Two closely related casein kinase I (CKI) isoforms, CKIδ and CKIε, are ubiquitously expressed in many human tissues, but their specific biologic function remains to be clarified. Here, we provide the first evidence that CKIε is involved in hematopoietic cell differentiation. CKIε, but not CKIδ, was down-regulated along with human granulocytic differentiation. The specific down-regulation was observed in granulocyte colony-stimulating factor (G-CSF)-induced cell differentiation of murine interleukin-3 (IL-3)-dependent myeloid progenitor 32D cells. Introduction of wild-type (WT)-CKIε into 32D cells inhibited the G-CSF-induced cell differentiation, whereas kinase-negative (KN)-CKIε promoted the differentiation. Neither WT- nor KN-CKIε affected IL-3-dependent cell growth. Moreover, introduction of WT- or KN-CKIδ, did not affect the cytokine-induced cell growth and differentiation. While G-CSF-induced activation of signal transducers and activators of transcription 3 (STAT3) was sustained by KN-CKIε1, STAT3 activation was attenuated by WT-CKIε. This may be explained by the fact that the suppressor of cytokine signaling 3 (SOCS3) was stabilized by its physical association with CKIε. Such stabilization by CKIε was also seen in IL-3-induced β-catenin. The stabilization of downstream components of cytokine and Wnt signaling by CKIε might be critical for integration of several intracellular signaling pathways to a cell-specific biologic response in hematopoietic cell self-renewal.