CRISPR/Cas9 screens for the identification of essential host factors for Toxoplasma gondii infection

Toxoplasma gondii is an apicomplexan parasite infecting virtually all warm-blooded animals, including birds. Currently, none of the treatments available are sufficient to eliminate quiescent tissue cysts and eradicate the parasite from its host. Notably, obligate intracellular parasites such as Toxoplasma heavily rely on the host cell, e.g. for the acquisition of crucial metabolites and the disposal of toxic waste products. Still, to date little is known regarding the host factors that this pathogen requires for intracellular development and egress, as much of the existing research in the field of host-parasite interaction predominantly focuses on the parasite itself rather than the host. Here, we performed unbiased and fluorescence-activated cell sorting based genome-wide CRISPR/Cas9 knockout screens with the aim of identifying host factors that potentially contribute to initiation and progression of the lytic cycle in human cells. Based on our initial results, we designed a custom library of sgRNAs targeting 3549 enriched genes and we performed secondary small-scale pooled CRISPR screens at a higher coverage to increase confidence. These screens will be validated by in-depth phenotypic characterization of the most promising hits using single-gene KO and microscopy-based arrayed approaches to shed new light on Toxoplasma host-dependency. Some of the metabolic factors identified may also be required by other parasites of the phylum Apicomplexa, constituting shared unifying fingerprints across different species. If successful, our results would broaden existing knowledge on host-pathogen interactions and offer new promising avenues for the treatment of toxoplasmosis by specifically targeting the host instead of the parasite.