Arginase Is Essential for Survival of Leishmania donovani Promastigotes but Not Intracellular Amastigotes

Abstract

Studies of Leishmania donovani have shown that both ornithine decarboxylase and spermidine synthase, two enzymes of the polyamine biosynthetic pathway, are critical for promastigote proliferation and required for maximum infection in mice. However, the importance of arginase (ARG), the first enzyme of the polyamine pathway in Leishmania, has not been analyzed in L. donovani. To test ARG function in intact parasites, we generated Δarg null mutants in L. donovani and evaluated their ability to proliferate in vitro and trigger infections in mice. The Δarg knockout was incapable of growth in the absence of polyamine supplementation, but the auxotrophic phenotype could be bypassed by addition of either millimolar concentrations of ornithine or micromolar concentrations of putrescine or by complementation with either glycosomal or cytosolic versions of ARG. Spermidine supplementation of the medium did not circumvent the polyamine auxotrophy of the Δarg line. Although ARG was found to be essential for ornithine and polyamine synthesis, ornithine decarboxylase appeared to be the rate-limiting enzyme for polyamine production. Mouse infectivity studies revealed that the Δarg lesion reduced parasite burdens in livers by an order of magnitude but had little impact on the numbers of parasites recovered from spleens. Thus, ARG is essential for proliferation of promastigotes but not intracellular amastigotes. Coupled with previous studies, these data support a model in which L. donovani amastigotes readily salvage ornithine and have some access to host spermidine pools, while host putrescine appears to be unavailable for salvage by the parasite.

The outcome of arginase activity inhibition in BALB/c mice hosting Leishmania tropica

“…23,24 In contrast, in visceral form of Leishmaniasis, L. donovani Δarg-null mutants showed that the proliferation of promastigotes is ARG-dependent. 25 Δarg L. amazonensis strain indicated a comparable relation between NO, ARG and parasite proliferation in mice. 22 Another study has shown that the nor-NOHA is able to inhibit both mammalian and L. mexicana ARG.…”

“…Δ arg L. major has no effect on parasite burden or parasite infection, but Δ arg L. mexicana is able to reduce the parasite load by increasing NO production . In contrast, in visceral form of Leishmaniasis , L. donovani Δ arg ‐null mutants showed that the proliferation of promastigotes is ARG‐dependent . Δ arg L. amazonensis strain indicated a comparable relation between NO, ARG and parasite proliferation in mice .…”

The Polyamine Pathway as a Potential Target for Leishmaniases Chemotherapy

“…Their intracellular concentration may be regulated at the level of their biosynthesis, interconversion, degradation, and transport [103]. In Leishmania, the hydrolysis of l-arginine by L-ARG to produce ornithine and urea is a crucial initial step for polyamines production [12,102,[104][105][106] and for parasite growth and survival in promastigote and amastigote forms [107][108][109][110]. Thus, an inhibition of polyamine synthesis represents a promisor target for leishmaniases chemotherapy (Figure 2).…”

“…In addition, the importance of polyamine pathway has been described with the generation of L. donovani, L. major, L. mexicana, and L. amazonensis null mutants of essential enzymes involved in this pathway, such as ARG, ODC, AdoMetDC, or SpdS. Knockout parasites of these enzymes have been providing data of how these parasites synthesize polyamines and depend on supplementation of products of these enzymes for survival [7,8,102,105,106,138]. The role of polyamine pathway in intracellular amastigotes is controversial; in vitro and in vivo infections with L. amazonensis arginase knockout parasites present lower infection index [102,119].…”

“…The role of polyamine pathway in intracellular amastigotes is controversial; in vitro and in vivo infections with L. amazonensis arginase knockout parasites present lower infection index [102,119]. In contrast, in vivo infection with L. donovani arginase knockout parasites reduces the parasite burden in the liver of mice but does not impact the parasite burden in the spleen [106]. Supplementation with putrescine recovered the levels of infectivity of L. amazonensis in murine macrophages with reduced levels of l-arginine transport [126].…”