Treatment of Trypanosoma cruzi with 2-bromopalmitate alters morphology, endocytosis, differentiation and infectivity

Abstract

BackgroundThe palmitate analogue 2-bromopalmitate (2-BP) is a non-selective membrane tethered cysteine alkylator of many membrane-associated enzymes that in the last years emerged as a general inhibitor of protein S-palmitoylation. Palmitoylation is a post-translational protein modification that adds palmitic acid to a cysteine residue through a thioester linkage, promoting membrane localization, protein stability, regulation of enzymatic activity, and the epigenetic regulation of gene expression. Little is known on such important process in the pathogenic protozoan Trypanosoma cruzi, the etiological agent of Chagas disease.ResultsThe effect of 2-BP was analyzed on different developmental forms of Trypanosoma cruzi. The IC50/48 h value for culture epimastigotes was estimated as 130 μM. The IC50/24 h value for metacyclic trypomastigotes was 216 nM, while for intracellular amastigotes it was 242 μM and for cell derived trypomasigotes was 262 μM (IC50/24 h). Our data showed that 2-BP altered T. cruzi: 1) morphology, as assessed by bright field, scanning and transmission electron microscopy; 2) mitochondrial membrane potential, as shown by flow cytometry after incubation with rhodamine-123; 3) endocytosis, as seen after incubation with transferrin or albumin and analysis by flow cytometry/fluorescence microscopy; 4) in vitro metacyclogenesis; and 5) infectivity, as shown by host cell infection assays. On the other hand, lipid stress by incubation with palmitate did not alter epimastigote growth, metacyclic trypomastigotes viability or trypomastigote infectivity.ConclusionOur results indicate that 2-BP inhibits key cellular processes of T. cruzi that may be regulated by palmitoylation of vital proteins and suggest a metacyclic trypomastigote unique target dependency during the parasite development.Electronic supplementary materialThe online version of this article (10.1186/s12860-018-0170-3) contains supplementary material, which is available to authorized users.

Complementary crosstalk between palmitoylation and phosphorylation events in MTIP regulates its role during Plasmodium falciparum invasion

“…The phenotypic effect of crosstalk was further studied by measuring P. falciparum growth inhibition against individual and combinatorial treatments with ST092793 and 2-BMP which are generic inhibitors of phosphorylation and palmitoylation, respectively. Although 2-BMP is known to have non-specific effects ( DeJesus and Bizzozero, 2002 ; Zheng et al., 2015 ), it has been used extensively to target palmitoylation-based pathways in L. donovani , T. cruzi , and Toxoplasma ( Foe et al., 2015 ; Ayana et al., 2018 ; Batista et al., 2018b ; Batista et al., 2018a ) for reducing infectivity. Plausibly, one of our recent unpublished data has also shown that 2-BMP has no impact on overall morphology of host RBCs.…”

Eggs of the mosquito Aedes aegypti survive desiccation by rewiring their polyamine and lipid metabolism

“…A similar titrated inhibitor-based approach as described above was followed to inhibit fatty acid oxidation in fresh and desiccated Aedes eggs, and 0.8 mM of 2-bromopalmitic acid (Sigma Aldrich, 238422) was added to fresh blood used for feeding 10 days old female mosquitoes. This concentration was specifically chosen based on our optimised range for DFMO treatment which served as a reference and from published data by [ 77 ]. Higher doses of the drug (>1.2 mM) led to the mortality of adult females.…”

Synergistic effect and ultrastructural changes in Trypanosoma cruzi caused by isoobtusilactone A in short exposure of time

“…Vero cells (CCL-81) isolated from the kidney of the African green monkey Cercopithecus aethiops [ 17 ] were purchased from ATCC® (Washington DC, USA). The cells were inoculated in 200 μL of Dulbecco’s modified Eagle (DMEM) medium in 96-well plates at a concentration of 2 × 10 4 cells/well and incubated at 37°C for 24 h for adherence of the cells [ 18 ]. Six different concentrations of the compounds (12 to 900 μM for butanolides, and 200 to 1,600 μM for benznidazole) were added to the wells.…”