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Lung M2 macrophages are regulators of airway inflammation, associated with poor lung function in allergic asthma. Previously, we demonstrated that IL-4-induced M2 gene expression correlated with tyrosine phosphorylation of the insulin receptor substrate (IRS)-2 in macrophages. We hypothesized that negative regulation of IRS-2 activity following IL-4 stimulation is dependent upon serine phosphorylation of IRS-2. Herein, we describe an inverse relationship between tyrosine phosphorylation (pY) and serine phosphorylation (pS) of IRS-2 following IL-4 stimulation. Inhibiting serine phosphatase activity increased pS-IRS-2 and decreased pY-IRS-2 leading to reduced M2 gene expression (CD200R, CCL22, MMP12, and TGM2). We found that inhibition of p70S6K, downstream of TORC1, resulted in diminished pS-IRS-2 and prolonged pY-IRS-2 as well. Inhibition of p70S6K increased expression of CD200R and CCL22 indicating that p70S6K negatively regulates some, but not all, human M2 genes. Knocking down GRB10, another negative regulatory protein downstream of TORC1, enhanced both pY-IRS-2 and increased expression of all four M2 genes. Furthermore, GRB10 associated with IRS-2, NEDD4.2 (an E3-ubiquitin ligase), IL-4Rα and γC following IL-4 stimulation. Both IL-4Rα and γC were ubiquitinated after 30 minutes of IL-4 treatment, suggesting that GRB10 may regulate degradation of the IL-4 receptor signaling complex through interactions with NEDD4.2. Taken together, these data highlight two novel regulatory proteins that could be manipulated therapeutically to limit IL-4-induced IRS-2 signaling and polarization of M2 macrophages in allergic inflammation. [ABSTRACT FROM AUTHOR]