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Parasitology

Single cell expression and chromatin accessibility of the Toxoplasma gondii lytic cycle identifies AP2XII-8 as an essential ribosome regulon driver

Abstract

The apicomplexan parasite Toxoplasma gondii undergoes a lytic replication cycle that is characterized by a coordinated gene expression program orchestrated by stage-specific transcription factors. Here, we generated and analyzed comprehensive single-cell multi-omic profiles (scRNA-seq and scATAC-seq) across the lytic cycle to understand the regulatory mechanisms governing this complex developmental process. Our analysis identified distinct cell states corresponding to G1, S/M, and cytokinesis phases with specific transcriptional signatures. By integrating chromatin accessibility and gene expression data, we identified key regulatory elements and transcription factor binding sites. We characterized the essential ApiAP2 transcription factor AP2XII-8 using CUT&RUN and conditional knockdown approaches, revealing its role as a master regulator of ribosomal gene expression. AP2XII-8 depletion causes severe growth defects and dysregulation of hundreds of genes, particularly affecting ribosome biogenesis. This study provides unprecedented insights into the regulatory mechanisms underlying T. gondii development and highlights AP2XII-8 as a critical regulator of ribosome biosynthesis, establishing a foundation for understanding apicomplexan developmental biology through multi-omic approaches.

Comparative single-cell transcriptional atlases of Babesia species reveal conserved and species-specific expression profiles

Abstract

Babesia is a genus of apicomplexan parasites that infect red blood cells in vertebrate hosts. Pathology occurs during rapid replication cycles in the asexual blood stage of infection. Current knowledge of Babesia replication cycle progression and regulation is limited and relies mostly on comparative studies with related parasites. Due to limitations in synchronizing Babesia parasites, fine-scale time-course transcriptomic resources are not readily available. Single-cell transcriptomics provides a powerful unbiased alternative for profiling asynchronous cell populations. Here, we applied single-cell RNA sequencing to 3 Babesia species (B. divergens, B. bovis, and B. bigemina). We used analytical approaches and algorithms to map the replication cycle and construct pseudo-synchronized time-course gene expression profiles. We identify clusters of co-expressed genes showing "just-in-time" expression profiles, with gradually cascading peaks throughout asexual development. Moreover, clustering analysis of reconstructed gene curves reveals coordinated timing of peak expression in epigenetic markers and transcription factors. Using a regularized Gaussian graphical model, we reconstructed co-expression networks and identified conserved and species-specific nodes. Motif analysis of a co-expression interactome of AP2 transcription factors identified specific motifs previously reported to play a role in DNA replication in Plasmodium species. Finally, we present an interactive web application to visualize and interactively explore the datasets.