BIOL8291: PfATP4, Na+ Regulation, and Antimalarials Presentation

This presentation focuses on the research investigating sodium ion (Na+) regulation within the malaria parasite Plasmodium falciparum, specifically examining the role of the cation ATPase PfATP4. The study highlights how P. falciparum alters the permeability of the host erythrocyte membrane, leading to increased Na+ influx. PfATP4, a plasma membrane protein, actively extrudes Na+ to maintain low intracellular concentrations. The presentation discusses how mutations in PfATP4 confer resistance to spiroindolone antimalarials, which target Na+ homeostasis. Key findings include the identification of PfATP4 as a crucial factor in maintaining Na+ balance and the implications of its mutations on drug resistance. Future research directions involve developing new antimalarial drugs that target Na+ influx and efflux mechanisms in mutated P. falciparum strains. The presentation also touches on the unknown mechanisms underlying Na+ increases in response to K+ removal, suggesting further areas of exploration. The document is available on Desklib, a platform offering study tools and solved assignments for students.

Na+ regulation in the malaria parasite Plasmodium falciparum involves the
cation ATPase PfATP4 and is a target of the spiroindolone antimalarials
Introduction
Research Questions
Methods Results
References
Discussion
Spillman, N.J., Allen, R.J., McNamara, C.W., Yeung, B.K., Winzeler, E.A., Diagana, T.T. and
Kirk, K., 2013. Cell host & microbe, 13(2), pp.227-237.
Plamodium falciparun, formulates novel pathways
permeability in the host erythrocyte plasma membrane,
resulting in alteration in nutrient absorption by the cell
infected. Such pathways are also associated with an
increased intake of sodium (Na +) ions resulting in
excessively high levels of the mineral (Dangi 2016).
The following study examines the extrusion of Na + by
the parasite, with the aid of PfATP4 – a plasma
membrane protein of the parasite similar to NA + -
ATPases. The presence of mutations in PfATP4 confers
resistant to antimalarial spiroindolones – medications
aiming to hinder Na+ homoeostasis in malarial parasites
(Spillman et al. 2013).
Figure 1: Schematic Representation
Showing the Proposed Role of PfATP4 in
Na+ Homeostasis in the Intraerythrocytic
P. falciparum Trophozoite-Stage Parasite
A: It is estimated that PtATP4 functions as an ENA
Na+ -ATPase associated with eactive extrusion of
sodium ions from the intracellular matrix of the
malarial parasite, against the influx of Na+, hence
maintaining 10 times lower concentration.
B: Inhibition of PtATP4 by spiroindolones resulting in
increased influx of Na+ (Spillman et al. 2013).
Aim: To investigate Na+ regulation in Plamodium
falciparun(1).
Hypothesis: PfATP4 is an Na + efflux pump of the
plasma membrane, which is similar to Na + -ATPases
and is a major target of the antimalarial medications
(1).
For the purpose of investigation of the processes
underlying regulation of Na+ in the malarial parasite,
mature strains of P. falciparum 3D7 trophozoites were
isolated functionally, from host cells using saponin
permeabilization of the cell membrane of the host along
with incorporation of SBFL, a fluorescent dye sensitive to
Na+. For the purpose of investigation of the processes
underlying maintenance of low concentrations of Na + in
the malarial parasite, a number of ion transport inhibitors
and ionophones were evaluation for their reactions with
Na+. Suspending the parasites, which had been isolated, in
medium free from glucose (environments associated with
depletion of ATP in parasites) led to an increase in Na +
gradually (1).
Figure 2. Na+-Dependence and Spiroindolone-Sensitivity of
Membrane-Associated ATPase Activity in P. falciparum
Infected
Black: The membrane preparations were kept in a solution high
in Na+ (100mM, indicated by black bars) White: Solution low in
Na+ (0.5 mM. as indicated by white bars, where equimolar
choline was used in place of Na +), in the presence, and absence
of 50 nM NITD2 46 (Spillman et al. 2013).
Figure 3. Effects of an Ionophore and Ion Transport
Inhibitors on
[Na+]i in Saponin-Isolated, SBFI-loaded P. falciparum
Trophozoites. A to E: Trace of [Na]1 indicating the
consequences of addition (as displayed at the point
symbolized by the closed triangle) of A) gramicidin (5 mM),
(B) EIPA (20 mM), (C) ouabain (2 mM), (D) furosemide (100
mM), and (E) orthovanadate (100 mMr (1).
•Sequence analysis revealed that PfATP4 plays a key
role in the maintenance of low intracellular and high
extracellular Na+ concentrations and the extrusion of
Na+ ions within the malarial parasite P. falciparum.
•PfATP4 being located in the cell membrane of the
malarial parasite, justifies its role in the extrusion of
Na+ and the study findings that PfATP4 mutations is
associated with alterations in Na+ concentrations.
•This results in high resting Na +, reductions in Na +
efflux in response to high load of intracellular Na+ and
increments towards loss of parasite resistance to the
hindrances in growth caused by high levels of Na + in
the extracellular matrix (Spillman et al. 2013).
Dangi, P., 2016. A novel spiroindoline
kills human malaria parasites via
modulation of Na ion influx mediated
autophagy and apoptosis.
International Journal of Infectious
Diseases, 45, p.357.
Spillman, N.J., Allen, R.J.,
McNamara, C.W., Yeung, B.K.,
Winzeler, E.A., Diagana, T.T. and
Kirk, K., 2013. Na+ regulation in the
malaria parasite Plasmodium
falciparum involves the cation ATPase
PfATP4 and is a target of the
spiroindolone antimalarials. Cell host
& microbe, 13(2), pp.227-237.
Veiga, M.I., Dhingra, S.K., Henrich,
P.P., Straimer, J., Gnädig, N.,
Uhlemann, A.C., Martin, R.E.,
Lehane, A.M. and Fidock, D.A., 2016.
Globally prevalent PfMDR1
mutations modulate Plasmodium
falciparum susceptibility to
artemisinin-based combination
therapies. Nature communications, 7,
p.11553.
• The concentration of resting Na+, in a 125mM Na +
suspension, was found to be 11.0 ± 0.6 mM.
• After replacing extracellular Na + with alternative cations
the concentration of Na+ reduced rapidly within a few
minutes. Alternatively, after increasing the concentration
of [Na+]o, there was a slight increase in [Na+]1.
• Increments in [Na+]1 was observed to be less than 20 mM,
after administration of increments in [Na +]o, twice normal
physiological values from 175 mM to 300 mM.
• It was found that the selective monovalent-cation
ionophore, Gramicidin (5μM) resulted in a significant
increase in Na+ with the approaching 125 mM Na +
concentration.
• The phosphate analog, sodium orthovanadate, which
inhibits P-type ATPases, was observed to exert a
prolonged time-dependent increase in Na+ (1).
Future Aspects
•From the findings, it can hence be implied that
mutations result in a resistance of the parasite to
spiroindolones.
•Such findings confirm the hypothesis that the target
for such antimalarial medications is PfATP4.
•Hence, considering such the same, this researched
paves the way for indicating the need to develop and
administer novel antimalarial medications which
target the mechanisms underlying high rates of influx
and efflux of Na+ ions with mutated P. falciparum
strains with mutations in PfATP4 (Spillman et al.
2013).
•The procedures underlying increase in Na+ in
response to K + removal is not known (Veiga et al.
2016)..
•Hence, this implicates the need to conduct future
research underlying extensive exploration of the same.