The effect of enforced Notch signaling on TCR beta, positive, and negative selection of developing T cells

The Notch receptors and signaling cascade represent evolutionarily conserved developmental pathways that mediate cell fate decisions through cell-cell interactions. The role of the Notch pathways during thymocyte development has been controversial. While clearly indicated in T cell versus B cell lineage commitment, the physiological relevance of Notch1 has been less established during further stages of T cell development in the thymus. Using staining protocols for surface Notch1 and downstream targets, we assayed for physiologically relevant Notch1 expression and activation in mouse thymocytes. Surface Notch1 is highly expressed and activated in double negative (DN) thymocytes subsets and Notch1 expression is downregulated as thymocytes mature into the double positive (DP) then single positive (SP) thymocytes, suggesting that other Notch family members or downstream Notch activation targets are important at later stages of development, such as during the DP to SP transition. We investigated the role of the Notch pathways during the DN to DP transition and the DP to SP transition by expressing a constitutively active Notch1 under a thymic specific promoter. Enforced Notch1 signaling promotes the proliferative expansion after the β-selection checkpoint during the DN to DP transition. This may provide some insight into the cellular mechanisms by which Notch1 promotes T cell acute lymphoblastic leukemia (T-ALL). During the DP to SP transition enforced activation of the Notch pathway is able promote the development of SP thymocytes by augmenting positive selection rather than affecting negative selection. These data suggest that Notch pathways do not modulate TCR signals to dictate survival and may play a role in maturation of thymocytes after they have been positively selected.