A proteomic approach was used to study the responses of Taxus cuspidata cells to local microenvironments in different zones of immobilized support matrices. Analysis of protein spots by 2-DE revealed significant differences in the abundance of 31 spots, 28 spots, and 23 spots in outer, middle, and central zone cells between the immobilized and suspended cells. Six of these proteins, identified by MALDI-TOF-MS, were involved in the regulation of carbohydrate, nitrogen, and sulfur metabolisms. Immobilization triggered an increase in taxol production of the immobilized cells in the middle and central zones compared to that of the suspended cells. A negative relation between taxol production and the mitotic index was observed in the cells in the immobilization support matrix. Cells in the outer zone had high mitotic index and low taxol production, while cells in the middle and central zones showed low mitotic index and high taxol production. The abundance of S-adenosylmethionine synthetase, which was identified as one of the differentially expressed proteins, was positively correlated to the cell division activity in the immobilized cell cultures.