Dataset Information

Glycolytic profile shift and antioxidant triggering in symbiont-free and H2O2-resistant Strigomonas culicis

ABSTRACT: During their life cycle, trypanosomatids are exposed to stress conditions and adapt their energy and antioxidant metabolism to colonize their hosts. Strigomonas culicis is a monoxenous protist found in invertebrates with an endosymbiotic bacterium that completes essential biosynthetic pathways for the trypanosomatid. Our research group previously generated a wild-type H2O2-resistant (WTR) strain that showed improved mitochondrial metabolism and antioxidant defenses, which led to higher rates of Aedes aegypti infection. Here, we assess the biological contribution of the S. culicis endosymbiont and reactive oxygen species (ROS) resistance to oxidative and energy metabolism processes. Using high-throughput proteomics, several proteins involved in glycolysis and gluconeogenesis, the pentose phosphate pathway and glutathione metabolism were identified. The results suggest that ROS resistance decreases glucose consumption and indicate that the metabolic products from gluconeogenesis are key to supplying the protist with high-energy and reducing intermediates. Our hypothesis was confirmed by biochemical assays showing opposite profiles for glucose uptake and hexokinase and pyruvate kinase activity levels in the WTR and aposymbiotic strains, while the enzyme glucose-6P 1-dehydrogenase was more active in both strains. Regarding the antioxidant system, ascorbate peroxidase has an important role in H2O2 resistance and may be responsible for the high infection rates previously described for A. aegypti. In conclusion, our data indicate that the energy-related and antioxidant metabolic processes of S. culicis are modulated in response to oxidative stress conditions, providing new perspectives on the biology of the trypanosomatid-insect interaction as well as on the possible impact of resistant parasites in accidental human infection.

INSTRUMENT(S):

ORGANISM(S): Strigomonas Culicis

TISSUE(S): Cell In Vitro, Cell Culture

DISEASE(S): Trypanosomiasis

SUBMITTER: Richard Hemmi Valente

LAB HEAD: Richard Hemmi Valente

PROVIDER: PXD015717 | Pride | 2019-11-27

REPOSITORIES: Pride

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			Action	DRS
	20171012_C01_02ug_01.RAW	Raw
	20171012_C01_02ug_01.sqt	Other
	20171012_C01_02ug_02.RAW	Raw
	20171012_C01_02ug_02.sqt	Other
	20171012_C01_02ug_03.RAW	Raw

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Publications

Glycolytic profile shift and antioxidant triggering in symbiont-free and H2O2-resistant Strigomonas culicis.

Bombaça Ana Cristina Souza ACS Brunoro Giselle Villa Flor GVF Dias-Lopes Geovane G Ennes-Vidal Vitor V Carvalho Paulo Costa PC Perales Jonas J d'Avila-Levy Claudia Masini CM Valente Richard Hemmi RH Menna-Barreto Rubem Figueiredo Sadok RFS

Free radical biology & medicine 20191121

During their life cycle, trypanosomatids are exposed to stress conditions and adapt their energy and antioxidant metabolism to colonize their hosts. Strigomonas culicis is a monoxenous protist found in invertebrates with an endosymbiotic bacterium that completes essential biosynthetic pathways for the trypanosomatid. Our research group previously generated a wild-type H2O2-resistant (WTR) strain that showed improved mitochondrial metabolism and antioxidant defenses, which l ...[more]

