Carbonic anhydrase II (CAII) catalyzes the reversible hydration of carbon dioxide and plays key roles in acid base homeostasis in mammals. We found that human CAII gene promoter could be activated in human cells such as HeLa and T47D cells when the CAII promoter-luciferase gene was transfected with v-Src and assayed as a reporter of the promoter activity. Kinase negative mutants of Src, in contrast, showed little activation. The activation was completely suppressed with the introduction of a dominant-negative Ras in T47D cells, while no suppression was observed in HeLa cells. Introduction of various kinds of deletions into the CAII promoter revealed two essential regions responsible for this activation. No activation, however, was observed in activated Fyn-transfected human cells or in v-Src-transfected rodent cells. These findings suggest that Src can modulate the human CAII promoter by exerting its tyrosine kinase activity in certain human cells, and that two types of Src signaling pathways, Ras-dependent and -independent, exist in a cell type dependent manner.