A MultiOmics time-resolved systems map of gene regulation in Toxoplasma gondii

Understanding how T. gondii builds an invasion-competent cell has been limited by its rapid, asynchronous division cycle, incomplete gene function annotation, and studies that have examined one regulator or complex at a time. Here, we placed acute perturbation of 21 transcriptional and epigenetic factors, promoter occupancy, chromatin accessibility, and molecular validation on a common single-cell clock. The resulting consensus map links the lytic-cycle programs to candidate activating and repressive inputs, organizes genes into 108 co-regulated communities, and nominates regulatory partnerships that would not have emerged from any single data type. The work bridges three longstanding gaps: it resolves perturbation responses in cell-cycle time, distinguishes promoter occupancy from perturbation-linked regulation, and turns a large multidimensional dataset into an experimentally navigable map. We indeed confirmed known factor interactions such as MORC/HDAC3 and recovered combinations such as the Cdc5/SNIP1 and DMAP1/EPL1 as coregulatory nodes (that in other systems have been described to cooperate). Furthermore, we discovered a novel apicomplexan specific combination of AAF1/AP2XI-5 that provides mechanistic insights of the unique AP2 transcription factor family.