The alpha-51D gene encodes surface protein 51D, which specifies serotype 51D of Paramecium tetraurelia stock 51. Previously, we isolated the gene as one expressed at much higher levels in a short-lived mutant than in its parental wild-type stock and characterized its expression to be up-regulated with increased clonal age of the wild-type stock. Here we investigated the transcriptional mechanism for the serotype expression in light of its possible causal relationship to the Paramecium clonal life span. DNA-protein binding analyses of the upstream of the alpha-51D gene identified a stretch of DNA sequence that interacted specifically with macronuclear proteins prepared from the 51D-expressing mutant. The DNA sequence was mapped to the 23 bp between -310 and -288, counting from the initiation position of 51D transcription. A protein with a binding ability for this DNA element was purified to homogeneity from the macronuclear proteins by chromatography using the specific DNA-protein interaction. In vitro transcriptional analyses showed that both the purified protein and its target domain are essential for increased synthesis of the 51D transcript. These results showed that the DNA-protein interaction is required for induction of the 51D expression in the mutant. However, aged wild-type paramecia, in which the serotype is also highly expressed, lacked the same specific transcriptional activity as detected in the mutant, suggesting another DNA-protein interaction involved in the 51D up-regulation.