Costimulatory signals lower the threshold for T cell activation, and tune the immune response, driving T cells towards distinct effector lineages and establishing the balance between immunity and tolerance. Because of their central role in shaping the immune response, and because of the conserved nature of costimulatory receptor-ligand interactions, costimulatory molecules are powerful therapeutic targets. Interaction of T cells with pMHC and costimulatory ligands such CD80/86 and ICAM-1 takes place at the immunological synapse. Several pieces of evidence suggest that the DC cytoskeleton plays an important role in modulating signaling events at the immunological synapse, but the molecular basis for this is not understood. We have discovered that while pMHC moves relatively freely on the DC membrane, the DC actin cytoskeleton constrains the mobility of ICAM-1 and CD80, and we have identified two actin-binding proteins, moesin and ?-actinin, that play a key role in this process. We hypothesize that cytoskeletal constraints to mobility of ICAM-1 and CD80 on the DC surface promote costimulatory signaling, by creating tension on receptors and modulating dynamics of signaling microclusters on the T cell side of the synapse. This hypothesis will be tested by carrying out three specific aims. First, we will characterize the interactions of moesin and ?- actinin with basic sequences in the cytoplasmic tails of ICAM-1 and CD80, and test the effects of perturbing these interactions on lateral diffusion of ICAM-1 and CD80. Second, in collaboration with membrane biophysicist Tobias Baumgart, we will use novel mixed mobility surfaces to test the effects of varying ICAM-1 mobility and patterning while maintaining pMHC in a mobile state. Several aspects of the T cell response to these surfaces will be assessed, including specific signaling events, LFA-1 conformational change, and dynamics of the T cell cytoskeleton and associated signaling molecules. Finally, we will engineer DCs in which ICAM-1 mobility is altered, either by perturbing actin-binding proteins or by mutating interacting residue in the ICAM-1 tail, and test the effects on T cell activation and lineage development in vitro. Adoptive transfer experiments will also be performed to test the effects of ICAM-1 mobility on T cell-DC interactions in vivo using intravital imaging. Taken together, these studies will test an unexplored aspect of costimulatory signaling, and will provide important new insights into how the DC actin cytoskeleton modulates the immune response. This information will guide clinical efforts to manipulate DC function in vaccine development, transplantation medicine and treatment of allergy and autoimmune disease.

Public Health Relevance

Activation of T cells by dendritic cells is essential for the initiation of adaptive immunity, and this process is highly dependent on stimulation by costimulatory molecules. This proposal tests an unexplored aspect of this process, the mobility of costimulatory ligands in the DC plasma membrane. The knowledge gained from these studies will be important for the development of effective dendritic cell-based vaccines, and for modulating immune responses in transplant recipients and autoimmune disease patients.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM104867-01
Application #
8431504
Study Section
Cellular and Molecular Immunology - A Study Section (CMIA)
Program Officer
Marino, Pamela
Project Start
2013-07-01
Project End
2017-04-30
Budget Start
2013-07-01
Budget End
2014-04-30
Support Year
1
Fiscal Year
2013
Total Cost
$349,602
Indirect Cost
$131,202
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104