Involvement of nucleoside diphosphate kinase b and elongation factor 2 in Leishmania braziliensis antimony resistance phenotype

Douglas S Moreira; Silvane M F Murta

doi:10.1186/s13071-016-1930-6

Involvement of nucleoside diphosphate kinase b and elongation factor 2 in Leishmania braziliensis antimony resistance phenotype

Parasit Vectors. 2016 Dec 13;9(1):641. doi: 10.1186/s13071-016-1930-6.

Authors

Douglas S Moreira¹, Silvane M F Murta²

Affiliations

¹ Centro de Pesquisas René Rachou CPqRR, Fundação Oswaldo Cruz - FIOCRUZ, Avenida Augusto de Lima 1715, Belo Horizonte, MG, Brazil.
² Centro de Pesquisas René Rachou CPqRR, Fundação Oswaldo Cruz - FIOCRUZ, Avenida Augusto de Lima 1715, Belo Horizonte, MG, Brazil. silvane@cpqrr.fiocruz.br.

Abstract

Background: Nucleoside diphosphate kinase b (NDKb) is responsible for nucleoside triphosphates synthesis and it has key role in the purine metabolism in trypanosomatid protozoans. Elongation factor 2 (EF2) is an important factor for protein synthesis. Recently, our phosphoproteomic analysis demonstrated that NDKb and EF2 proteins were phosphorylated and dephosphorylated in antimony (Sb^III)-resistant L. braziliensis line compared to its Sb^III-susceptible pair, respectively.

Methods: In this study, the overexpression of NDKb and EF2 genes in L. braziliensis and L. infantum was performed to investigate the contribution of these proteins in the Sb^III-resistance phenotype. Furthermore, we examined the role of lamivudine on Sb^III susceptibility in clones that overexpress the NDKb gene, and the effect of EF2 kinase (EF2K) inhibitor on the growth of EF2-overexpressing parasites.

Results: Western blot analysis demonstrated that NDKb and EF2 proteins are more and less expressed, respectively, in Sb^III-resistant line of L. braziliensis than its wild-type (WTS) counterpart, corroborating our previous phosphoproteomic data. NDKb or EF2-overexpressing L. braziliensis lines were 1.6 to 2.1-fold more resistant to Sb^III than the untransfected WTS line. In contrast, no difference in Sb^III susceptibility was observed in L. infantum parasites overexpressing NDKb or EF2. Susceptibility assays showed that NDKb-overexpressing L. braziliensis lines presented elevated resistance to lamivudine, an antiviral agent, but it did not alter the leishmanicidal activity in association with Sb^III. EF2-overexpressing L. braziliensis clone was slightly more resistant to EF2K inhibitor than the WTS line. Surprisingly, this inhibitor increased the antileishmanial effect of Sb^III, suggesting that this association might be a valuable strategy for leishmaniasis chemotherapy.

Conclusion: Our findings represent the first study of NDKb and EF2 genes overexpression that demonstrates an increase of Sb^III resistance in L. braziliensis which can contribute to develop new strategies for leishmaniasis treatment.

Keywords: Antimony resistance; Chemotherapy; Elongation factor 2; Leishmania spp.; Nucleoside diphosphate kinase b.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Antimony / toxicity*
Blotting, Western
Drug Resistance*
Gene Expression Profiling
Leishmania braziliensis / drug effects*
Leishmania braziliensis / genetics*
Leishmania infantum / drug effects
Leishmania infantum / genetics
NM23 Nucleoside Diphosphate Kinases / analysis*
NM23 Nucleoside Diphosphate Kinases / genetics
Peptide Elongation Factor 2 / analysis*
Peptide Elongation Factor 2 / genetics

Substances

NM23 Nucleoside Diphosphate Kinases
Peptide Elongation Factor 2
Antimony