Effects of DFMO in the ultraestructural cell organization of Trypanosoma cruzi

CEJAS S; CARRILLO C; VALENTIM C; ALGRANATI ID; VANNIER DOS-SANTOS MA

Caxambú, Brazil

Congreso; XXIX Reunião Anual de Pesquisa em Doença de Chagas e XVIII Reunião Anual de Protozoologia; 2002

Trypanosoma cruzi lacks ornithine decarboxylase (ODC), the enzyme which converts ornithine into putrescine, the first step of polyamine biosynthesis in several eukaryotic cells. The presence of arginine decarboxylase (ADC), which converts arginine into agmatine, an alternative route, is yet under discussion in this parasite. In previous work, wild type T cruzi epimastigotes were transfected with the ODC gene from Crithida fasciculata. The resulting transgenic cultures became autotrophic for polyamines and their proliferation was sensitive to DFMO, a specific and irreversible ODC inhibitor, when they grew in a semi-defined medium essentially free of polyamines. After some time of exposure to high levels of the inhibitor, a DFMO resistant subpopulation was naturally selected. This resistant culture presented higher levels of ODC activity because an amplification in the copy number of ODC gene than sensitive cultures. With the aim of analyze the effect of DFMO in the ultrastructural cell organization, wild type and ODC-transfected Tulahuen 2 strain cells were cultured with 5 mM DFMO and studied under electronic microscopy. The transfected cells showed the kinetoplast-DNA altered in several ways, elongated, fragmented or in bizarre arrangements. Also unusual basal body numbers were observed in DFMO – treated cells. Interestingly, although DFMO is a specific ODC inhibitor, the drug also affected the mitochondria structure in wild type cells (that did not have the ODC); the effect was markedly lower though. Our results suggest that polyamines may contribute to the arrangement and organization of k-DNA in the mitochondria of T. cruzi.