OVERVIEW: Our research
Toxoplasma gondii is a protozoan parasite that is able to infect and persist in many warm-blooded hosts, from birds to humans. In some mammalian hosts, including humans and mice, Toxoplasma persists primarily in the central nervous system (CNS). Toxoplasma’s ability to cause a lifelong infection in the CNS requires that the CNS and Toxoplasma co-evolve to avoid an overly-exuberant CNS immune response that could destroy both the host (the CNS) and the microbe. By understand the molecular mechanism that underlie this persistence, we hope to: (1.) determine new targets for treating chronic CNS toxoplasmosis, for which there is currently no therapy, and (2.) define a new mechanism for driving and modulating the CNS immune response. Ultimately, the latter goal may help us develop treatments for diseases such as Alzheimer’s and stroke, in which increased CNS inflammation plays a significant role.
THE MICROBE: Toxoplasma gondii
Toxoplasma gondii is a eukaryotic, obligate intracellular parasite that infects up to 1/8 of the world’s human population. Congenital infections with Toxoplasma or reactivation of the parasite in severely immunocompromised patients (e.g. AIDS patients) can lead to devastating neurologic consequences but most people infected with Toxoplasma experience no effects from this life-long CNS infection. Thus, Toxoplasma has evolved mechanisms to avoid provoking a CNS immune response that would lead to its clearance and it is these mechanisms we would like to understand. In addition to being a medically important neurotropic microbe, Toxoplasma has certain characteristics that make it a great tool for understanding CNS-microbe interactions. These include:
- Genetic tractability & asexual haploid life cycle (i.e. you can knock in and out genes relatively easily)
- Strain differences in acute and chronic stages (these differences will make it much easier to detect how Toxoplasma differentially manipulates host cells and then enables us to determine which Toxoplasma proteins are critical to these manipulations)
- Natural pathogen in mice (which allows us to use mice to answer questions about the Toxoplasma-CNS interaction)
- Toxoplasma community is a friendly, fun-loving group that likes to collaborate and do science! Out of this group has grown ToxoDB (which is part of Eukaryotic DB) which is database/repository for the ‘omics that are being performed in Toxo.
THE HOST: Mice and the CNS
While studies have been done on immune cells (who, how, why) that enter the CNS during neuro-inflammation, it has only recently been recognized that the cells of the CNS likely have a function in modulating the inflammatory response that occurs in the CNS. There are a variety of cells in the CNS but we focus on the major parenchymal cells of the CNS: neurons, which are the cells in which Toxoplasma resides in the chronic state; astrocytes, which are cells that have essential functions in synapse pruning, development of coordinated neuronal networks, neurotransmitter recycling, and innate immunity of the CNS; and microglia, which are the resident macrophage of the CNS. How Toxoplasma interacts and manipulates these cells is the focus of our lab.
We use mice to study this interaction because they are natural hosts for Toxoplasma, are mammals (Iike humans) and unlike humans, there are many genetically modified mice which enables us to figure things out in vivo (in live animals).
Ultimately, since Toxoplasma is an obligate intracellular parasite, we hypothesize that a major part of how the parasite persists in the brain is mediated by the host cell-parasite interaction and so we focus on molecularly interrogating this interaction.
Setting up the Toxoplasma-Cre system:
In order to molecularly define the Toxoplasma-CNS interaction, we needed to identify the chronically infected cells in the CNS of mice. Previously, most attempts to do this have used electron microscopy, which makes the tissue unusable for other purposes such as collecting cells of interest for RNA expression analysis. Anita sought to address this issue during her postdoctoral fellowship with John Boothroyd where a very smart colleague of hers, Gusti Zeiner, suggested that she use site specific recombination (SSR) triggered by the parasite. Basically, by engineering parasites to inject in an enzyme called Cre recombinase, these Toxoplasma-Cre parasites could be used to infect Cre-reporter mice, which turn on specific genes only if Cre-mediated recombination in that cell has occurred. The beauty of this assay is that it marks cells that have interacted with Toxoplasma, and since the modification occurs at the level of the DNA, the gene is permanently turned on even if the cell was infected weeks or months before (and so is very useful for studying chronic CNS infections).
With this tool in hand (the Toxoplasma-Cre system), the Koshy lab was born. We moved to the University of Arizona in 2012, where we continue to have two goals: performing great science that will improve human health and having fun along the way.