The Parker laboratory is interested in the cell-surface molecules and intracellular signaling pathways which determine whether an encounter between helper T cells and B cells or other antigen presenting cells results in immunity or tolerance. In a simplified model of peripheral tolerance to self, the Parker lab found that a signal through OX40 (CD134) blocks functional anergy in transferred T cells responding to transgenic or allogeneic antigens, drives the T cells to differentiate into cytokine-secreting effector cells, and results in fatal acute graft versus host disease in unirradiated recipient animals. Current work is focused on the role of the alternative pathway of NFkB activation in that model. In a second project, the lab is exploring the possibility that certain subsets of weakly autoreactive B cells play an essential role as antigen-presenting cells in inducing tolerance to self antigens in T cells. In a third project, the lab is exploring the "immunological synapse", the structure that forms in the contact zone between a T cell and an antigen presenting cells. We are examining the role of the synapse in the specific delivery of effector cytokines from T cells to antigen presenting cells, with an emphasis on the membrane-bound TNF family members, CD40L (CD154) and FasL. We are also exploring differences among T cell subsets in the structure of the synapse and the functional consequences of those differences.
Stop and start signals in the immune system
The interactions between T cells and antigen presenting cells can result in an immune response or in immunological tolerance, depending on the state of activation and differentiation of the interacting cells. In my laboratory, we attempt to identify and characterize the cell-surface molecules and intercellular interactions which determine whether an encounter between helper T cells and B cells or other antigen presenting cells results in immunity or tolerance.
Peripheral tolerance in T cells
One aim is to explore a role for resting B cells as antigen presenting cells in the induction of helper T cell tolerance to self and foreign antigens, using mice transgenic for antigen or antigen receptors and deficient in other molecules required for intercellular communication. In a simplified model of peripheral tolerance to self, we found that a signal through OX40 (CD134) blocks functional anergy in transferred T cell antigen receptor transgenic T cells responding to transgenic self antigen, and results in fatal acute graft versus host disease. In collaboration with Phillip Stork in the Vollum Institute at OHSU, we use mice transgenic for intracellular signaling proteins to investigate protein kinase signaling pathways in T cell tolerance.
In collaboration with James Rosenbaum of the Casey Eye Institute at OHSU, we are used intravital fluorescent microscopy in the eye to follow antigen specificity in extravasion over time in inflammatory sites, using adoptive transfer of labeled T cell antigen receptor transgenic T cells.
The immunological synapse
T lymphocytes see antigen only as small peptides bound to MHC molecules on the surface of antigen presenting cells (APC). In addition to the T cell antigen receptor (TCR) and peptide-loaded MHC, dozens of other cell surface molecules play necessary or modulating roles in determining the outcome of the T cell/APC interaction. Antigen recognition is accompanied by large-scale, cytoskeleton-dependent rearrangements of these molecules to form an organized contact interface between the T cell and the APC termed the "immunological synapse".
T cells loaded with a Ca++ sensitive intracellular dye are added to the fibroblasts, on which the peptide-loaded MHC molecules are shown in blue.
We are using video fluorescent microscopy to follow calcium signals and synapse formation between murine TCR transgenic T cells and fibroblasts bearing fluorescent MHC molecules loaded with covalently attached antigenic peptides. T cells loaded with a Ca++ sensitive intracellular dye are added to the fibroblasts, on which the peptide-loaded MHC molecules are shown in blue. When initial antigen recognition occurs, the T cells change color from yellow to green as the intracellular Ca++ concentration suddenly increases. Following antigen recognition, MHC molecules accumulate in the contact region between the T cell and the APC, forming an immunological synapse.
T cell dissociates from the APC and captures MHC:peptide complexes directly from the immunological synapse.
T cells stop when they recognized antigen. If they start moving again, they drag the accumulated MHC molecules with them as they move across the APC surface. If they dissociate from the APC, they take a portion of the accumulated MHC molecules with them. The T cell and associated synapse (peptide-loaded MHC molecules are shown in green) migrate across the APC. The T cell then dissociates from the APC and captures MHC:peptide complexes directly from the immunological synapse. In the minutes following separation, the spot on the APC diminishes significantly while the MHC spot transferred to the T cell remains unchanged. We are focusing on the role of the immunological synapse in delivering the membrane-associated cytokines, CD40L (CD154) and FasL (CD178) from CD4 T cells to APC. These two cytokines are central in the regulatory interactions and fate decisions that determine the balance between an effective immune response and autoimmune disease. We will test the hypothesis that preformed CD40L, like preformed FasL, is delivered rapidly upon antigen recognition by effector and memory CD4 T cells directly to the APC, with important functional consequences distinct from those of newly synthesized CD40L. We will also test the hypothesis that the function of the ring of adhesion molecules in the mature immunological synapse in CD4 T cells is to ensure antigen-specific delivery of cytolytic molecules to the APC and prevent delivery to surrounding cells.