Complement component C3a causes a robust degranulation in human mast cells. Whether C3a also stimulates chemokine production in human mast cells and what signaling pathway it activates is not known. In the present study, we demonstrate that CD34⁺cell-derived primary mast cells and a human mast cell line LAD 2 express surface C3a receptors at similar levels. Furthermore, C3a caused ∼50% internalization of cell surface C3a receptors in both cell types. We therefore used LAD 2 cells as a model to study C3a-induced biological responses and signaling in human mast cells. We found that C3a stimulated substantial degranulation and induced chemokine monocyte chemoattractant protein 1 (MCP-1/CCL2) and regulated upon activation, normal T-cell expressed and secreted (RANTES/CCL5) production in LAD 2 mast cells. C3a caused a rapid and sustained extracellular-signal-regulated kinase (ERK) phosphorylation and Akt phosphorylation in LAD 2 mast cells. Furthermore, U0126 and LY294002, which respectively inhibit MEK-induced ERK phosphorylation and PI3 kinase-mediated Akt phosphorylation had distinct effects on C3a-induced responses. Thus, U0126, which blocked C3a-induced RANTES/CCL5 production by 50.6±2.3%, inhibited MCP-1/CCL2 generation by 85.2±0.6%. In contrast, LY294002 had no effect on C3a-induced RANTES/CCL5 production but blocked MCP-1/CCL2 generation by 83.7±1.5%. These data demonstrate that C3a activates divergent signaling pathways to induce chemokine production in human mast cells.