Abstract
The trafficking of T-cells through peripheral tissues and into afferent lymphatic vessels is essential for immune surveillance and an adaptive immune response. Glycogen synthase kinase 3β (GSK3β) is a serine/threonine kinase and regulates numerous cell/tissue-specific functions, including cell survival, metabolism, and differentiation. Here, we report a crucial involvement of GSK3β in T-cell motility. Inhibition of GSK3β by CHIR-99021 or siRNA-mediated knockdown augmented the migratory behavior of human T-lymphocytes stimulated via an engagement of the T-cell integrin LFA-1 with its ligand ICAM-1. Proteomics and protein network analysis revealed ongoing interactions among GSK3β, the surface receptor Notch1 and the cytoskeletal regulator CRMP2. LFA-1 stimulation in T-cells reduced Notch1-dependent GSK3β activity by inducing phosphorylation at Ser9 and its nuclear translocation accompanied by the cleaved Notch1 intracellular domain and decreased GSK3β-CRMP2 association. LFA-1-induced or pharmacologic inhibition of GSK3β in T-cells diminished CRMP2 phosphorylation at Thr514. Although substantial amounts of CRMP2 were localized to the microtubule-organizing center in resting T-cells, this colocalization of CRMP2 was lost following LFA-1 stimulation. Moreover, the migratory advantage conferred by GSK3β inhibition in T-cells by CHIR-99021 was lost when CRMP2 expression was knocked-down by siRNA-induced gene silencing. We therefore conclude that GSK3β controls T-cell motility through interactions with CRMP2 and Notch1, which has important implications in adaptive immunity, T-cell mediated diseases and LFA-1-targeted therapies.
Highlights
In a healthy human under physiological conditions, T-lymphocytes continuously recirculate between the peripheral lymphoid tissues via the blood and lymphatic systems to perform an active immune surveillance as well as mount an adaptive immune response
We have previously reported that lymphocyte function-associated antigen-1 (LFA-1)/intercellular adhesion molecule-1 (ICAM-1) ligation in human peripheral blood lymphocyte (PBL) T-cells promotes Th1 polarization through a Glycogen synthase kinase 3b (GSK3b)-dependent pathway [9]
The current study demonstrates a crucial involvement of GSK3b in T-cell migration, which is important for T-cells to respond to environmental cues, such as chemokines, in order to mount an effective immune response
Summary
In a healthy human under physiological conditions, T-lymphocytes continuously recirculate between the peripheral lymphoid tissues via the blood and lymphatic systems to perform an active immune surveillance as well as mount an adaptive immune response. The T-cell aLb2 integrin lymphocyte function-associated antigen-1 (LFA-1; CD11a/CD18) binds to its ligand intercellular adhesion molecule-1 (ICAM-1) expressed on the endothelium, and this adhesive interaction is crucial for T-cell migration and effector functions [3]. The molecular machinery and the downstream pathways triggered by LFA-1 attachment to the ICAM-1 facilitating T-cell motility remain unclear. The glycogen synthase kinase 3 (GSK3) is a ubiquitous constitutively active serine/threonine kinase that exists in two isoforms, GSK3a and GSK3b, and targets over hundred proteins to regulate context-specific cellular functions [4]. Uncovered as a key enzyme involved in glycogen synthesis, GSK3b is known to regulate cell cycle, development, survival, metabolism, and inflammation in multiple cell types [5,6,7,8]. GSK3b involvement in T-cell motility is yet to be fully understood
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