The Role of the Mitochondrial Glycine Cleavage Complex in the Metabolism and Virulence of the Protozoan Parasite Leishmania major

Abstract

For the human pathogen Leishmania major, a key metabolic function is the synthesis of thymidylate, which requires 5,10-methylenetetrahydrofolate (5,10-CH 2 -THF). 5,10-CH 2 -THF can be synthesized from glycine by the mitochondrial glycine cleavage complex (GCC). Bioinformatic analysis revealed the four subunits of the GCC in the L. major genome, and the role of the GCC in parasite metabolism and virulence was assessed through studies of the P subunit (glycine decarboxylase (GCVP)). First, a tagged GCVP protein was expressed and localized to the parasite mitochondrion. Second, a gcvP ؊ mutant was generated and shown to lack significant GCC activity using an indirect in vivo assay after incorporation of label from [2-14 C]glycine into DNA. The gcvP ؊ mutant grew poorly in the presence of excess glycine or minimal serine; these studies also established that L. major promastigotes require serine for optimal growth. Although gcvP ؊ promastigotes and amastigotes showed normal virulence in macrophage infections in vitro, both forms of the parasite showed substantially delayed replication and lesion pathology in infections of both genetically susceptible or resistant mice. These data suggest that, as the physiology of the infection site changes during the course of infection, so do the metabolic constraints on parasite replication. This conclusion has great significance to the interpretation of metabolic requirements for virulence. Last, these studies call attention in trypanosomatid protozoa to the key metabolic intermediate 5,10-CH 2 -THF, situated at the junction of serine, glycine, and thymidylate metabolism. Notably, genome-based predictions suggest the related parasite Trypanosoma brucei is totally dependent on the GCC for 5,10-CH 2 -THF synthesis.

The contribution of the glycine cleavage system to the pathogenesis of Francisella tularensis

“…This finding is likely explained by the presence of an additional serine biosynthetic pathway, but may also indicate that the gut is not a serine-limiting environment. Surprisingly, our findings with F. tularensis are similar to in vitro and in vivo phenotypes in the trypanosomatid, Leishmania major [32]. Despite differences between prokaryotes and eukaryotes, these human pathogens both lack alternative serine biosynthetic pathways and thus appear to rely on the GCS during infection.…”

Lipoic Acid Metabolism in Microbial Pathogens

“…Among these, only the role of the GCV in Leishmania biology has been characterized, and this complex is necessary for normal replication and pathology in vivo. Genetic disruption of the gene encoding the L. major GCV P protein reduces parasite virulence and leads to attenuated infections in mice (206). In contrast, the GCV is not required for the in vitro growth of parasites in macrophages, although these parasites do show increased sensitivity to elevated glycine or reduced serine levels (206).…”

“…Animal pathogens are also able to scavenge the free lipoate generated by host digestion (3,29,164). 206 SPALDING AND PRIGGE MICROBIOL. MOL.…”

“…Genetic disruption of the gene encoding the L. major GCV P protein reduces parasite virulence and leads to attenuated infections in mice (206). In contrast, the GCV is not required for the in vitro growth of parasites in macrophages, although these parasites do show increased sensitivity to elevated glycine or reduced serine levels (206). One reason for a severe in vivo, but not in vitro, phenotype may be that the GCV allows L. major to adapt to metabolic perturbations associated with inflammatory responses in vivo.…”

Effect of Human Urine on Cell Cycle and Infectivity of Leismania Species Promastigotes In Vitro

“…36 In a study of Trypanosoma cruzi and T. rangeli , an increase in the number of infective tripomastigotes was detected in urine-containing culture. 37 In our study, infectivity of Leishmania promastigotes increased in cultures containing urine compared with control cultures that did not contain urine.…”