A plasma membrane localized protein phosphatase in Toxoplasma gondii, PPM5C, regulates attachment to host cells

Chunlin Yang,Emily Sampson,Gustavo Arrizabalaga,William J Blakely,Caia Dominicus,Malgorzata Broncel,Moritz Treeck

doi:10.1038/s41598-019-42441-1

Chunlin Yang, Emily Sampson + Show 5 more

Open Access

https://doi.org/10.1038/s41598-019-42441-1

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Abstract

The propagation of Toxoplasma gondii is accomplished by repeated lytic cycles of parasite attachment to a host cell, invasion, replication within a parasitophorous vacuole, and egress from the cell. This lytic cycle is delicately regulated by calcium-dependent reversible phosphorylation of the molecular machinery that drives invasion and egress. While much progress has been made elucidating the protein kinases and substrates central to parasite propagation, little is known about the relevant protein phosphatases. In this study, we focused on the five protein phosphatases that are predicted to be membrane-associated either integrally or peripherally. We have determined that of these only PPM5C, a PP2C family member, localizes to the plasma membrane of Toxoplasma. Disruption of PPM5C results in a slow propagation phenotype in tissue culture. Interestingly, parasites lacking PPM5C divide and undergo egress at a normal rate, but have a deficiency in attaching to host cells. Both membrane localization and phosphatase activity are required for PPM5C’s role in attachment. Phosphoproteomic analysis show relatively few phosphorylation sites being affected by PPM5C deletion in extracellular parasites of which several are found on proteins involved in signaling cascades. This implies that PPM5C is part of a wider regulatory network important for attachment to host cells.

Highlights

Toxoplasma gondii, an obligate intracellular parasite of the phylum Apicomplexa, infects warm-blooded animals including an estimated two billion people[1,2]
Immunofluorescence assay (IFA) of the resulting strains showed that PPM2A and PPM2B are both distributed throughout the cytosol (Fig. 2A)
We detected PPM5C in the residual body, a structure that arises during parasite division and www.nature.com/scientificreports tagged protein phosphatases were stained with antibodies against the HA epitope and the surface protein

Summary

Introduction

Toxoplasma gondii, an obligate intracellular parasite of the phylum Apicomplexa, infects warm-blooded animals including an estimated two billion people[1,2]. The symptoms of severe toxoplasmosis are in great part the result of the tissue damage caused by repeated cycles of parasite entry into cells, replication, and lytic egress These steps, along with attachment and motility, are exquisitely regulated by coordinated secretion from three unique organelles, micronemes, rhoptries, and dense granules[10]. Calcium dependent reversible phosphorylation has been shown to play a critical role in regulating microneme secretion and activation of the motility system during egress and invasion[12,13,14]. Since most phosphorylated sites detected in lytic cycle effector proteins are primarily serine and threonine residues, phosphatases involved in their dephosphorylation are more likely to be PSPs16 Among these PSPs, only calcineurin, a phosphoprotein phosphatase (PPP) family PSP, has been shown to be involved in the regulation of the lytic cycle of Toxoplasma[21]. Calcineurin plays a role in microneme independent host cell attachment[21]

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