Toxoplasma F-box protein 1 is required for daughter cell scaffold function during parasite replication

Abstract

By binding to the adaptor protein SKP1 and serving as substrate receptors for the SKP1 Cullin, F-box E3 ubiquitin ligase complex, F-box proteins regulate critical cellular processes including cell cycle progression and membrane trafficking. While F-box proteins are conserved throughout eukaryotes and are well studied in yeast, plants, and animals, studies in parasitic protozoa are lagging. We have identified eighteen putative F-box proteins in the Toxoplasma genome of which four have predicted homologs in Plasmodium . Two of the conserved F-box proteins were demonstrated to be important for Toxoplasma fitness and here we focus on an F-box protein, named TgFBXO1, because it is the most highly expressed by replicative tachyzoites and was also identified in an interactome screen as a Toxoplasma SKP1 binding protein. TgFBXO1 interacts with Toxoplasma SKP1 confirming it as a bona fide F-box protein. In interphase parasites, TgFBXO1 is a component of the Inner Membrane Complex (IMC), which is an organelle that underlies the plasma membrane. Early during replication, TgFBXO1 localizes to the developing daughter cell scaffold, which is the site where the daughter cell IMC and microtubules form and extend from. TgFBXO1 localization to the daughter cell scaffold required centrosome duplication but before kinetochore separation was completed. Daughter cell scaffold localization required TgFBXO1 N-myristoylation and was dependent on the small molecular weight GTPase, TgRab11b. Finally, we demonstrate that TgFBXO1 is required for parasite growth due to its function as a daughter cell scaffold effector. TgFBXO1 is the first F-box protein to be studied in apicomplexan parasites and represents the first protein demonstrated to be important for daughter cell scaffold function.

Toxoplasma gondii UBL‐UBA shuttle proteins contribute to the degradation of ubiquitinylated proteins and are important for synchronous cell division and virulence

“…38 Disruption of TgOTUD3A results in dysregulation of cell cycle progression, suggesting the important roles of ubiquitin-mediated proteins degradation in cell cycle regulation. 39 In addition, studies about ECR1 and FBXO1, two putative E3 ligase which are important factors for Toxoplasma replication, have elucidated partial molecular basis for Toxoplasma endodyogeny.. 4,40 All these results show that ubiquitinylation plays key roles in Toxoplasma cell cycle regulation. Our present works that deletion of UBL-UBA shuttle proteins results in K48-linked polyubiquitinylated proteins accumulation and cell division asynchronous also confirmed this conclusion, however, the precise mechanisms are unknown.…”

“…necessary to study the mechanisms of T gondii cell division, which is a unique process termed endodyogeny where two daughter parasites develop within the mother cell. 4,5 Protein ubiquitinylation, a universal posttranslational modification (PTM) in eukaryotic cells, plays vital roles in regulating cell division of T gondii by timely destroying proteins that are short-lived, unneeded, damaged, or misfolded, such as the components of replisome and inner membrane complex. 6,7 Ubiquitinylated proteins are degraded by the ubiquitin proteasome system (UPS), which employs numbers of enzymes to ubiquitinate substrate proteins and then, delivers the ubiquitinylated proteins to the proteasome for degradation.…”

Myristoylation, an Ancient Protein Modification Mirroring Eukaryogenesis and Evolution

“…TgFBO1 is an inner membrane complex MYRed component with an F-box domain that might bind SKIP1 [69]. In humans, ten such MYRed E3 cullins were identified [7,70].…”

COP9 signalosome is an essential and druggable parasite target that regulates protein degradation

“…Cullin proteins form a scaffold on which cullin-RING E3 ubiquitin ligases (CRLs) assemble. The ubiquitin ligase activity of CRLs is controlled by cycles of attachment and removal of the Nedd8 [29,31,[40][41][42][43][44][45][46]. A conserved cullin lysine residue forms an isopeptide bond with the https://doi.org/10.1371/journal.ppat.1008952.g002…”