ABSTRACT
Malaria parasite infection in anopheline mosquitoes is limited by inflammatory levels of nitric oxide metabolites. To assess the mechanisms of parasite stasis or toxicity, we investigated the biochemistry of these metabolites within the blood-filled mosquito midgut. Our data indicate that nitrates, but not nitrites, are elevated in the Plasmodium-infected midgut. Although levels of S-nitrosothiols do not change with infection, blood proteins are S-nitrosylated after ingestion by the mosquito. In addition, photolyzable nitric oxide, which can be attributed to metal nitrosyls, is elevated after infection and, based on the abundance of hemoglobin, likely includes heme iron nitrosyl. The persistence of oxyhemoglobin throughout blood digestion and changes in hemoglobin conformation in response to infection suggest that hemoglobin catalyzes the synthesis of nitric oxide metabolites in a reducing environment. Provision of urate, a potent reductant and scavenger of oxidants and nitrating agents, as a dietary supplement to mosquitoes increased parasite infection levels relative to allantoin-fed controls, suggesting that nitrosative and/or oxidative stresses negatively impact developing parasites. Collectively, our results reveal a unique role for nitric oxide in an oxyhemoglobin-rich environment. In contrast to facilitating oxygen delivery by hemoglobin in the mammalian vasculature, nitric oxide synthesis in the blood-filled mosquito midgut drives the formation of toxic metabolites that limit parasite development.
Subject(s)
Anopheles/metabolism , Anopheles/parasitology , Malaria/prevention & control , Nitric Oxide/metabolism , Oxidative Stress/drug effects , Plasmodium berghei/metabolism , Allantoin/pharmacology , Animals , Dermatologic Agents/pharmacology , Dietary Supplements , Hemoglobins/chemistry , Hemoglobins/metabolism , Malaria/metabolism , Mice , NG-Nitroarginine Methyl Ester/administration & dosage , Nitrites/toxicity , Nitrogen Oxides/toxicity , Oxygen/metabolism , Reactive Nitrogen Species/toxicity , Uric Acid/pharmacologyABSTRACT
Malaria parasite infection in anopheline mosquitoes induces nitrosative and oxidative stresses that limit parasite development, but also damage mosquito tissues in proximity to the response. Based on these observations, we proposed that cellular defenses in the mosquito may be induced to minimize self-damage. Specifically, we hypothesized that peroxiredoxins (Prxs), enzymes known to detoxify reactive oxygen species (ROS) and reactive nitrogen oxide species (RNOS), protect mosquito cells. We identified an Anopheles stephensi 2-Cys Prx ortholog of Drosophila melanogaster Prx-4783, which protects fly cells against oxidative stresses. To assess function, AsPrx-4783 was overexpressed in D. melanogaster S2 and in A. stephensi (MSQ43) cells and silenced in MSQ43 cells with RNA interference before treatment with various ROS and RNOS. Our data revealed that AsPrx-4783 and DmPrx-4783 differ in host cell protection and that AsPrx-4783 protects A. stephensi cells against stresses that are relevant to malaria parasite infection in vivo, namely nitric oxide (NO), hydrogen peroxide, nitroxyl, and peroxynitrite. Further, AsPrx-4783 expression is induced in the mosquito midgut by parasite infection at times associated with peak nitrosative and oxidative stresses. Hence, whereas the NO-mediated defense response is toxic to both host and parasite, AsPrx-4783 may shift the balance in favor of the mosquito.
Subject(s)
Insect Proteins/biosynthesis , Oxidative Stress/drug effects , Peroxidases/biosynthesis , Reactive Nitrogen Species/toxicity , Amino Acid Sequence , Animals , Anopheles/chemistry , Cells, Cultured , Drosophila melanogaster , Hydrazines/toxicity , Hydrogen Peroxide/toxicity , Insect Proteins/physiology , Malaria/physiopathology , Malaria/prevention & control , Molecular Sequence Data , Nitrites/toxicity , Nitrogen Oxides/toxicity , Peroxidases/physiology , Peroxiredoxins , Peroxynitrous Acid/toxicity , Plasmodium berghei , RNA Interference , Sequence Alignment , TransfectionABSTRACT
During the process of bloodfeeding by Anopheles stephensi, mammalian latent transforming growth factor beta1 (TGF-beta1) is ingested and activated rapidly in the mosquito midgut. Activation may involve heme and nitric oxide (NO), agents released in the midgut during blood digestion and catalysis of L-arginine oxidation by A. stephensi NO synthase (AsNOS). Active TGF-beta1 persists in the mosquito midgut to extended times postingestion and is recognized by mosquito cells as a cytokine. In a manner analogous to the regulation of vertebrate inducible NO synthase and malaria parasite (Plasmodium) infection in mammals by TGF-beta1, TGF-beta1 regulates AsNOS expression and Plasmodium development in A. stephensi. Together, these observations indicate that, through conserved immunological cross talk, mammalian and mosquito immune systems interface with each other to influence the cycle of Plasmodium development.
