The roles of ERK pathway localization and the regulation of [beta]-catenin/Bcl-XL kinetic in thymic selection

Abstract

Thymocytes undergo a process of development that serves to favor the differentiation of T-cells bearing self-MHC (major histo-compatibility complex) restricted T-cell receptors (TCR). Signaling through the TCR is crucial for the development from double positive (CD4+CD8+, referred to as DP) thymocytes into mature T-cells. Strong or no affinity interaction between MHC and the TCR leads to apoptosis and is considered as negative selection or death by neglect respectively. A weak to moderate affinity signal initiates differentiation of DP cells into single positive cells (positive for either CD4 or CD8, SP) termed positive selection. The importance of several pathways for either negative or positive selection has been described. However, the molecular events that allow the differentiation of a positive versus negative selecting signal are unclear because, engagement of the TCR leads to activation of all pathways known to play a role in selection. It has been suggested that differential localization of the extra cellular regulated kinase 1/2 (ERK) pathway plays a key role in selection. Here we examine the localization of ERK activation in thymic selection by the manipulating calcium levels leading alterations in RasGRP location and through a Raf construct targeted to either the plasma membrane or the golgi. We also examine differences in [beta]-catenin expression and its role in the expression of Bcl-XL, a pro-survival molecule critical for survival at the DP stage. We find that activation of the ERK pathway from the plasma membrane converts positive selection to negative selection, while the activation of ERK from the golgi upon stimulation by a negative selector causes a deviation of these cells from apoptosis into the CD8[alpha][alpha]+ lineage. In addition we show that the kinetics of [beta]-catenin and Bcl-XL are initially consistent with a role in mediating survival. However, later in selection [beta]-catenin may be more important in preventing cells that have been negatively signaled from maturing further, while Bcl-XL levels remain low, leading to their eventual death.