One striking feature of the biology of trypanosomes is the changes in mitochondrial structure and function that occur as these parasites transition from one life cycle stage to another. Our laboratory has been interested in the role the mitochondrial ATP synthase plays in mitochondrial changes through the life cycle. Analysis of the recently completed T. brucei genome suggested that there may be multiple putative genes encoding ATP synthase subunit c. While homologous in their 3' ends, these genes differ in their 5' ends and, if expressed, would result in three distinct proteins. Our analysis showed that all three of the possible transcripts were detected in both procyclic and bloodstream stages, although the c-3 transcript was less abundant than that for c-1 or c-2. The three isoforms of subunit c are produced in both the bloodstream and procyclic stages and their mature protein products possess distinct N-terminal regions of the protein as found within mitochondria. All three isoforms are also incorporated into the assembled ATP synthase complex from procyclic cells. Although multiple subunit c genes have been found in other organisms, they produce identical polypeptides and the finding of significant differences in the mature proteins is unique to T. brucei.