부산대학교 도서관

The inner membrane complex (IMC) of Toxoplasma gondii is essential for all phases of the parasite's life cycle. One of its most critical roles is to act as a scaffold for the assembly of daughter buds during replication by endodyogeny. While many daughter IMC proteins have been identified, most are recruited after bud initiation and are not essential for parasite fitness. Here, we report the identification of IMC43, a novel daughter IMC protein that is recruited at the earliest stages of daughter bud initiation. Using an auxin-inducible degron system we show that depletion of IMC43 results in aberrant morphology, dysregulation of endodyogeny, and an extreme defect in replication. Deletion analyses reveal a region of IMC43 that plays a role in localization and a C-terminal domain that is essential for the protein's function. TurboID proximity labelling and a yeast two-hybrid screen using IMC43 as bait identify 30 candidate IMC43 binding partners. We investigate two of these: the essential daughter protein IMC32 and a novel daughter IMC protein we named IMC44. We show that IMC43 is responsible for regulating the localization of both IMC32 and IMC44 at specific stages of endodyogeny and that this regulation is dependent on the essential C-terminal domain of IMC43. Using pairwise yeast two-hybrid assays, we determine that this region is also sufficient for binding to both IMC32 and IMC44. As IMC43 and IMC32 are both essential proteins, this work reveals the existence of a bud assembly complex that forms the foundation of the daughter IMC during endodyogeny. Author summary: Toxoplasma gondii is an obligate intracellular parasite that causes disease in immunocompromised individuals and congenitally infected neonates. Toxoplasma replicates using a unique process of internal budding in which two daughter buds form inside a single maternal parasite. This process is facilitated by an organelle called the inner membrane complex (IMC) which is found in Toxoplasma and other parasites in the phylum Apicomplexa. Although the IMC is known to be required for parasite replication, only a few early-recruiting IMC proteins have been identified. In this study, we identify and functionally analyze a novel IMC protein which is one of the earliest components of daughter buds and plays an essential role in parasite replication. We also identify its binding partners and demonstrate how their interactions impact the construction of daughter cells. This work reveals the existence of an essential protein complex formed in the earliest stages of parasite replication, expands our understanding of the IMC's role in Toxoplasma replication, and identifies potential targets for therapeutic intervention. [ABSTRACT FROM AUTHOR]