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.
Theiler’s murine encephalomyelitis virus (TMEV or Theiler’s virus) is a neurotropic picornavirus that can persist lifelong in the central nervous system of infected mice, thereby causing a chronic ...inflammatory demyelinating disease. The leader (L) protein of the virus is an important determinant of viral persistence. It was shown to inhibit transcription of type I interferon (IFN) genes and to cause nucleocytoplasmic redistribution of host proteins. Here, we report that expression of L shuts-off the synthesis of green fluorescent protein and of firefly luciferase, suggesting that it induces a global shut-off of host protein expression. L inhibited neither the transcription nor the translation of the reporter genes but blocked cellular mRNA export from the nucleus. This activity correlated with the phosphorylation of nucleoporin 98 (Nup98) an essential component of the nuclear pore complex. On the other hand, our data confirm that L inhibits IFN expression at the transcriptional level and show that transcription of other chemokine or cytokine genes is affected by L. This transcriptional inhibition correlated with inhibition of interferon regulatory factor 3 (IRF-3) dimerization. Whether inhibition of IRF-3 dimerization and dysfunction of the nuclear pore complex are related phenomena remains an open question. In vivo, IFN antagonism appears to be an important role of L early during infection since a virus bearing a mutation in the zinc finger of L replicated as well as the wild-type virus in type I IFN receptor-deficient mice (IFNAR-KO) but had impaired fitness in IFN-competent mice.
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