Type III protein secretion is essential for the pathogenicity of Pseudomonas syringae on its host plants. Expression of HrpA, a major component of the type III secretion system (T3SS)-associated pilus, was studied both in plant leaves and in vitro using reporter genes. We found that induction of the hrpA promoter was stronger in plants than in vitro, and that the induction was enhanced by both host and nonhost plants of P. syringae pv. tomato. In vitro, the expression was enhanced by cell-free exudates from plant cell suspension cultures, added into the minimal medium. Further analysis of the plant-cell-derived, hrpA-inducing factors showed that they were small and water-soluble compounds, which could signal P. syringae the proximity of living plant cells. We also studied the production and secretion of native HrpA protein in vitro, and detected a plant-signal-dependent increase in HrpA secretion. In contrast to HrpA, the intracellular accumulation or secretion of the other T3SS-dependent proteins were not significantly increased, despite the presence of plant cell-derived, promoter-inducing factors. Thus, the accumulation of HrpA pilin seems to be subjected to a distinct post-transcriptional regulation.