Effect of BFD2 (TGME49_311100) knockout on T. gondii gene expression during BFD1 induction

The acute-to-chronic stage transition in Toxoplasma gondii involes a global restructuring of the parasite transcriptome. Prior work identified a single transcription factor (BFD1) that is both necessary and sufficient to establish the chronic-stage program, making it the master regulator of this process. Recently, we found that parasites lacking a CCCH-type zinc finger RNA-binding protein (BFD2) are also unable to produce chronic forms. To understand how BFD2 aids in establishing the BFD1 transcriptional program, we knocked out BFD2 in a BFD1-conditional expression strain, wherein BFD1 is induced by treatment with the small molecule Shield-1. Overall design: Comparative RNA-seq analysis was performed on conditional BFD1 expression strains in the presence (DD-BFD1) and absence of BFD2 (DD-BFD1/?bfd2), after 48h of treatment with either 3 µM Shield-1 to induce BFD1 expression or an equivalent volume of ethanol (i.e. vehicle). Experiments were performed in biological triplicate (separate infections) and technical duplicate (i.e. samples split in two at the time of harvest and processed in parallel for all subsequent steps, from RNA isolation through library prepartion and sequencing).