Basophils mediate many of their biological functions by producing IL-4. However, it is unknown how the Il4 gene is regulated in basophils. Here, we report that CCAAT/enhancer-binding protein α (C/EBPα), a major myeloid transcription factor, was highly expressed in basophils. We show that C/EBPα selectively activated Il4 promoter-luciferase reporter gene transcription in response to IgE cross-linking, but C/EBPα did not activate other known Th2 or mast cell enhancers. We found that the PI3K pathway and calcineurin were essential in C/EBPα-driven Il4 promoter-luciferase gene transcription. Our mutation analyses revealed that C/EBPα drove Il4 promoter-luciferase activity depending on its DNA binding domain. Mutation of the C/EBPα-binding site in the Il4 promoter region abolished C/EBPα-driven Il4 promoter-luciferase activity. Our results further showed that a mutation in nuclear factor of activated T cells (NFAT)-binding sites in the Il4 promoter also negated C/EBPα-driven Il4 promoter-luciferase activity. Our study demonstrates that C/EBPα, in cooperation with NFAT, directly regulates Il4 gene transcription.