Background: Protein kinase C (PKC) theta, a serine/threonine kinase, is involved in T(H)2 cell activation and proliferation. Type 2 innate lymphoid cells (ILC2s) resemble T(H)2 cells and produce the T(H)2 cytokines IL-5 and IL-13 but lack antigen-specific receptors. The mechanism by which PKC-theta drives innate immune cells to instruct T(H)2 responses in patients with allergic lung inflammation remains unknown. Objectives: We hypothesized that PKC-theta contributes to ILC2 activation and might be necessary for ILC2s to instruct the T(H)2 response. Methods: PRKCQ gene expression was assessed in innate lymphoid cell subsets purified from human PBMCs and mouse lung ILC2s. ILC2 activation and eosinophil recruitment, T(H)2-related cytokine and chemokine production, lung histopathology, interferon regulatory factor 4 (IRF4) mRNA expression, and nuclear factor of activated T cells (NFAT1) protein expression were determined. Adoptive transfer of ILC2s from wild-type mice was performed in wild-type and PKC theta-deficient (PKC-theta(-/-)) mice. Results: Here we report that PKC-theta is expressed in both human and mouse ILC2s. Mice lacking PKC-theta had reduced ILC2 numbers, T(H)2 cell numbers and activation, airway hyperresponsiveness, and expression of the transcription factors IRF4 and NFAT1. Importantly, adoptive transfer of ILC2s restored eosinophil influx and IL-4, IL-5 and IL-13 production in lung tissue, as well as T(H)2 cell activation. The pharmacologic PKC-theta inhibitor (Compound 20) administered during allergen challenge reduced ILC2 numbers and activation, as well as airway inflammation and IRF4 and NFAT1 expression. Conclusions: Therefore our findings identify PKC-theta as a critical factor for ILC2 activation that contributes to T(H)2 cell differentiation, which is associated with IRF4 and NFAT1 expression in allergic lung inflammation.