The TORC1-activated proteins, p70S6K and GRB10, regulate IL-4 signaling and M2 macrophage polarization by modulating phosphorylation of insulin receptor substrate-2 (CCR3P.214)

Abstract

Abstract The abundance of M2 macrophages in the airways of both human asthmatics and mouse models correlates with disease severity. M2 macrophage differentiation is induced by IL-4 and IL-13. The cytosolic adaptor, IRS-2, is tyrosine phosphorylated (pY-IRS-2) following engagement of the type I IL-4 receptor and it mediates M2 gene expression. Therefore, the goal of this study was to determine negative regulators of IRS-2 activity. We found an inverse relationship between tyrosine and serine phosphorylation of IRS-2. IRS-2 was highly serine phosphorylated (pS-IRS-2) in unstimulated cells and pS-IRS-2 levels decreased as pY-IRS-2 increased following IL-4 stimulation. Inhibiting serine phosphatase activity caused a dramatic increase in IRS-2 serine phosphorylation and IL-4-induced M2 gene expression was ablated. By using phospho-serine kinase arrays and inhibitors of serine/threonine kinases, we showed that TORC1 and p70S6K inhibition but not TORC2, JNK or GSK3β inhibition, resulted in prolonged pY-IRS-2. p70S6K inhibition also augmented expression of the hallmark human M2 macrophage genes, CD200R, CCL22, and TGM2. Lastly, we showed interaction between IL-4Rα, γC, IRS-2 and the TORC1-activated protein GRB10. Using siRNAs to GRB10, we enhanced pY-IRS-2, prolonged IL-4 receptor expression and increased M2 gene expression in human monocytes after IL-4 stimulation. These data highlight p70S6K and GRB10, downstream of TORC1, as key negative regulators of IL-4 signaling in human monocytes.