signalling, survival, maturation and migration in myeloid cells and thereby contribute to immune homeostasis in vivo.

The ß2 integrin-kindlin-3 interaction is essential for T-cell homing but dispensable for T-cell activation in vivo.

Kindlin-3 is mutated in the rare genetic disorder, leukocyte adhesion deficiency type III, which is characterized by deficient integrin-mediated adhesion of leukocytes and platelets. However, the specific roles of kindlin-3-ß2-integrin interactions in T-cell adhesion and homing and immune responses in vivo remain unclear. Here, we show that the TTT motif in ß2 integrins controls kindlin-3 binding. Mutation of the kindlin-3 binding site in ß2 integrins caused a loss of firm adhesion of T cells under both static and shear flow conditions and a reduction of T-cell homing to lymph nodes in vivo. However, atomic force microscopy studies of integrin-ligand bonds revealed that initial ligand binding could still occur, and 2-dimensional T-cell migration was reduced but not abolished by the TTT/AAA mutation in the ß2 integrin. Importantly, dendritic cell-mediated T-cell activation in vivo was normal in TTT/AAA ß2 integrin knock-in mice. Our results reveal a selective role of the kindlin-3-integrin association for lymphocyte functions in vivo; the integrin-kindlin-3 interaction is particularly important in adhesion strengthening under shear flow, and for T-cell homing to lymph nodes, but dispensable for T cell activation which occurs in a shear-free environment.