Centrosome docking at the immunological synapse is controlled by Lck signaling.

Docking of the centrosome at the plasma membrane directs lytic granules to the immunological synapse. To identify signals controlling centrosome docking at the synapse, we have studied cytotoxic T lymphocytes (CTLs) in which expression of the T cell receptor-activated tyrosine kinase Lck is ablated. In the absence of Lck, the centrosome is able to translocate around the nucleus toward the immunological synapse but is unable to dock at the plasma membrane. Lytic granules fail to polarize and release their contents, and target cells are not killed. In CTLs deficient in both Lck and the related tyrosine kinase Fyn, centrosome translocation is impaired, and the centrosome remains on the distal side of the nucleus relative to the synapse. These results show that repositioning of the centrosome in CTLs involves at least two distinct steps, with Lck signaling required for the centrosome to dock at the plasma membrane.

Reduced functional avidity promotes central and effector memory CD4 T cell responses to tumor-associated antigens.

The effect of TCR signals on the differentiation of memory T cells is poorly defined. Conventional wisdom suggests that high-avidity interactions are best for the selection of vaccine Ag candidates or T cell specificities for adoptive T cell therapy to stimulate robust responses. However, in conditions of Ag persistence, high-avidity clones might exhaust and fail to form long-lived protective memory. We have manipulated the functional avidity of CD4 T cells by reducing expression of Lck, a key kinase involved in TCR triggering. Using a mouse model, we followed tetramer-positive T cells responding to a tumor Ag expressed by an adenocarcinoma. We show that reducing the functional avidity increased effector-effector memory responses and improved the generation of self-renewing, recirculating, tumor Ag-specific memory phenotype CD4 T cells. Moreover, such cells together with wild type CD8 T cells were better able to control tumor growth. Mechanistically, reducing Lck prolonged IL-2 production and cell turnover in the central memory population while reducing expression of exhaustion markers in the face of chronic Ag. Our data indicate that, in situations of persistent Ag challenge, generating T cells with reduced functional avidity may elicit more effective immune responses.

Fyn regulates the duration of TCR engagement needed for commitment to effector function.

In naive T cells, engagement of the TCR with agonist peptide:MHC molecules leads to phosphorylation of key intracellular signaling intermediates within seconds and this peaks within minutes. However, the cell does not commit to proliferation and IL-2 cytokine production unless receptor contact is sustained for several hours. The biochemical basis for this transition to full activation may underlie how T cells receive survival signals while maintaining tolerance, and is currently not well understood. We show here that for CD8 T cells commitment to proliferation and cytokine production requires sustained activation of the Src family kinase Lck and is opposed by the action of Fyn. Thus, in the absence of Fyn, commitment to activation occurs more rapidly, the cells produce more IL-2, and undergo more rounds of division. Our data demonstrate a role for Fyn in modulating the response to Ag in primary T cells.