Protein phosphatase 2A plays a crucial role in Giardia lamblia differentiation

Abstract

The ability of Giardia lamblia to undergo two distinct differentiations in response to physiologic stimuli is central to its pathogenesis. The giardial cytoskeleton changes drastically during encystation and excystation. However, the signal transduction pathways mediating these transformations are poorly understood. We tested the hypothesis that PP2A, a highly conserved serine/threonine protein phosphatase, might be important in giardial differentiation. We found that in vegetatively growing trophozoites, gPP2A-C protein localizes to basal bodies/centrosomes, and to cytoskeletal structures unique to Giardia: the ventral disk, and the dense rods of the anterior, posterior-lateral, and caudal flagella. During encystation, gPP2A-C protein disappears from only the anterior flagellar dense rods. During excystation, gPP2A-C localizes to the cyst wall in excysting cysts but is not found in the wall of cysts with emerging excyzoites. Transcriptome and immunoblot analyses indicated that gPP2A-C mRNA and protein are upregulated in mature cysts and during the early stage of excystation that models passage through the host stomach. Stable expression of gPP2A-C antisense RNA did not affect vegetative growth, but strongly inhibited the formation of encystation secretory vesicles (ESV) and water-resistant cysts. Moreover, the few cysts that formed were highly defective in excystation.Thus, gPP2A-C localizes to universal cytoskeletal structures and to structures unique to Giardia. It is also important for encystation and excystation, crucial giardial transformations that entail entry into and exit from dormancy.

Nuclear Inheritance and Genetic Exchange without Meiosis in the Binucleate ParasiteGiardia intestinalis

“…However, because we cannot distinguish between old and new discs, it remains possible that the disc fragments in the cyst are derived from the parental disc and are retained as raw material for the construction of new discs during excystation. This is the first study, to our knowledge, to use immunofluorescence staining to examine the Giardia cytoskeleton in depth during excystation, although previous studies have looked at the localizaton of two proteins, PP2A and beta-giardin, in excyzoites (Palm et al, 2005;Lauwaet et al, 2007b). As reported previously (Coggins and Schaefer, 1984;Coggins and Schaefer, 1986;Buchel et al, 1987;Erlandsen et al, 1989), we observed that the excyzoite exits the cyst at one pole, starting with its flagella.…”

Glucosylceramide synthesis inhibition affects cell cycle progression, membrane trafficking, and stage differentiation in Giardia lamblia

“…55 ). Interestingly, putative giardial protein homologs of components of the ceramide signaling cascade, including protein phosphatases PP1, PI3K/AKT, and PP2A ( 56 ), are annotated in the Giardia genome database, and the latter was shown to be involved in the differentiation process of the parasite ( 57 ). Thus it is likely that signaling by ceramide, and possibly by other bioactive sphingolipids, is also highly conserved in this simple eukaryote.…”

Microvesicles released from Giardia intestinalis disturb host-pathogen response in vitro

“…Three encystation-specific cyst wall proteins (CWP1, 2 and 3) are expressed and concentrated in encystation-specific-vesicles (ESVs) that circulate within the parasite before being transported to the cyst wall (Luján et al, 1996;Reiner et al, 1990;Lauwaet et al, 2007;Faso and Hehl, 2011;Benchimol and De Souza, 2011). Synthesis of ESVs starts 4-6 h after encystation is induced and is completed with the cyst formation by approximately 24 h (Reiner et al, 1990).…”