Toxoplasma: Sphingolipids in host-parasite interactions syntheses versus scavenging

The apicomplexans are a large group of related, single-celled microscopic organisms that cause a range of diseases (including malaria) in both humans and economically important domestic animals. One of these, Toxoplasma gondii, can infect most species of warm-blooded animals causing a disease known as toxoplasmosis. In recent times Toxoplasma has come to prominence as causing serious diseases in patients whose defences (immune response) against such invading organisms have been damaged, such as those suffering from acquired immunodeficiency syndrome ¿ AIDS. In addition, infection of unborn lambs with Toxoplasma is a major cause of miscarriage, leading to annual multi-million pound losses to UK sheep farmers. Humans and other animals usually become infected with Toxoplasma following ingestion of contaminated food or faeces. Subsequently, the parasites can invade almost any of the cells that make up the body of an animal. However, in most cases this infection does not cause serious disease, instead Toxoplasma is controlled by the immune response of the animal and retreats into cysts in the brain or in muscle tissue. These cysts can remain throughout the animal¿s life without causing any problems. However, a damaged (in an AIDS patient) or undeveloped (in an unborn animal) immune response is unable to control the parasite in this way leading to disease. Within the animal cell Toxoplasma manufactures many substances it needs for growth, however, it is also able to scavenge various materials that it uses for its own purposes. This research proposal aims to investigate how the parasite manufactures or acquires an important component of its plasma membrane (a fatty barrier that separates the inside of the parasite cell from the external environment) ¿ sphingolipid. This will help us understand how Toxoplasma grows within animal cells and how it subsequently causes disease. In addition, the observation that some of the machinery needed to make sphingolipids in Toxoplasma (and in other apicomplexans) is very unusual may eventually allow the development of new drugs against these organisms.

The important human and agricultural apicomplexan, protozoan parasite Toxoplasma gondii can infect the nucleated cells of almost any warm blooded animal, where it subsequently proliferates within a parasitophorous vacuole (PV). Although the PV is known to exhibit an intimate relationship with the host cell and Toxoplasma is able to scavenge many essential, particularly lipid, macromolecules from it, the mechanisms of this interaction and the balance between parasite de novo biosynthesis and salvage pathways remains unclear. Unlike the vacuoles harbouring many intracellular pathogens, the non-fusagenic Toxoplasma PV is not believed to interact directly with either the exo- or endocytic pathways of the host cell, and therefore is thought to be unable to scavenge essential molecules via vesicular traffic from either of these routes. Sphingolipids perform a diverse array of functions, from acting as secondary signalling molecules to forming part of micro-domains, termed lipid rafts, which have been proposed to function in processes from cell signalling to polarised trafficking. Within the context of this research proposal the roles of de novo synthesised and host acquired sphingolipid in Toxoplasma invasion and proliferation will be investigated. In the first instance this will be achieved by characterising the parasite serine palmitoyltransferase (SPT), an enzyme which mediates the first, rate-limiting step in sphingolipid biosynthesis. Unlike in all other eukaryotes studied to date, the putative SPT from Toxoplasma and other apicomplexans resembles the homodimeric enzyme from the bacterium Sphingomonas paucimobilis and is thought to be targeted to the apicoplast, a vestigal plastid which like all other plastid and mitochondrial organelles is derived from an ancient bacterial endosymbiont. Functional analyses and localisation of this unusual putative enzyme will shed new light on the evolution of sphingolipids in the Eukaryota. However, it will also allow the role of de novo sphingolipid biosynthesis in infection and proliferation to be delineated by the creation of parasites in which SPT function is ablated. Similar investigations assaying for the ability of Toxoplasma to proliferate within host cells deficient in SPT activity themselves will address whether host derived sphingolipid is required for the establishment of an infection. As such the research outlined here will establish the roles of de novo synthesised and host derived sphingolipid in Toxoplasma invasion and proliferation, thereby greatly adding to our understanding of how this parasite interacts with its host.