Dataset Information

Endosymbiont elimination enhances de novo heme synthesis by the overexpression of a ferrous iron transporter in Strigomonas culicis

ABSTRACT: 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.

ORGANISM(S): Candidatus Kinetoplastidibacterium Blastocrithidiae (ex Strigomonas Culicis) Strigomonas Culicis

SUBMITTER: Rubem Menna-Barreto

PROVIDER: PXD044748 | panorama | Mon Jan 12 00:00:00 GMT 2026

REPOSITORIES: PanoramaPublic

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Publications

Heme metabolism in Strigomonas culicis: Implications of H2O2 resistance induction and symbiont elimination.

Bombaça Ana Cristina Souza ACS Caminha Marcelle Almeida MA Barbosa Juliana Magalhães Chaves JMC Pedra-Rezende Yasmin Y Ennes-Vidal Vitor V Brunoro Giselle Villa Flor GVF Archanjo Bráulio Soares BS d'Avila Claudia Masini CM Valente Richard Hemmi RH Menna-Barreto Rubem Figueiredo Sadok RFS

The Journal of biological chemistry 20240817 9

Monoxenous trypanosomatid Strigomonas culicis harbors an endosymbiotic bacterium, which enables the protozoa to survive without heme supplementation. The impact of H2O2 resistance and symbiont elimination on intracellular heme and Fe2+ availability was analyzed through a comparison of WT strain with both WT H2O2-resistant (WTR) and aposymbiotic (Apo) protozoa. The relative quantification of the heme biosynthetic pathway through label-free pa ...[more]

PMID: 39159809

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Project description:The two-component system (TCS) SrrAB responds to the oxidative stress in Staphylococcus epidermidis. To study its regulatory function in oxidative stress, a srrAB deletion mutant (∆srrAB) was constructed using S. epidermidis strain 1457 (SE1457) as the parent strain. Compared to SE1457, the viable cell counts of the ∆srrAB mutant significantly decreased in the post-stationary phase culture, coinciding with a sharp increase in ROS accumulation. The impaired growth of the ∆srrAB mutant was partially restored by shifting the culture from oxic to microaerobic conditions. Consistently, growth of the ∆srrAB mutant in TSB medium containing H2O2 was notably inhibited compared to parent strain SE1457, and the mutant showed significantly decreased resistance (100- to 1000-fold) to H2O2 and CHP in both oxic and microaerobic conditions, which was fully rescued by the addition of ROS inhibitor DIP. Furthermore, deletion of srrAB resulted in decreased intracellular survival in the Ana-1 macrophages due to intracellular ROS accumulation. Complementation of srrAB in the ∆srrAB mutant restored ROS resistance and intracellular survival to wild-type levels. RNA-seq analysis revealed that srrAB deletion affected the transcription levels of 610 genes, including those involved in oxidative stress, respiratory and energy metabolism, and transition ion homeostasis. These findings were corroborated by quantitative real-time reverse transcription-PCR (qRT-PCR). In the ∆srrAB mutant, expressions of ROS-scavenging genes katA, ahpC, scdA, serp1797 and serp0483 were downregulated compared to SE1457. Electrophoretic mobility shift assay further demonstrated that phosphorylated SrrA bound to the promoter regions of srrAB, katA, ahpC, scdA, serp1797 and serp0483 genes. This study elucidates that in S. epidermidis, SrrAB is the first TCS to sense and respond upon the oxidants, directly regulating transcription levels of the genes involved in ROS scavenging and ion homeostasis, thereby facilitating S. epidermidis detoxification of ROS and adaption to the commensal environment.

Dataset Information

Endosymbiont elimination enhances de novo heme synthesis by the overexpression of a ferrous iron transporter in Strigomonas culicis

Publications

Heme metabolism in Strigomonas culicis: Implications of H<sub>2</sub>O<sub>2</sub> resistance induction and symbiont elimination.

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