O papel de proteínas da família Y-box de parasitos na resposta ao estresse oxidativo

Abstract

Y-box binding proteins (YBP) are considered multifunctional regulators of gene expression that present two characteristic domains: a highly conserved nucleic acid binding domain (cold shock domain, CSD) and a less conserved C-terminal domain (tail domain). Several studies have demonstrated the role of some YBP in the cellular response to stress caused by genotoxic agents, the ability to interact with injuries and DNA mismatch repair, as well as interaction with some proteins of the DNA repair pathways showing that the YBP may participate in DNA repair processes. SMYB1 is a Schistosoma mansoni protein that presents a CSD highly similar to CSDs from other YBP family members. S. mansoni, the etiologic agent of schistosomiasis, is a trematode parasite with a complex life cycle that alternates between two different hosts and also lives in diverse environments. The characterization of proteins involved in the regulation of gene expression is of great importance for the understanding of molecular events that control physiological and morphological changes in such organisms. The present study aims to increase the current knowledge about the SMYB1 protein and its biological function. To this end, immunolocalization experiments were performed in different S. mansoni life cycle stages, revealing the presence of this protein in the citoplasm of eggs, male and female adult worms, cercaria, miracidia and schistosomulae. The citoplasmic localization of SMYB1 is in agreement with previous findings from our research group that have shown its ability to interact with RNA molecules. The interaction with mRNAs by other YBP members is well-established and suggests a possible function in the regulation of post-transcriptional gene expression events. The role of SMYB1 in the protection against oxidative stress was also evaluated. To reach this goal, we have obtained MSH2 single knockout lines of Trypanosoma cruzi MSH2 double knockout lines of Trypanosoma brucei and verified that SMYB1 is able to complement the function of MSH2 in both parasites. SMYB1-expressing strains were more resistant to hydrogen peroxide treatment, suggesting that this protein may recognize oxidative lesions in the DNA (including 8-oxoG, which is increased in the presence of hydrogen peroxide) and a response to that injury. RBP16, an ortholog of SMYB1, is a mitochondrial RNA binding protein from Trypanosoma brucei that contains a CSD 33-38% identical to CSDs of other eukaryotic YBPs. Therefore, another aim of this work was to evaluate the role of RBP16 in wildtype parasites and the behaviour of such organisms in the presence of genotoxic agents. Evaluating the effect of RBP16 silencing in T. brucei, we have verified that this is an essential gene for parasite growth and response to oxidative stress. Nevertheless, the decreasing of protein levels did not increase cell sensitivity to DNA damages caused by cisplatin and metilmetanosulfonate. DNA repair processes are crucial to the parasitic success of both T. cruzi and T. brucei and, therefore, a better understanding of the mechanisms involved in the repair of damaged DNA may help to explain the biology of Trypanosomatids and lead to the development of new strategies for the treatment of diseases caused by such parasites.