A New ABC Half-Transporter in Leishmania major Is Involved in Resistance to Antimony

Abstract

The characterization of ABCI4, a new intracellular ATP-binding cassette (ABC) half-transporter in Leishmania major, is described. We show that ABCI4 is involved in heavy metal export, thereby conferring resistance to Pentostam, to Sb(III), and to As(III) and Cd(II). Parasites overexpressing ABCI4 showed a lower mitochondrial toxic effect of antimony by decreasing reactive oxygen species production and maintained higher values of both the mitochondrial electrochemical potential and total ATP levels with respect to controls. The ABCI4 half-transporter forms homodimers as determined by a coimmunoprecipitation assay. A combination of subcellular localization studies under a confocal microscope and a surface biotinylation assay using parasites expressing green fluorescent protein-and FLAG-tagged ABCI4 suggests that the transporter presents a dual localization in both mitochondria and the plasma membrane. Parasites overexpressing ABCI4 present an increased replication in mouse peritoneal macrophages. We have determined that porphyrins are substrates for ABCI4. Consequently, the overexpression of ABCI4 confers resistance to some toxic porphyrins, such as zinc-protoporphyrin, due to the lower accumulation resulting from a significant efflux, as determined using the fluorescent zinc-mesoporphyrin, a validated heme analog. In addition, ABCI4 has a significant ability to efflux thiol after Sb(III) incubation, thus meaning that ABCI4 could be considered to be a potential thiol-X-pump that is able to recognize metal-conjugated thiols. In summary, we have shown that this new ABC transporter is involved in drug sensitivity to antimony and other compounds by efflux as conjugated thiol complexes.

The LABCG2 Transporter from the Protozoan Parasite Leishmania Is Involved in Antimony Resistance

“…One of the most characteristic mechanisms of antimony resistance in Leishmania is drug efflux mediated by ABC (ATP-binding cassette) transporters such as MRPA (formerly PGPA)/ABCC3 (3,4) or ABCI4 (5), which results in a reduced degree of antimony accumulation in parasites. ABC transporters are one of the largest protein families known; they are highly evolutionarily conserved from bacteria to humans and are involved in the transport of different compounds through biological membranes.…”

The Oral Antimalarial Drug Tafenoquine Shows Activity against Trypanosoma brucei

“…ROS production has also been described as a hallmark for cell death by necrosis (30). We used the ROS-sensitive probe H 2 DCF-DA, a probe not highly specific but extensively used in the parasites Leishmania (49,50) and T. brucei (51,52), in order to determine TFQ's potential as an inductor of ROS production in parasites. for 10 min with 1, 5, and 10 M TFQ.…”

“…Fixed cells were sedimented onto electron microscopy stubs previously covered with poly-L-lysine at 4°C over 24 h and in the presence of humidity. The samples were postfixed with 1% (wt/vol) osmium tetroxide (OsO 4 ) and then dehydrated using a graded series of ethanol solutions (50,70, and 90% and three times at 100%) over a period of 15 min at room temperature. The critical point was reached in a Leica EM CPD300 automated dryer, before the samples were fully dried in the presence of carbon using an EMITECH K975X Turbo Evaporator and then observed using an Auriga (FIB-FESEM) Carl Zeiss SMT scanning electron microscope.…”

Antiulcer Activity after Oral Administration of the Wormwood Ethanol Extract on Lesions due to <I>Leishmania major</I> Parasites in BALB/C Mice

“…CL treatment is often impeded by the development of drug resistance 11 , side effects like cardiac and renal toxicities, variable efficacy, necessity to endure long courses of parenteral administration, the absence of an effective vaccine, as well as the formidable task of developing a simple, cheap, sustainable and swift drug-evaluating system globally effective on the different Leishmania species/strains infecting humans; though this being too optimistic 20 . Attempts to identify alternative search strategies are thus conspicuously essential 25 . About 80% of the world's inhabitants depend on folk medicines for their healthcare 9 .…”