PMID: 31760093

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Project description:The monoxenous trypanosomatid Strigomonas culicis inhabits several mosquito species, including Aedes aegypti. Previously, our group described that the infection of A. aegypti females by this trypanosomatid triggers the insect's immune response, reduces its reproductive fitness, and could potentially influence the vector competence. S. culicis maintains a mutualistic relationship with an endosymbiotic bacterium, which enables the protozoa to survive without heme supplementation. Here, the impact of H2O2 resistance and symbiont elimination on intracellular heme and Fe2+ availability was analyzed, comparing both the wild-type H2O2-resistant (WTR) and aposymbiotic (Apo) with the wild-type (WT) strain. Relative quantification through label-free PRM targeted mass spectrometry approach pointed out that H2O2 resistance induction does not influence the abundance of heme biosynthetic pathway; however, symbiont elimination increases the abundance of all components of de novo heme synthesis, except for protoporphyrinogen oxidase (PPOX). Confirming the limited role of PPOX in this biossynthetic pathway, the addition of protoporphyrin IX (PPIX) to heme-decreased medium (HDM) leads to higher growth rates and enhances intracellular heme content in Apo strain. A putative ferrous iron transporter homologous to LIT1 and TcIT from Leishmania major and Trypanosoma cruzi, was also detected for the first time. Targeted mass spectrometry approach indicated that H2O2 resistance and symbiont elimination increased S. culicis iron transporter (ScIT) abundance by 1.6- and 16.4-fold in comparison to WT. Accordingly, antibody-mediated blockage of ScIT decreased 26.7 and 41.9% intracellular Fe2+ concentration in both WTR and Apo strains, whereas no effect was detected in WT. Gene expression analysis showed that Apo growth in HDM increases 1.8-fold ScIT transcript levels, comparing to protozoa maintained in standard medium. Also, addition of 10 µM hemin to HDM significantly decreased intracellular Fe2+ concentration and ferric iron reduction in Apo strain. Together, our data indicate molecular mechanisms of S. culicis to deal with symbiont elimination and reduced heme availability. Since heme and Fe2+ are essential cofactors in many metabolic pathways, such as oxidative phosphorylation and antioxidant system, this study also provides novel mechanistic insights associated with S. culicis H2O2 resistance.

Project description:Salmonella causes inflammation in infected hosts. Inflammation is a well-characterized defensive mechanism of innate immunity. The recognition and engagement of lipopolysaccharide (LPS) endotoxins in the outer membranes of Salmonella to Toll-like receptor 4 of immune cells (macrophages and dendritic cells) trigger inflammatory responses characterized by secretion of pro-inflammatory cytokines, including TNF-beta, IL-1 and IL-6. These cytokines cause fever, anorexia, bodyweight losses, and catabolism of skeletal muscles and adipose tissues. However, molecular events underlying innate immune responses and metabolic activities during the later stage of inflammation are poorly understood. Additionally, the effects of prebiotics and antibiotics on innate immunity and nutrient metabolism are not yet reported. The objective of this study is to investigate the effects of a mannanoligosaccharide (MOS) prebiotic and virginiamycin (VIRG) sub-therapeutic antibiotic on innate immunity and glucose metabolism during late inflammation. We induced Salmonella LPS-systemic inflammation in a chicken model. Differentially regulated gene expressions were measured using 2 colour focussed oligonucleotide chicken-specific microarrays. Microarray analysis was performed on liver, intestinal and skeletal muscle tissues. We found that late inflammation was principally modulated by interleukin 3 (IL 3) and that glucose was mobilized from gluconeogenesis occurring in the intestines only. MOS and VIRG modulated innate immunity and metabolic genes differently. In contrast to VIRG, MOS terminated inflammatory responses earlier. Our results indicate IL 3 gene up-regulation in VIRG-fed chickens. To meet the higher energy requirements of VIRG chickens, genes for intestinal gluconeogenesis and liver glycolysis were respectively induced. Our study reveals the potential mechanisms by which prebiotic and antibiotic modulated innate immunity and glucose metabolism during late inflammation. 14-day old chickens were injected i.p. with saline or LPS. For each tissue and experimental conditions (saline or LPS challenge), a total of 12 microarrays (6 MOS birds + 6 VIRG birds) were used in a 2 x 2 factorial design and complete interwoven loop arrangement. We compared gene expression from prebiotic-fed birds with antibiotic-fed birds without including reference RNA. LPS challenge, antibiotic or prebiotic, innate immunity, glucose metabolism

Dataset Information

Glycolytic profile shift and antioxidant triggering in symbiont-free and H2O2-resistant Strigomonas culicis

Dataset's files

Publications

Glycolytic profile shift and antioxidant triggering in symbiont-free and H<sub>2</sub>O<sub>2</sub>-resistant Strigomonas culicis.

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