PMID:30845261

Citation	Messina, V, Loizzo, S, Travaglione, S, Bertuccini, L, Condello, M, Superti, F, Guidotti, M, Alano, P, Silvestrini, F and Fiorentini, C (2019) The bacterial protein CNF1 as a new strategy against Plasmodium falciparum cytoadherence. PLoS ONE 14:e0213529
Abstract	Plasmodium falciparum severe malaria causes more than 400,000 deaths every year. One feature of P. falciparum-parasitized erythrocytes (pRBC) leading to cerebral malaria (CM), the most dangerous form of severe malaria, is cytoadherence to endothelium and blockage of the brain microvasculature. Preventing ligand-receptor interactions involved in this process could inhibit pRBC sequestration and insurgence of severe disease whilst reversing existing cytoadherence could be a saving life adjunct therapy. Increasing evidence indicate the endothelial Rho signaling as a crucial player in malaria parasite cytoadherence. Therefore, we have used the cytotoxic necrotizing factor 1 (CNF1), an Escherichia coli protein able to modulate the activity of Cdc42, Rac, and Rho, three subfamilies of the Rho GTPases family, to study interactions between infected erythrocytes and cerebral endothelium in co-culture models. The main results are that CNF1 not only prevents cytoadherence but, more importantly, induces the detachment of pRBCs from endothelia monolayers. We first observed that CNF1 does affect neither parasite growth, nor the morphology and concentration of knobs that characterize the parasitized erythrocyte surface, as viewed by scanning electron microscopy. On the other hand, flow cytometry experiments show that cytoadherence reversion induced by CNF1 occurs in parallel with a decreased ICAM-1 receptor expression on the cell surface, suggesting the involvement of a toxin-promoted endocytic activity in such a response. Furthermore, since the endothelial barrier functionality is compromised by P. falciparum, we conducted a permeability assay on endothelial cells, revealing the CNF1 capacity to restore the brain endothelial barrier integrity. Then, using pull-down assays and inhibitory studies, we demonstrated, for the first time, that CNF1 is able not only to prevent but also to cause the parasite detachment by simultaneously activating Rho, Rac and Cdc42 in endothelial cells. All in all our findings indicate that CNF1 may represent a potential novel therapeutic strategy for preventing neurological complications of CM.
Links	PubMed Online version:10.1371/journal.pone.0213529
Keywords
edit table

Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
ECOLX:A0A1W5T566		GO:2000251: positive regulation of actin cytoskeleton reorganization	ECO:0006062: wide-field fluorescence microscopy evidence used in manual assertion		P		In Figure 4 they use florescence microscopy to confirm the CNF1 protein in E.coli induces changes in the organization of the actin cytoskeleton in stress fibers and membrane protrusions, which prevents cytoadherence. This is shown through the addition of CNF1 protein and other inhibitors. With the addition of CNF1 compared to the control you can see the rearrangement of the cytoskeleton but, in the presence of inhibitors along with CNF1 the modifications of rearrangement are decreased.	complete CACAO 13556
ECOLX:A0A1W5T566		GO:0020035: cytoadherence to microvasculature, mediated by symbiont protein	ECO:0001143: adhesion assay evidence used in manual assertion		P		In Figure 1, adhesion assays were used to determine that CNF1 decreased pRBC cytoadherence in both cell line models by at least 40 percent. This is proven by comparing the pre- and post-treatment of the CNF1 protein with the TNF-alpha activated control. In figure 1A, CNF1 is used as pre-treatment to prevent the attachment of pRBC. In figure 1B, CNF1 is used as post-treatment to reverse adherence of pRBCs to endothelial cells.	complete CACAO 13658
ECOLX:A0A1W5T566		GO:0034446: substrate adhesion-dependent cell spreading	ECO:0007046: scanning electron microscopy evidence used in manual assertion		P		Figure 3A is a scanning electron micrograph that shows the response of brain cells to treatment with CNF1 protein, a mutant version of the protein (CNF1 C866S), or a control (TNF-alpha). Cells treated with CNF1 had increased monolayer confluence, as CNF1 promoted cell spreading and flattening. Cells treated with CNF1 C866S or TNF-alpha had an unchanged surface morphology in comparison.	complete CACAO 13721

References

See Help:References for how to manage references in GONUTS.