Abstract
Protein kinase C θ (PKCθ) is expressed in T lymphocytes, in which it plays a crucial role in cell activation, proliferation, and differentiation. Previously, we discovered that the Thr335-Pro motif within the PKCθ-V3 regulatory domain serves as a critical site for PKCθ activation and demonstrated that phosphorylation of the Thr335-Pro motif creates a binding site for the peptidyl-prolyl cis-trans isomerase (PPIase), Pin1. Herein, we elaborate on the functional consequences of the Pin1-PKCθ association and identify Pin1 as a regulator of PKCθ catalytic activity. Pin1 downregulated the activity of PKCθ in vitro in PMA-stimulated human Jurkat T cells and C57BL/6J mouse spleen- and thymus-derived T lymphocytes, an effect that was reversed by juglone, a Pin1-activity inhibitor. Utilizing Pin1 isomerase-deficient mutants, we demonstrate that the functionality of the Pin1 catalytic domain is essential for its regulation of PKCθ, suggesting that Pin1 mediates its inhibitory effect on PKCθ via cis-trans isomerization. In silico docking analysis supported the role of critical residues within the Pin1-PPIase domain that enables the cis-trans interconversion of the PKCθ phospho-Thr335-Pro motif. Retroviral knockdown of Pin1 in Jurkat T cells led to an elevation in PKCθ kinase activity. Furthermore, stimulation of [Lckcre × Pin1lox] F1 mice-derived Pin1-deficient T cells augmented the phosphorylation of SPAK kinase, a bona fide PKCθ downstream substrate. Together, our results support a role for the Pin1 isomerase as a regulator of PKCθ activity and highlight the potential contribution of Pin1 to the fine-tuning of the T cell activation dynamics.