Protein Phosphatase 2A Regulates Interleukin-2 Receptor Complex Formation and JAK3/STAT5 Activation

Abstract

Reversible protein phosphorylation plays a key role in interleukin-2 (IL-2) receptor-mediated activation of Janus tyrosine kinase 3 (JAK3) and signal transducer and activator of transcription 5 (STAT5) in lymphocytes. Although the mechanisms governing IL-2-induced tyrosine phosphorylation and activation of JAK3/STAT5 have been extensively studied, the role of serine/threonine phosphorylation in controlling these effectors remains to be elucidated. Using phosphoamino acid analysis, JAK3 and STAT5 were determined to be serine and tyrosinephosphorylated in response to IL-2 stimulation of the human natural killer-like cell line, YT. IL-2 stimulation also induced serine/threonine phosphorylation of IL-2R␤, but not IL-2R␥. To investigate the regulation of serine/threonine phosphorylation in IL-2 signaling, the roles of protein phosphatase 1 (PP1) and 2A (PP2A) were examined. Inhibition of phosphatase activity by calyculin A treatment of YT cells resulted in a significant induction of serine phosphorylation of JAK3 and STAT5, and serine/threonine phosphorylation of IL-2R␤. Moreover, inhibition of PP2A, but not PP1, diminished IL-2-induced tyrosine phosphorylation of IL-2R␤, JAK3, and STAT5, and abolished STAT5 DNA binding activity. Serine/threonine phosphorylation of IL-2R␤ by a staurosporine-sensitive kinase also blocked its association with JAK3 and IL-2R␥ in YT cells. Taken together, these data indicate that serine/threonine phosphorylation negatively regulates IL-2 signaling at multiple levels, including receptor complex formation and JAK3/STAT5 activation, and that this regulation is counteracted by PP2A. These findings also suggest that PP2A may serve as a therapeutic target for modulating JAK3/STAT5 activation in human disease.

Selective IL-2 Responsiveness of Regulatory T Cells Through Multiple Intrinsic Mechanisms Supports the Use of Low-Dose IL-2 Therapy in Type 1 Diabetes

“…Protein phosphatase (PP)2A has been implicated in promoting IL-2R signaling in the YT cell line through its ability to dephosphorylate serine and threonine residues in IL-2Rb, JAK3, and STAT5 (20). Therefore, we tested whether the PP1/PP2A inhibitor calyculin A affected IL-2-dependent pSTAT5 activation in various populations of primary human T cells.…”

“…Tregs express high levels of FOXP3-dependent microRNA miR-155 that lowers SOCS1 and enhances IL-2R signaling (32). Another mechanism may be related to the levels of serine/threonine phosphorylation associated with IL-2Rb, JAK3, and STAT5, where this modification downregulates IL-2R signaling (20). Indeed, we showed that inhibition of the serine/threonine phosphatase activity with calyculin A, which targets PP1 and PP2A, highly inhibited IL-2-dependent pSTAT5 at low doses of IL-2 in Tregs.…”

LB100, a small molecule inhibitor of PP2A with potent chemo- and radio-sensitizing potential

“…For example, aberrant activation of the JAK/ STAT pathway is associated with numerous human malignancies, 38 and PP2A negatively regulates the activating phosphorylation of JAK3 and STAT5 in the context of IL-2 stimulation. 39 Likewise, PP2A regulates the major signaling pathway, Ras/Raf/ MEK/ERK, by directly associating with ERK2 and MEK1 and indirectly with Ras and Raf. [40][41][42][43] Constitutive activation of the Ras/Raf/MEK/ERK pathway has been well characterized in malignant transformation of susceptible cells.…”

Interleukin-2 Receptor β Thr-450 Phosphorylation Is a Positive Regulator for Receptor Complex Stability and Activation of Signaling Molecules

“…In addition, a reduction in protein migration was observed in the sample stimulated with IL-2 as well as in the CA-treated sample (Fig. 1B, lanes b and c, bottom panel), a phenomenon observed frequently in phosphorylated proteins (18). The bands corresponding to non-stimulated (75-kDa), IL-2-stimulated (80-kDa), and CA-treated (100-kDa) IL-2R␤ (Fig.…”

“…Inhibition of protein phosphatase type 2A (PP2A) diminishes STAT3, STAT6, STAT5, JAK3, and IL-2R␤ activation (18, 24 -26). Serine phosphorylation of STAT5, JAK3, and IL-2R␤ was enhanced, whereas IL-2-induced tyrosine phosphorylation was attenuated (18).…”