Leishmania amazonensis Arginase Compartmentalization in the Glycosome Is Important for Parasite Infectivity

Maria Fernanda Laranjeira Da Silva,Lucile M Floeter-Winter,Stephen M Beverley,Sandra M Muxel,Ricardo Andrade Zampieri,Fabio T M Costa

doi:10.1371/journal.pone.0034022

Maria Fernanda Laranjeira Da Silva, Lucile M Floeter-Winter + Show 4 more

Open Access

https://doi.org/10.1371/journal.pone.0034022

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Abstract

In Leishmania, de novo polyamine synthesis is initiated by the cleavage of L-arginine to urea and L-ornithine by the action of arginase (ARG, E.C. 3.5.3.1). Previous studies in L. major and L. mexicana showed that ARG is essential for in vitro growth in the absence of polyamines and needed for full infectivity in animal infections. The ARG protein is normally found within the parasite glycosome, and here we examined whether this localization is required for survival and infectivity. First, the localization of L. amazonensis ARG in the glycosome was confirmed in both the promastigote and amastigote stages. As in other species, arg− L. amazonensis required putrescine for growth and presented an attenuated infectivity. Restoration of a wild type ARG to the arg − mutant restored ARG expression, growth and infectivity. In contrast, restoration of a cytosol-targeted ARG lacking the glycosomal SKL targeting sequence (argΔSKL) restored growth but failed to restore infectivity. Further study showed that the ARGΔSKL protein was found in the cytosol as expected, but at very low levels. Our results indicate that the proper compartmentalization of L. amazonensis arginase in the glycosome is important for enzyme activity and optimal infectivity. Our conjecture is that parasite arginase participates in a complex equilibrium that defines the fate of L-arginine and that its proper subcellular location may be essential for this physiological orchestration.

Highlights

Leishmaniasis is the second most important infection caused by a protozoan and affects 12 million people worldwide (WHO)
L. amazonensis ARG remains in the glycosome in the amastigote form during macrophage infection
Infected J774A 1 macrophages were probed with anti-ARG polyclonal antibodies to determine ARG localization in the L. amazonensis amastigote form by electron microscopy (EM) immunolocalization experiments (Figure 1A)

Summary

Introduction

Leishmaniasis is the second most important infection caused by a protozoan and affects 12 million people worldwide (WHO). There is no vaccine to prevent leishmaniasis, and the drug arsenal to treat this disease presents many problems, including toxicity and increasing parasite resistance to common chemotherapies. The causative agents of leishmaniasis are protozoan parasites of the genus Leishmania, family Trypanosomatidae. The life cycle of Leishmania includes the extracellular promastigote form that resides in the midgut of the phlebotomine sand fly vector. Leishmania is able to survive and replicate in these different environments by adapting to a wide range of temperatures, pH levels and nutrient availability and by escaping the anti-proliferative defense molecules produced by the host cell [1,2]

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