Sunday, January 29, 2023
1/29/2023
PROJECT SUMMARY/ABSTRACT Toxoplasma gondii has the remarkable ability to infect virtually any cell type of almost all warm-blooded animals and is arguably the most successful parasite on earth, having infected an estimated one-third of humans globally. While initial infection typically resolves without complication, the parasite is able to persist for the life of its host, and can re-emerge in the immunocompromised and immunosuppressed to cause fatal disease. Toxoplasma, like other apicomplexan parasites, has an unusual cell cycle in which daughter cells are fully assembled within a mother before cytokinesis. Because of its unusual features and regulatory checkpoints distinct from mammalian hosts, the parasite cell cycle is an ideal target for therapy In the final steps of Toxoplasma division, the mother cell organelles and cytoskeleton are degraded to make room for the daughter cells. Because the daughter cells must insert their own cytoskeleton, including the host cell invasion machinery, into the plasma membrane, this is a critical point in parasite assembly. Once divided, the daughter cells remain connected through a poorly understood organelle called the residual body. While the residual body was first described over 60 years ago, little is know about it. We have found that maternal cytoskeleton is actively stripped from the plasma membrane and delivered to the residual body, where it is then degraded. We have identified a ubiquitin E3 ligase that localizes to the residual body, and is essential to the turnover of maternal cytoskeleton. Moreover, we have shown that mistargeting of the E3 outside of the residual body results in premature degradation of the daughter cells' invasion machinery, blocking the infective cycle. We have therefore identified the sequestration of protein turnover as a major cellular function for the residual body. The goal of the proposed studies is to (i) understand the functional role of the residual body in Toxoplasma cytokinesis and the turnover of maternal materials, and (ii) to delineate the components of the ubiquitination cascade that targets the parasite cytoskeleton. We will determine what other organelles and structures are delivered to the residual body for turnover and identify the proteins that sequester E3 ligase activity and cytoskeleton degradation within the residual body. Furthermore, we will build the components required for the creation of an innovative reconstitution of the E3 ligase activity that will enable future studies towards generating a mechanistic model of maternal cytoskeleton turnover.
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