Metabolism of Polyamines in Trypanosoma cruzi transfected with a recombinant plasmid bearing the C. fasciculata Ornithine decarboxylase (ODC) coding region

CARRILLO C; HUBER MA; CEJAS S; GONZÁLEZ NS; ALGRANATI ID

Salzburgo, Austria

Congreso; XI International Congress of Protozoology; 2001

Trypanosoma cruzi, a pathogenic protozoan causing Chagas disease, is an organism naturally auxotrophic for polyamines. Previous work from our laboratory have shown that T. cruzi doesn’t synthesize de novo polyamines because it lacks the ornithine decarboxylase (ODC) gene. Our recent results would indicate that T. cruzi does have neither the arginine decarboxilasa (ADC), alternative enzyme in biosynthesis of poliaminas. This activity couldn’t be detected either in soluble extracts or in membranes. On the other hand, the addition of arginine to a polyamine-free medium was not able to stimulate the proliferation of the parasite and the DNA amplification by PCR using primers with sequence highly conserved in several ADC genes failed to give any product. Epimastigotes of T. cruzi were transfected with a recombinant plasmid containing the ODC gene of Crithidia fasciculata. These parasites became autotrophic for polyamines, expressed a high level of ODC activity and could grow continuously in a semi-defined medium. Furthermore, the transfected parasites were specifically sensitive to the ODC inhibitor, alfa-difluoromethylornitine (DFMO). Growing transfected T. cruzi in the presence of the inhibitor by a long time, a DFMO resistant sub-population was developed. This culture contained a higher ODC activity (2 -5 times) than the ODC sensitive parasites. The biochemical parameters were similar in DFMO-resistant and sensitive parasites. Consistently, there was a higher number of copies of ODC gene in DFMO-resistant T. cruzi. The global comprehension of polyamine metabolism in trypanosomatid parasites could be useful to find evolution feats and to design new strategies against parasitic diseases.