Caracterização das subunidades catalíticas do proteassoma 20S em cepas de Trypanosoma cruzi susceptíveis e resistentes ao benzonidazol

AUTOR(ES)
DATA DE PUBLICAÇÃO

2007

RESUMO

Chagas disease is caused by Trypanosoma cruzi. According to WHO, in Latin America, 16-18 million people is infected with this protozoan. Due the involvement of the proteasome in the cellular biology of the T. cruzi and the low efficiency of the drugs used in the individuals treatment with Chagas disease, we believed that the characterization of the catalytic subunits of the 20S proteasome, would help in the development of future drugs that could be used in the therapeutics of the Chagas disease. This work shows the molecular and biochemical characterization of the catalytic subunits of the 20S proteasome in a T. cruzi population with in vitro induced (17WTS and 17LER), in vivo (BZS and BZR) and strains naturally sensitive and resistant to benznidazole (CL and Colombian). Chromosomal localization by using the pulse field gel electrophoresis and hybridization profiles with specific probes showed several pattern in the different strains, suggesting that these genes are not associated with the drugs phenotype resistance in T. cruzi. Southern blot analyses showed that these genes are present in at least two copies for genome haploid of the parasite. The expression of those genes was evaluated through of Northern blot analyses and the levels of the 20S proteasome identified using Western blot and antibodies against alpha subunits. The mRNA levels did not vary among the strains, but a larger level of 20S proteasome were detected in the sensitive strains (17WTS, BZS and CL) when compared the resistant strains (17LER, BZR and Colombian). The absent of accumulation of poly-ubiquitin conjugated, suggest that the poly-ubiquitination pathway and degradation by 26S proteasome, are strongly regulated in the strains of T. cruzi studied. The measure of the peptidase activity using specific substrate for the subunit Beta-1, -2 and -5, showed that the main proteolytic activity is tripsin-like in all strains. Moreover, the strains 17WTS and 17LER also presented a high quimotripsin-like activity. These results obtained until the moment suggest that, in the epimastigote development stage of the T. cruzi the post-translation modifications are more efficient by the regulation of 20S proteasome function than the transcriptional or posttranscriptional modification.

ASSUNTO(S)

biofisica molecular 1.trypanosoma cruzi - teses. 2. drogas - resistência - teses.

ACESSO AO ARTIGO

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