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  1. Transcriptional dynamics of a large family of surface protein genes in toxoplasma gondii

    Theisen, Terence Charles
    [Stanford, California] : [Stanford University], 2020

    Toxoplasma gondii is a eukaryotic intracellular parasite that has evolved numerous large, paralogous gene families. These families are critical for supporting Toxoplasma's unparalleled host range which consists of nearly any nucleated cell in almost any warm-blooded animal. The SRS (SAG1-related sequence) gene family encodes over 100 surface proteins, some of which are involved in parasite attachment and evading the immune response of a host. For most SRS genes, however, little is understood about their function and expression profile. Additionally, Toxoplasma undergoes a complex and poorly understood developmental process that is critical for establishing a chronic infection in its intermediate hosts. In chapter 1, I review the current literature on the SRS family and present novel analyses on how SRS gene expression changes across the life stages of Toxoplasma. In chapter 2, I describe efforts to apply single-cell RNA-sequencing to Toxoplasma parasites to create a comprehensive atlas of cell cycle and asexual development based on data from over 5,400 parasites. This data reveals unexpected heterogeneity in SRS gene expression among individual parasites: in a population of the rapidly growing tachyzoite stage nearly all > 100+ SRS genes are detected despite only an average of 7 SRS genes being "on" in any individual tachyzoite at a given time. From this dataset, AP2IX-1 is identified as a parasite transcription factor that triggers downregulation of SAG1, a ubiquitous surface antigen and SRS, and upregulation of transcripts typically only expressed in the sexual stages of Toxoplasma's lifecycle. In chapter 3, I describe efforts to determine whether heterogeneous SRS expression signatures are inherited over generations of parasite replication by comparing transcriptomics from clonally derived parasites. Preliminary data from these studies suggests that SRS expression patterns are not strongly inherited by progeny and that heterogeneity of SRS expression arises within 10 generations between subcloned parasites. In chapter 4, I present work that identified new parasite factors involved in the secretion of Toxoplasma effector proteins into the host cell during infection. In chapter 5, I conclude with remarks on future directions for the study of the SRS gene family and understanding heterogeneity in expression patterns in Toxoplasma

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