4-1BB Signaling Activates the T Cell Factor 1 Effector/β-Catenin Pathway with Delayed Kinetics via ERK Signaling and Delayed PI3K/AKT Activation to Promote the Proliferation of CD8+ T Cells

4-1BB (CD137), an inducible costimulatory molecule, strongly enhances the proliferation and effector function of CD8+ T cells. Since the serine/threonine kinase, glycogen synthase kinase-3 (GSK-3), is involved in a variety of signaling pathways of cellular proliferation, migration, immune responses, and apoptosis, we examined whether 4-1BB signaling activates GSK-3/β-catenin signaling and downstream transcription factors to enhance the proliferation of CD8+ T cells. 4-1BB signaling induces rapid activation of ERK and IκB degradation, and shows delayed activation of AKT at 24 h post 4-1BB stimulation on anti-CD3 activated T cells. ERK and AKT signals were required for sustained β-catenin levels by inactivating GSK-3, which was also observed with delayed kinetics after 4-1BB stimulation. As a transcriptional partner of β-catenin, 4-1BB signaling decreased levels of FOXO1 and increased levels of stimulatory TCF1 in CD8+ T cells at 2–3 days but not at early time points after 4-1BB engagement. The enhanced proliferation of CD8+ T cells due to 4-1BB signaling was completely abolished by treatment with the TCF1/β-catenin inhibitor quercetin. These results show that 4-1BB signaling enhances the proliferation of activated CD8+ T cells by activating the TCF1/β-catenin axis via the PI3K/AKT/ERK pathway. As effects of 4-1BB on AKT, FOXO1, β-catenin and GSK-3β showed delayed kinetics it is likely that an intervening molecule induced by 4-1BB and ERK signaling in activated T cells is responsible for these effects. These effects were observed on CD8+ but not on CD4+ T cells. Moreover, 4-1BB appeared to be unique among several TNFRs tested in inducing increase in stimulatory over inhibitory TCF-1.