ERM and Rho Signal Pathways in T Cell Immune Senescence

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DESCRIPTION (provided by applicant): Cytoskeletal proteins of the ezrin/radixin/moesin family (ERM proteins) contribute to T cell activation in responses to antigen, and also to T cell polarization in response to connective tissue matrix proteins and chemokine gradients. Previous work has shown that T cells from aged mice are defective in their ability to develop molecular linkages between surface macromolecules and the underlying cytoskeletal framework, both for proteins that move to the synapse and those that are excluded from the site of T cell/APC interaction. T cells from aged mice also show defective cytoskeletal rearrangements and lamellipodia formation when placed in contact with slides coated with antibodies to the TCR/CD3 complex. New data suggest that aged T cells differ from young T cells in several aspects of ERM biochemistry, including ERM phosphorylation and ERM associations with binding proteins, and also show defects in the activation of Rho-family GTPases that control ERM function. Three aims are proposed to clarify, at a molecular level, the basis for age-dependent defects in ERM and Rho-based signals, and their contribution to defective T cell polarization responses. Aim 1 will focus on T cell cytoskeletal polarization in responses to fibronectin signals and to stimulation by a gradient of the chemokine SDF-1a, testing the idea that those T cells from aged mice that fail to respond appropriately to these signals will show corresponding defects in re-localization of ERM proteins, ERM-binding proteins, and ERM-associated surface markers. Aim 2 will evaluate age- dependent change in ERM association with kinases, phosphatases, and surface linker proteins that control ERM position and function during TCR-dependent and TCR-independent polarization responses. This aim will also include membrane fractionation protocols to evaluate age effects on ERM localization with respect to high viscosity membrane microdomains, and studies of Cbp and Cbl-b proteins known to regulate association of ERM with the plasma membrane. Work for Aim 3 will investigate two hypotheses about the age-related defect in RhoA activity, one involving altered Vav signal pathways and the other evaluating the inhibition of RhoA reactivation by members of the GDI family of RhoA dissociation inhibitors. The work proposed should confirm the importance of altered ERM and RhoA pathways in T cell immune senescence, determine which of the plausible control pathways is actually critical to altered T cell responses to connective tissue and chemokine signals, and suggest new lines of experimentation that would merit attention in long term studies. Aging of the immune system contributes to many of the diseases and disabilities of old age, but too little is known about factors that regulate the decline of this system in old age. This research plan is aimed at exploring several ideas about the molecular basis for age-dependent immune failure by studies of the biochemistry of T cell activation in cells from aged mice.