Trypanosoma cruzi, the etiologic agent of
Chagas' disease, is a
polyamine auxotroph organism because its genome contains neither
ornithine decarboxylase (ODC) nor
arginine decarboxylase (ADC) genes, presumably lost during evolution. After transformation with a recombinant plasmid bearing the complete coding region of Crithidia fasciculata ODC gene, the transgenic parasites were able to synthesize
putrescine and simultaneously became susceptible to
alpha-difluoromethylornithine (DFMO), an irreversible inhibitor of ODC. We have studied the emergence of DFMO-resistant T. cruzi after one-step selection of ODC-transformed parasites cultivated in the presence of high levels of the
drug (5 mM). Our results have indicated a duplication of the ODC gene copy number in the
drug-resistant cell line. The ODC transcripts and the corresponding translation products showed very significant increases (about 7- and 25-fold, respectively) in DFMO-resistant parasites, while the ODC enzymatic activity was 5 times higher than in
drug-sensitive T. cruzi. The unequal increases of ODC
protein and enzymatic activity in DFMO-resistant protozoa strongly suggest that in addition to gene amplification and enhanced transcription and translation, the assembly of ODC
polypeptide chains into dimeric active
enzyme molecules might also contribute to regulate the development of DFMO resistance.