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PLoS Biol. 2019 Jun 10;17(6):e3000308. doi: 10.1371/journal.pbio.3000308. eCollection 2019 Jun.

Phosphorylation of the VAR2CSA extracellular region is associated with enhanced adhesive properties to the placental receptor CSA.

Author information

1
Université de Paris, Biologie Intégrée du Globule Rouge, UMR_S1134, BIGR, INSERM, Paris, France.
2
Institut National de la Transfusion Sanguine, Paris, France.
3
Unité de Biologie des Interactions Hôte-Parasite, Institut Pasteur, Paris, France.
4
INSERM U1201, Paris, France.
5
CNRS ERL9195, Paris, France.
6
Proteomics Core Facility, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.
7
CNRS5290, IRD224, University Montpellier, MIVEGEC, Montpellier, France.
8
Centre for Chronic Infectious and Inflammation Disease, Biomedical Sciences Cluster, School of Health and Biomedical Sciences, RMIT University, Bundoora, Victoria, Australia.

Abstract

Plasmodium falciparum is the main cause of disease and death from malaria. P. falciparum virulence resides in the ability of infected erythrocytes (IEs) to sequester in various tissues through the interaction between members of the polymorphic P. falciparum erythrocyte membrane protein 1 (PfEMP1) adhesin family to various host receptors. Here, we investigated the effect of phosphorylation of variant surface antigen 2-CSA (VAR2CSA), a member of the PfEMP1 family associated to placental sequestration, on its capacity to adhere to chondroitin sulfate A (CSA) present on the placental syncytium. We showed that phosphatase treatment of IEs impairs cytoadhesion to CSA. MS analysis of recombinant VAR2CSA phosphosites prior to and after phosphatase treatment, as well as of native VAR2CSA expressed on IEs, identified critical phosphoresidues associated with CSA binding. Site-directed mutagenesis on recombinant VAR2CSA of 3 phosphoresidues localised within the CSA-binding region confirmed in vitro their functional importance. Furthermore, using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein-9 nuclease (CRISPR/Cas9), we generated a parasite line in which the phosphoresidue T934 is changed to alanine and showed that this mutation strongly impairs IEs cytoadhesion to CSA. Taken together, these results demonstrate that phosphorylation of the extracellular region of VAR2CSA plays a major role in IEs cytoadhesion to CSA and provide new molecular insights for strategies aiming to reduce the morbidity and mortality of PM.

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