Negative Regulation of Memory Phenotype CD8 T Cell Conversion by Adhesion and Degranulation–Promoting Adapter Protein

Abstract

The maintenance of T cell repertoire diversity involves the entry of newly developed T cells, as well as the maintenance of memory T cells generated from previous infections. This balance depends on competition for a limited amount of homeostatic cytokines and interaction with self-peptide MHC-I. In the absence of prior infection, memory-like or memory phenotype (MP) CD8 T cells can arise from homeostatic cytokine exposure during neonatal lymphopenia. Aside from downstream cytokine signaling, little is known about the regulation of the conversion of naïve CD8 T cells to MP CD8 T cells during acute lymphopenia. We have identified a novel negative regulatory role for the adapter protein ADAP in CD8 T cell function. We show that in the absence of ADAP, naïve CD8 T cells exhibit a diminished response to stimulatory Ag, but an enhanced response to weak agonist altered peptide ligands. ADAP-deficient mice exhibit an increased number of MP CD8 T cells that occurs following thymic emigration and is largely T cell intrinsic. Naïve ADAP-deficient CD8 T cells are hyperresponsive to lymphopenia in vivo and exhibit enhanced activation of STAT5 and homeostatic antigen-independent proliferation in response to IL-15. Our results indicate that ADAP dampens naïve CD8 T cell responses to lymphopenia and IL-15, and demonstrates a novel antigen-independent function for ADAP in the suppression of MP CD8 T cell generation.

Adhesion- and Degranulation-Promoting Adapter Protein Promotes CD8 T Cell Differentiation and Resident Memory Formation and Function during an Acute Infection

“…Although we and others have demonstrated defects in T-APC contacts, entry of T cells into the cell cycle, proliferation, differentiation and survival in the absence of ADAP (13–17, 19, 21), several recent reports have documented a negative regulatory function for ADAP in T cells (22, 23). We have recently shown that ADAP-deficient T cells have an enhanced response to weak agonist peptide ligands in naïve CD8 T cells (22).…”

“…Ex vivo conjugate assays were performed by modifying our previously described conjugate assay (14, 15, 18, 21, 22), using splenic dendritic cells enriched from mice treated for 14 days with a B16 tumor expressing FLT3 ligand (33). Briefly, single cell suspensions obtained from spleens of FLT3 conditioned mice, which contained 25–35% CD11c + MHCII + dendritic cells, were labeled with Cell Tracker Orange (CTO, Thermo Fisher) as previously described (14, 15, 18, 21, 22) and then pulsed with the indicated doses of SIINFEKL peptide for 30 mins at 37C in warm TCPM, washed, and resuspended at 5 x 10 6 cells/ml in pre-warmed TCPM.…”

“…Briefly, single cell suspensions obtained from spleens of FLT3 conditioned mice, which contained 25–35% CD11c + MHCII + dendritic cells, were labeled with Cell Tracker Orange (CTO, Thermo Fisher) as previously described (14, 15, 18, 21, 22) and then pulsed with the indicated doses of SIINFEKL peptide for 30 mins at 37C in warm TCPM, washed, and resuspended at 5 x 10 6 cells/ml in pre-warmed TCPM. 100 μl (0.5 x 10 6 ) of peptide-pulsed splenocytes were added to triplicate wells of a 96-well round-bottom plate, followed by the addition of 100 μl of pre-warmed unlabeled lymphocytes isolated from the spleens, SG, or FRT of VSV-OVA infected mice isolated as described above.…”

“…We have recently shown that ADAP-deficient T cells have an enhanced response to weak agonist peptide ligands in naïve CD8 T cells (22). In addition, uninfected ADAP-deficient mice have an enhanced frequency and number of CD8 T cells with a memory phenotype that is associated with enhanced responsiveness of ADAP-deficient CD8 T cells to the homeostatic cytokine IL-15 (22).…”

“…We have recently shown that ADAP-deficient T cells have an enhanced response to weak agonist peptide ligands in naïve CD8 T cells (22). In addition, uninfected ADAP-deficient mice have an enhanced frequency and number of CD8 T cells with a memory phenotype that is associated with enhanced responsiveness of ADAP-deficient CD8 T cells to the homeostatic cytokine IL-15 (22). Additional recent reports indicate that ADAP −/ − CTLs show enhanced cytolytic function both in vitro and in vivo (23), although CD8 CTLs lacking ADAP exhibit normal cytotoxicity in response to an allogeneic graft (24, 25).…”